Tesi: "Pitx2" – Grafiati

1

Quentien, Marie-Hélène. "Différenciation des cellules du lignage antéhypophysaire somatolactotrope : un r^ole pour les facteurs de transcription Pitx1 et Pitx2". Aix-Marseille 2, 2002. http://www.theses.fr/2002AIX22080.

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2

Kulak, Stephen. "Mutational analysis of the homeobox transcription factor PITX2". Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1999. http://www.collectionscanada.ca/obj/s4/f2/dsk1/tape9/PQDD_0021/MQ47053.pdf.

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3

Doerdelmann, Thomas. "Structural and Biophysical Studies of the Pitx2 Homeodomain". University of Cincinnati / OhioLINK, 2011. http://rave.ohiolink.edu/etdc/view?acc_num=ucin1307443112.

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4

Fung, Khe Cheong Frederic, e 馮啟昌. "Upregulation of PITX2 transcription factor is associated with ovarian tumorigenesis". Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2011. http://hub.hku.hk/bib/B45988183.

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5

Kieusseian, Aurélie. "Etude du rôle du facteur de transcription Pitx2 dans la régulation de l'hématopoïèse". Paris 7, 2005. http://www.theses.fr/2005PA077147.

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6

Yu, Wenjie. "A Pitx2-Irx1 regulatory network controls dental epithelial stem cell differentiation during tooth development". Diss., University of Iowa, 2017. https://ir.uiowa.edu/etd/6020.

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Tooth development is precisely controlled by epithelium-mesenchyme interactions, coordinated signaling pathways and associated transcription factors. Although the processes involved in tooth development are well established, details of the cellular and molecular mechanisms that control tooth development are not fully understood. One of the primary unknown mechanisms is the regulation of dental epithelial stem cells (DESCs), including DESC specification, proliferation and differentiation. In this dissertation, I have addressed this gap in knowledge by studying the role of Pituitary homeobox 2 (Pitx2) and Iroquois 1 (Irx1) in teeth at the cellular and molecular level in mice. PITX2 contains mutations of which are associated for Axenfeld-Rieger syndrome (ARS) in humans and is also required for early tooth development. All the background knowledge is included in Chapter I. In Chapter II, I describe the conditional ablation of Pitx2 in the dental epithelium using a Krt14Cre driver line (Pitx2cKO mice). Knocking out Pitx2 in teeth led to delayed epithelial invagination at bud stage and disruption of tooth morphogenesis at cap stage. At the cellular level, Pitx2 mediates DESC differentiation, daughter cell proliferation in bud stage tooth and regulates enamel knot formation in cap stage tooth. At the molecular level, Pitx2 acts as an upstream regulator of the sonic hedgehog (Shh) signaling pathway by regulating the expression of Shh in the dental epithelial signaling center during early tooth development. In addition, I demonstrated that Pitx2 directly controls the transcription of Irx1. In Chapter III, I determined the cellular and molecular mechanisms of Irx1 in mice. Irx1 general knockout mice were generated by replacing the entire Irx1 gene body with a LacZ reporter gene. Irx1 null mice are neonatal lethal and this lethality is due to pulmonary immaturity with defective surfactant protein secretion. In teeth, Irx1 is expressed in the outer enamel epithelium (OEE) and stratum reticulum (SR) and mediates DESC to OEE and SR differentiation through regulation of Forkhead box protein J1 (Foxj1) and Sex determining region Y-box9 (Sox9). In summary, I identified a Pitx2-Irx1 regulatory network that controls DESC differentiation in teeth, which provided the field with a better understanding of tooth development and tooth regeneration.

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7

Kendall, Jed. "Targeting ß-catenin in MPNSTs". University of Cincinnati / OhioLINK, 2017. http://rave.ohiolink.edu/etdc/view?acc_num=ucin149155952770148.

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8

Schmidt, Jennifer Verfasser], Wolfgang Arthur [Akademischer Betreuer] [Schulz e Vlada B. [Akademischer Betreuer] Urlacher. "Funktionelle Charakterisierung von PITX2 im Prostatakarzinom / Jennifer Schmidt. Gutachter: Wolfgang A. Schulz ; Vlada B. Urlacher". Düsseldorf : Universitäts- und Landesbibliothek der Heinrich-Heine-Universität Düsseldorf, 2015. http://d-nb.info/1065375433/34.

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9

Schmidt, Jennifer [Verfasser], Wolfgang Arthur [Akademischer Betreuer] Schulz e Vlada B. [Akademischer Betreuer] Urlacher. "Funktionelle Charakterisierung von PITX2 im Prostatakarzinom / Jennifer Schmidt. Gutachter: Wolfgang A. Schulz ; Vlada B. Urlacher". Düsseldorf : Universitäts- und Landesbibliothek der Heinrich-Heine-Universität Düsseldorf, 2015. http://d-nb.info/1065375433/34.

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10

Li, Xiao. "The molecular mechanisms of PITX2 in tooth development and enamel defects in Axenfeld-Rieger Syndrome". Diss., University of Iowa, 2013. https://ir.uiowa.edu/etd/5014.

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Patients with Axenfeld-Rieger Syndrome (ARS) present various dental abnormalities. ARS is genetically associated with mutations in the PITX2 gene, which encodes one of the earliest transcription factors to initiate tooth development. Thus, Pitx2 has long been considered as an upstream regulator of the transcriptional hierarchy in tooth development. However, it is unclear how its mutant forms cause ARS dental anomalies. In this report, we outline the transcriptional mechanism that is defective in ARS. We demonstrate that during normal tooth development Pitx2 activates Amelogenin (Amel) expression, whose product is required for enamel formation, and that this regulation is perturbed by missense PITX2 mutations found in ARS patients. We further show that Pitx2-mediated Amel activation is enhanced and controlled by co-factors and target genes of Pitx2. These co-factors include cooperative transcription factors such as Dlx2 and FoxJ1; chromatin-associated remodeler factor Hmgn2; and Wnt signaling components such as Lef-1, β-catenin and Dact2. We also unveil a novel Pitx2 target gene Irx1 that functions in dental epithelium differentiation. Consistent with a physiological significance to these modulations, we show that FoxJ1, Dact2, Irx1 knockout mice and K14-Hmgn2 transgenic mice display various types of amelogenesis defects including enamel hypoplasia - consistent with the human ARS phenotype. Collectively, these findings define transcriptional mechanisms and multi-level regulations involved in normal tooth development and shed light on the molecular underpinnings of the enamel defect observed in ARS patients who carry PITX2 mutations. Moreover, our findings validate the etiology of the enamel defect in novel mouse models of enamel hypoplasia. The impact of this study on current understanding of the dental epithelium development and the translational value lie in the gene network we identified. By manipulating components of the network, pluripotent dental cells can be reprogrammed and serve as new source for tooth regeneration. Our findings brought insights of novel gene therapy approach that can alleviate the dental problems of patients with ARS and other developmental anomalies.

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11

Barnabei, Tabitha Richards. "Association of Masseter Muscle PITX2, ENPP1 and ESR1 Expression, Muscle Fiber Type, Temporomandibular Joint Disorders and Subclassifications of Craniofacial Asymmetry". Master's thesis, Temple University Libraries, 2017. http://cdm16002.contentdm.oclc.org/cdm/ref/collection/p245801coll10/id/454652.

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Oral Biology
M.S.
Craniofacial asymmetry is a dentofacial deformity with genetic influences. The genes PITX2, ENPP1 and ESR1 have multiple genetic associations with functional properties in muscle and bone. The objectives of this study are to investigate how PITX2, ENPP1 and ESR1 gene expression associates with four subclassifications of craniofacial asymmetry, temporomandibular disorders and fiber type differences compared between right and left masseter muscles. We developed an asymmetry classification that diagnosed four types of asymmetry with distinctive growth patterns: Group 1 – menton deviation without ramal difference (“mandibular body asymmetry”); Group 2 –menton deviation with shorter ramal height on the deviated side (“typical asymmetry”); Group 3 – shorter ramal height on the opposite side of menton deviation (“atypical asymmetry”); Group 4 – menton deviation with shorter ramal height and maxillary canting on the deviated side (“C-shaped asymmetry”). Some of these patients are at high risk for TMD; therefore, temporomandibular joint functioning is assessed as a routine part of the pre-surgical evaluation. TMD was diagnosed using the Diagnostic Criteria for TMD (DC/TMD). The clinical examination includes mandibular range of motion, palpation for pain, joint noise and bruxism. In addition, the Jaw Pain and Function (JPF) questionnaire was used to assess patient reported symptoms as an indication of perceived severity before and one year after orthognathic surgery. Masseter muscle samples were collected from 174 subjects undergoing surgical treatment for correction of malocclusion. Muscle serial cross-sections were mounted for immunostaining with five antibodies specific for myosin heavy chain (MyHC) isoform. We classified masseter fibers into 4 fiber type groups: type I, type I/II hybrid, type IIA and/or IIX, neonatal and atrial. With the remaining muscle samples, total RNA was isolated and PITX2, ENPP1, and ESR1 expression was quantified using TaqMan qRT-PCR. Average relative quantity gene expression values and percent differences between left and right masseter samples were calculated. In this population, there is a high prevalence of facial asymmetry (48%). Pre-surgical mean JPF scores are significantly different between symmetric (JPF=1.97) and asymmetric (JPF=6.9; p<0.001) patients; with scores ≥ 6 diagnostic for presence of TMD. ENPP1 and ESR1 expression is differentially expressed between right and left masseter muscle in patients with asymmetry. ENPP1 is differentially expressed in asymmetry group 4 (p=0.01) and ESR1 is differentially expressed in asymmetry group 1 (p=0.048), group 2 (p=0.004) and group 4 (p=0.02). Masseter fiber type properties of type I, type I/II hybrid and type II fibers associate with facial asymmetry and specific subclassifications, suggesting functional differences between type I, type I/II and type II fibers may be important factors in the development of symmetry between facial sides. There are significant differences in the left-right percent differences of fiber area of type I fibers in asymmetry group 3 (p=0.05), type I/II hybrid fibers in group 3 (p=0.02), and type II fibers in asymmetric patients (p=0.03), asymmetry group 2 (p=0.05) and group 4 (p=0.005). Additionally, there are significant differences in the left-right percent differences of percent occupancy of type I fibers in asymmetric patients (p=0.04), asymmetry group 2 (p=0.01) and group 3 (p=0.05) and type II fibers in asymmetry group 2 (p=0.04). By comparing gene expression with masseter muscle fiber type properties, we found significant results for PITX2 and ENPP1 suggesting their roles as genetic factors influencing jaw bone length and masticatory muscle strength in malocclusion. There are significant positive correlations between left-right percent differences of PITX2 and type I fiber area (r=0.86; p=0.03), type I/II hybrid fiber area (r=0.94; p=0.006), and type I/II hybrid fiber percent occupancy (r=0.90; p=0.01). Also, there are positive correlations approaching significance between left-right percent differences of ENPP1 and type I fiber area (r=0.80; p=0.06) and type I/II hybrid fiber area (r=0.75; p=0.09). Given the high prevalence of TMD in a population of patients with facial asymmetry, we compared differences in gene expression in masseter muscle of patients with specific TMD diagnostic conditions. Average PITX2 expression is significantly increased (p=0.0375) and average ENPP1 is increased, but not significantly, in all TMD patients diagnosed by the clinician. Average ESR1 is slightly increased compared to JPF scores and may be an essential factor for patient reported TMD symptoms. With these results, PITX2, ENPP1, and ESR1 should be considered biomarkers for asymmetry and TMD; however, further studies are needed to provide a more thorough understanding of the genetic influences on the craniofacial complex.
Temple University--Theses

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12

Donate, Puertas Rosa. "Omic approach to atrial fibrillation". Thesis, Lyon, 2017. http://www.theses.fr/2017LYSE1164.

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La fibrillation atriale (FA) est un problème de santé publique majeur dans le monde entier. Le remodelage électrique, structurel et neuronal est sous-jacent à la myopathie atriale. La pharmacothérapie actuelle est souvent inefficace en raison du manque de connaissance de la pathophysiologie de la FA.Pour comprendre comment se réalise le remodelage atrial, une approche Omique qui explore le transcriptome, l'épigénome (méthylome et microOme) et le génome de patients atteints de FA a été réalisée. Parallèlement, le phénotype de vieux rats spontanément hypertendus (SHRs) a été caractérisé et une étude pharmacologique avec la décitabine (5-Aza-2'-deoxycitidine) a été menée. Les patients atteint de FA présentent un profil transcriptomique et d'expression de miRNA alteré dans l'oreillete gauche (OG), soulignant le rôle important d'un processus de "œmorphogénèse de la structure anatomique". L'expression réduite de Pitx2 était inversement corrélée à la taille de l'OG et ne pouvait pas être expliquée ni par le facteur de transcription ni par la surexpression de Smyd2, une cible de miR-519b. Les SHRs, similairement aux observations chez l'homme, ont développé des arythmies dépendantes de l'âge associées au remodelage atrial et ventriculaire gauche. La FA a été trouvée associée à l'hyperméthylation du promoteur de Pitx2 à la fois chez l'homme et chez les SHRs. L'agent hyperméthylant décitabine a amélioré le profil arhytmique de l'ECG et les activités SOD, et la réduction de la fibrose dans le ventricule gauche des SHRs. En utillisant une approche NGS basée sur un panel personnalisé de 55 gènes candidats à la myopathie atriale dans une cohorte de 94 patients atteints de FA, 11 nouvelles variantes faux-sens potentiellement pathogènes impliqués dans le remodelage structurel ont été identifiés. Des études fonctionnelles de ces variants ont débuté. Trois patients sont également des porteurs de variantes dans les gènes connus de FA. Les résultats actuels suggèrent que 1) la régulation épigénétique peut jouer un rôle dans la pathophysiologie de la FA 2) les agents hypométhylants doivent être considérés comme une nouvelle thérapie de la FA 3) une approche Omique peut aider à découvrir de nouveaux mécanismes sous-jacents à la myopathie atriale
Atrial fibrillation (AF) is a major public health care problem worldwide. Electrical, structural, and neural remodeling underlie atrial myopathy. Current pharmacotherapy is often ineffective due to the lack of knowledge of AF pathophysiology. To understand how atrial remodeling occurs, an Omic approach that explore the transcriptome, epigenome (methylome and microOme) and genome of AF patients was performed. In parallel, ageing spontaneously hypertensive rats (SHRs) were phenotypically characterised and a pharmacological study with decitabine (5-Aza-2’-deoxycitidine) was conducted. AF patients presented an altered transcriptomic and microRNA expression profile in the left atria (LA), emphasizing the important role of an "anatomical structure morphogenesis" process. The Pitx2 reduced expression was inversely correlated with LA size, and could not be explained by transcriptor factor. Smyd2 is a target of miR-519b-3p. SHRs, similar to what is observed in humans, developed age-dependent arrhythmias associated with left atrial and ventricular remodeling. AF was found to be associated with Pitx2 promoter hypermethylation both in humans and in SHRs. The hypomethylating agent decitabine improved ECG arrhythmic profiles and superoxide dismutase activities, and reduced fibrosis in the left ventricle of SHRs. Using a next-generation sequencing approach based on a custom panel of 55 atrial myopathy candidate genes in a cohort of 94 AF patients, 11 novel potentially pathogenic missense variants involved in structural remodeling were identified. Functional studies of these variants have started. Three patients were also carriers of variants in known AF-causing genes. The present results suggest that 1) epigenetic regulation may play a role in the pathophysiology of AF 2) hypomethylating agents have to be considered as a new AF therapy 3) an Omic approach may help to uncover new mechanisms underlying atrial myopathy

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13

Cella, Wener Passarinho. "Rastreamento de mutações nos genes PITX2, FOXC1 e GJA1 em pacientes com sindrome de Axenfeld-Rieger associada a glaucoma". [s.n.], 2005. http://repositorio.unicamp.br/jspui/handle/REPOSIP/310116.

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Orientadores: Jose Paulo Cabral de Vasconcellos, Vital Paulino Costa
Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Ciencias Medicas
Made available in DSpace on 2018-08-04T19:48:33Z (GMT). No. of bitstreams: 1 Cella_WenerPassarinho_M.pdf: 8448689 bytes, checksum: a8a8a7a3ca810183c1f5f3f2964e8c29 (MD5) Previous issue date: 2005
Resumo: A Síndrome de Axenfeld-Rieger é uma entidade clínica rara, de transmissão autossômica dominante e grande variabilidade clínica, podendo chegar à penetrância incompleta. Manifesta-se clinicamente por malformações do segmento anterior do olho acompanhada ou não de malformações extra-oculares, das quais as mais comuns são alterações dos ossos craniofaciais e dos dentes e falência de involução da pele peri-umbilical. O principal fator de morbidade da síndrome é a associação com glaucoma de desenvolvimento, presente em aproximadamente 50% dos indivíduos afetados. Dois genes, PITX2, localizado no cromossomo 4q25, e FOXCl, no cromossomo 6 p25, estão associados à Síndrome de Axenfeld-Rieger. Recentemente, identificou-se o gene GJAl associado à Síndrome de Displasia Oculodentodigital, a qual compartilha aspectos clínicos com a Síndrome de Axenfeld-Rieger tais como malformações do segmento anterior ocular, glaucoma e alterações ósseas e dentárias, tornando-se, assim também, mais um gene candidato para a Síndrome de Axenfeld-Rieger. O objetivo deste estudo foi avaliar a ffeqüência e os tipos de mutações nos genes PITX2, FOXCl e GJAl em pacientes portadores da Síndrome de Axenfeld-Rieger associada a glaucoma, além de correlacionar possíveis alterações moleculares com aspectos clínicos dos pacientes. Para tanto, foram examinados oito indivíduos (casos-índice) acometidos pela síndrome associada a glaucoma, assim como seus familiares. Após avaliação oftalmológica e sistêmica, foram coletados 5ml de sangue periférico para extração de DNA genômico, o qual foi amplificado por reação em cadeia de polimerase utilizando-se pares de iniciadores específicos para as regiões codificadoras e limites íntronJexon dos genes PITX2, FOXCl e GJAl, procedendo-se com o seqüenciamento automático para o rastreamento de mutações. Não foram encontradas mutações no - gene PITX2. No gene FOXCl foram encontradas uma inserção (1359-1360insGGC), uma deleção (718-719deICT) e uma mutação de ponto do tipo sem sentido (TrpI52STOP) entre as famílias estudadas. O paciente portador da deleção no gene FOXCl era um caso isolado e apresentava apenas alterações oculares da síndrome. As outras duas mutações ocorreram em pacientes com manifestações oculares e sistêmicas da doença, estando a inserção presente em um caso isolado e a mutação sem sentido em seis indivíduos da mesma família com padrão de transmissão autossômico dominante. O rastreamento de mutações no gene GJAl evidenciou uma mutação de ponto do tipo sentido trocado (Ala253Val) em três indivíduos da mesma família, sendo que um deles era clinicamente normal e dois sintomáticos que apresentavam concomitantemente a mutação Trp152STOP no gene FOXCl. Esta é a primeira descrição de uma mutação no gene GJAl em pacientes inequivocamente portadores da Síndrome de Axenfeld-Rieger. As ITeqüências de mutações observadas nos genes PITX2, FOXCl e GJAl em oito famílias brasileiras com Síndrome de Axenfeld-Rieger e glaucoma de desenvolvimento associado foram de 0%, 37,5% e 12,5%, respectivamente, sendo esta última em combinação com a mutação Trp152STOP no gene FOXCl. Apesar da amostragem reduzida, observou-se uma tendência a maior agressividade do glaucoma em pacientes portadores de mutação somente no gene FOXCl do que nos indivíduos portadores de mutação concomitante no gene FOXCl e no gene GJAl, sugerindo um possível efeito protetor da mutação Ala253Val no gene GJAl nesta família. Outros estudos são necessários, contudo, para definir a função do gene GJAl na etiopatogênese da Síndrome de Axenfeld-Rieger
Abstract: Axenfeld-Rieger Syndrome (ARS) is arare disorder, usually transmitted in an autosomal dominant pattem characterized by anterior segment dysgenesis and often associated with developmental glaucoma. In addition to the ocular changes observed in ARS, syndromic features can also occur, such as facial bone defects, teeth anomalies and peri-umbilical skin involution. Two transcription factor genes, PITX2 on chromosome 4q25 and FOXCl on chromosome 6p25, have been associated with the ARS phenotype through mutational events. Recently, the GJAl gene (connexin 43), associated with oculodentodigital dysplasia (ODDD) syndrome, which presents some similarities with ARS, was identified. The ODDD syndrome is characterized by malformations that involve the face, eyes, teeth and bones. The ocular abnormalities include microphthalmos and anterior segment dysgenesis that may lead to glaucoma as well. The main purpose of this study was to evaluate FOXCl, PITX2 and GJAl genes mutations in Brazilian patients with ARS. Eight unrelated patients atfected by ARS (all of them with glaucoma and 5 without systemic malformations) and their families were ophthalmologically evaluated and had their blood collected for DNA extraction purposes. The coding regions and íntron/exon boundaries of these genes were completely evaluated through direct sequencing. Among the 8 patients, 3 (37,5%) presented with ditferent structural alterations in the FOXCl gene. A deletion in heterozygosis oftwo bases downstream the forkhead domaÍn was observed in a patient with no systemic malformations (718-719deICT). An insertion ofthree bases, also downstream the forkhead domain, was identified in a patient with systemic malformation (1359-1360insGGC). A new nonsense mutation (Trp152STOP) was identified in the forkhead domain of the FOXCl gene in another patient with ARS and systemic alterations as well. One patient harbored the mutation Ala253Val in the GJAl gene (12,5%). No mutations were identified in the PITX2 gene among these individuaIs. Patients who carried the GJAl (Ala253Val) and FOXCl (TrpI52STOP) mutations (:&om the same family) developed less severe glaucoma compared with family members presenting FOXCl (TrpI52STOP) mutation alone. Three new structural alterations of the FOXCl gene and one new mutation in the GJAl gene were described in Brazilian patients with ARS and the :&equencyof mutations in the PITX2, FOXCl and GJAl genes in this study were 0%, 37,5% and 12,5%, respectively. Despite the small number of patients, we found a slight trend to more severe glaucoma in patients with FOXCl mutations compared to those without them, except in the two patients with FOXCl and GJAl associated mutations, suggesting an attenuation effect of GJAl gene mutation (Ala253Vai). However, other studies are necessary to define the exact role ofGJAl in ARS
Mestrado
Oftalmologia
Mestre em Ciências Médicas

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Chaney, Beth A. "DNA Recognition by the K50 Class Homeodomain PITX2: Solution Structure, Molecular Dynamics, and Implications for Mutations That Cause Rieger Syndrome". Cincinnati, Ohio : University of Cincinnati, 2005. http://www.ohiolink.edu/etd/view.cgi?acc%5Fnum=ucin1121276740.

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15

Kuhlmann, Stefan Michael [Verfasser]. "Defining the molecular cause of an altered response to anti-arrhythmic drugs in Pitx2-dependent atrial fibrillation / Stefan Michael Kuhlmann". Berlin : Medizinische Fakultät Charité - Universitätsmedizin Berlin, 2019. http://d-nb.info/120204154X/34.

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16

GIANNETTI, FEDERICA. "HIPS AND MES AS A CELLULAR MODEL TO STUDY THE MECHANISMS BEHIND ARRHYTHMIAS". Doctoral thesis, Università degli Studi di Milano, 2022. http://hdl.handle.net/2434/903405.

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Mouse embryonic stem cells (mESc) and human induced pluripotent stem cells (hiPSc) are commonly exploited in research for their ability to potentially differentiate into any cell subtype. Regarding cardiac differentiation, today's existing protocols are well advanced in recapitulating and promoting cardiomyocytes (CMs) differentiation, making CMs derived from mESc and hiPSc a good in vitro model for analyzing heart pathologies. In this thesis, these models have been used to characterize and better comprehend the functional output caused by genetic alterations. In the first part, mESc-derived cardiomyocytes help to elucidate the electrical dysregulation produced by the knock-out of Striatin (STRN), a scaffold protein with a role in organizing subcellular multiprotein complexes. STRN loss has been found in patients with dilated cardiomyopathy (DCM) and its mutation linked to DCM and ARVC in dogs. However, STRN physiological role in the cardiac context is not yet clear, for this reason the electrical characterization, by patch-clamp analysis, of mouse embryonic stem cells mESc-derived cardiomyocytes (mCMs) knock out for the STRN gene (STRN-KO) may help elucidating STRN role and its pathological implications. In this context, action potential (AP) analysis showed a higher rate and upstroke velocity of STRN-KO mCMs compared to its isogenic WT control. Accordingly, a significantly larger INa current density, responsible of the phase 0, was found in STRN-KO than in WT mCMs, while the other currents analyzed (If, ICaL and IKr) were similar. Literature data show that downregulation of STRN destabilize microtubules, and their disassembling increases sodium current in neonatal cardiomyocytes. Here is shown that treating STRN-KO mCMs with the microtubule stabilizer Taxol (10 μM) reverted INa density to values similar to WT cells. These data lead to the conclusion that STRN loss affects the electrical properties of mCMs likely acting on cytoskeleton stability and consequently on sodium channels trafficking and/or activity, creating a substrate more prone to develop arrhythmias. In the second part of the thesis, hiPSc-derived cardiomyocytes, obtained from a patient with atrial fibrillation (AF)carrying a heterozygous GOF point mutation in the PITX2 gene, are used to distinguish if this genetic condition predispose or AF. In GWAS studies, PITX2 is the genetic locus most associated with AF, it is often dysregulated in AF patients and its presence or absence correlates with electrical alterations. Moreover, PITX2 loss of function in zebrafish has been linked to perturbations of metabolic pathways forerunning AF-like phenotypes. APs recorded from small hiPSc-derived CMs (hpCMs) clusters reveals that PITX2-hpCMs are bradycardic, with a larger action potential amplitude and a shorter action potential duration than to three unrelated controls. Moreover, preliminary data obtained from atrial hiPSc-derived CMs, used to further elucidate the consequence of this mutation in the atrial background, showed no differences in ICaL and If densities; however, activation curve of If is negatively shifted with a higher inverse slope factor in PITX2 atrial hiPSc-derived CMs, implying a minor contribution of this current during the slow diastolic depolarization. Seahorse metabolic analysis indicates that PITX2-haCMs generates more ATP compared to isogenic control, and this difference is due to increased oxidative phosphorylation in mitochondria. In conclusion, although a more detailed characterization will be required, the PITX2 GOF mutation seems to enhance oxidative phosphorylation and a slower beating rate, altering not only the electrical activity but also the metabolism of cardiomyocytes. Overall, these in vitro models of cardiomyocytes were ideal to identify the functional outcome of genetic alteration, either to investigate the electrical mechanism behind arrhythmias or to characterize the role of a specific protein in the specific cardiac context.

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Vaideanu-Collins, Daniela. "An association study of PITX2 polymorphism in a cohort of patients with primary open angle glaucoma and considerations on the genetics of glaucoma". Thesis, University of Newcastle Upon Tyne, 2010. http://hdl.handle.net/10443/1061.

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Background: Glaucoma is a major cause of blindness world-wide. There is a need for methods to identify individuals at risk of developing glaucoma, so that early treatment can prevent visual loss. Genetic screening tests offer the prospect of pre-symptomatic diagnosis of at risk individuals. There is now strong evidence that a number of different genes are associated with glaucoma susceptibility. Mutations in the PITX2 homeobox transcription factor gene disrupt normal development of the anterior segment and cause overt structural abnormalities. It is possible that, as yet undetected mutations/polymorphisms in PITX2 may produce subtle and undetected abnormalities in anterior segment structure and function that could predispose to glaucoma. Purpose: The aim of this thesis is two fold: 1. Screening for the presence of single nucleotide polymorphisms in PITX2 gene in a cohort of 100 unrelated primary open angle glaucoma/ ocular hypertension patients, 10 Posterior embryotoxon subjects and 100 age and ethnically matched controls to establish the mutation spectrum. 2. Identification, phenotyping and recruitment for genetic studies of primary open angle glaucoma patients with strong family history of glaucoma. Materials and methods: 1. 100 primary open angle glaucoma patients and 60 age and ethnically matched controls were enrolled in the study. Patients and controls were phenotyped and a blood sample for DNA extraction collected. PITX2 exon-specific primers were used to PCR amplify patient and control DNA. Direct sequencing was used to screen for sequence alterations in the entire coding sequence of PITX2 gene. Concurrently, polymorphic sites reported in the PITX2 gene were identified from the NCBI and Ensemble databases and the frequency of polymorphic sites was investigated. The SHEsis and UNPHASED software packages were used for statistical analysis. 2. Patients diagnosed with glaucoma and strong family history were identified from Glaucoma Unit at Sunderland Eye Infirmary, phenotyped and enrolled in the study. The pedigrees were constructed and interested relatives enrolled in the study and phenotyped. A sample of blood for DNA extraction was collected from all people enrolled in the study. Results: 1. Direct sequencing did not identify any sequence variation in the coding region. 26 PITX2 polymorphic sites were identified from the internet databases, including five in the coding sequence. Sixteen non coding SNPs were confirmed within our study group and SNP frequencies were examined. None of the coding sequence SNPs was identified in our cohort, demonstrating a high degree of sequence conservation. Also, none of the SNPs confirmed in this study group showed an increased frequency in the primary open angle glaucoma group compared with the control group. 2. Thirty-three pedigrees were identified with strong family of glaucoma during the time allowed for patient recruitment. Of these, twenty-two agreed to take part in the study. Thirteen pedigrees are presented in this study, mostly demonstrating autosomal dominant inheritance. Conclusion: There is ample evidence to suggest that genetics play an important role in unravelling the pathogenesis of glaucoma. Identification and recruitment of patients for genetic studies is an essential step and the role of the clinician in this process is paramount. Also, developmental glaucoma genes are an important group of genes to be screened in primary open angle glaucoma/ocular hypertension patients, as they may play a role in the pathogenesis of this preventable blinding disease.

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18

Ziyadeh, Azza. "Nouveaux mécanismes contribuant à la variabilité phénotypique de mutations N- et C-terminales du canal sodique cardiaque". Phd thesis, Université Pierre et Marie Curie - Paris VI, 2014. http://tel.archives-ouvertes.fr/tel-01037923.

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Les mutations du gène SCN5A, codant la sous-unité ? du canal Na+ cardiaque Nav1.5, sont responsables d'arythmies cardiaques héréditaires. La pénétrance incomplète observée dans ces maladies suggère l'existence d'autres facteurs modulant le phénotype associé à ces mutations. Dans ce travail de thèse, nous avons caractérisé deux mutations identifiées dans SCN5A. Le mutant R104W, identifié chez un patient atteint du syndrome de Brugada, est retenu dans le réticulum endoplasmique (RE), dégradé par le protéasome et abolit le courant Na+. Co-exprimé avec le canal sauvage, R104W conduit à la rétention de celui-ci dans le RE, résultant en un effet dominant négatif sur les canaux sauvages. Nous avons démontré que ce nouveau mécanisme mettait en jeu une interaction entre les sous-unités ? de Nav1.5. La mutation R1860Gfs*12 a été identifiée dans une famille présentant des arythmies auriculaires. Dans un système d'expression hétérologue, ce mutant induit à la fois une perte et un gain de fonction de Nav1.5. La modélisation informatique nous a permis de montrer que la perte de fonction était plus prononcée dans les cellules auriculaires que ventriculaires. De plus, nous avons montré que la présence de polymorphismes en amont du gène PITX2 dans cette famille pouvait expliquer la variabilité des phénotypes observés. En conclusion, l'interaction entre les sous-unités ? de Nav1.5, les propriétés électriques différentes entre oreillette et ventricule et la présence de polymorphismes chez les patients porteurs de mutations SCN5A sont des facteurs importants dans l'interprétation des effets fonctionnels de ces mutations, contribuant à la variabilité phénotypique des canalopathies Na+.

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19

Acunzo, Julie. "Thérapie génique des adénomes hypophysaires humains in vitro : Evaluation du rôle du récepteur somatostatinergique sst2 et d'un dominant négatif du facteur de transcription Pitx2". Aix-Marseille 2, 2009. http://www.theses.fr/2009AIX20696.

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Les adénomes hypophysaires humains sont des tumeurs développées à partir d’un des cinq types cellulaires constituant l’antéhypophyse. Parmi ces adénomes, on retrouve les adénomes somatotropes, lactotropes, gonadotropes, corticotropes et thyréotropes. Ces tumeurs sont souvent à l’origine d’une hypersécrétion hormonale parfois responsable de complications cardiaques et respiratoires à l’origine d’une augmentation de la mortalité. Pour les adénomes non-fonctionnels (Non Functioning Pituitary Adenoma, NFPA), le plus souvent d’origine gonadotrope, il n’y a pas d’hypersécrétion hormonale et la découverte repose alors sur des signes de compressions des structures avoisinantes. Dans cette étude, nous nous sommes intéressés aux adénomes les plus fréquents, les adénomes lactotropes, somatotropes et les NFPA. Si la chirurgie est efficace pour réduire le volume tumoral, elle permet plus rarement une guérison complète. Pour les adénomes somatotropes et lactotropes des traitements pharmacologiques (les analogues de la somatostatine et les analogues de la dopamine) permettent d’inhiber l’hypersécrétion hormonale et parfois de réduire le volume tumoral, mais une proportion non négligeable de patients reste résistante à ces thérapeutiques pharmacologiques. Pour les NFPA, il n’existe aucune thérapeutique spécifique. Dans ce contexte, de nouvelles modalités de traitement semblent nécessaires. La thérapie génique pourrait alors représenter une nouvelle approche thérapeutique pour ce type de tumeur. Le facteur de transcription Pitx2 est surexprimé dans les adénomes gonadotropes (NFPA) et impliqué dans une voie de prolifération cellulaire. Pour obtenir un ciblage spécifique des adénomes non-fonctionnels, notre première stratégie a été d’utiliser un dominant négatif du facteur de transcription Pitx2 (R91P) et des siRNA dirigés contre Pitx2 dans le but de transduire et d’inhiber la viabilité cellulaire des NFPA. Ainsi, nous avons mis en évidence un effet apoptotique caspase dépendant de R91P et des siRNA sur une lignée gonadotrope murine et sur des NFPA humains in vitro. Le récepteur somatostatinergique sst2 est fortement impliqué dans l’inhibition de la sécrétion de GH et l’absence de ce récepteur dans les adénomes somatotropes est responsable de la résistance pharmacologique. Notre deuxième stratégie a donc été d’utiliser le récepteur somatostatinergique sst2 dans le but de réinduire son expression dans des adénomes somatotropes et lactotropes et de restaurer ainsi la sensibilité de ces tumeurs aux traitements médicaux. Suite à un transfert du gène sst2 in vitro, nous avons réussi pour la première fois à rétablir une sensibilité pharmacologique des adénomes somatotropes et lactotropes résistants aux analogues actuels. Notre étude met aussi en évidence un effet apoptotique caspase dépendant de sst2 lui même et une inhibition par sst2 de la sécrétion mais aussi de l’expression de la GH et de la PRL, indépendamment de l’activation par un ligand. Ces effets ligands indépendants suggèrent une activité constitutive de ce récepteur. A côté de cet objectif thérapeutique, cette étude sur le transfert de gènes nous a également permis de mieux comprendre le rôle de la surexpression de Pitx2 et de la perte d’expression du récepteur sst2 dans la tumorigénèse hypophysaire. Ces deux stratégies de transfert de gènes, l’une privilégiant la spécificité (Pitx2), l’autre l’amélioration des traitements actuels semblent prometteuses, mais nécessitent une validation sur des modèles précliniques.

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20

Rabe, Nadine. "Spinal Control of Locomotion : Developmental and Functional Aspects". Doctoral thesis, Uppsala universitet, Genetisk utvecklingsbiologi, 2010. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-112472.

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Neuronal networks are the central functional units of the nervous system. Knowledge about the identity of participating neurons and the assembly of these during development is crucial for the understanding of CNS function. A promising system to dissect the development and functionalities of a neuronal network is the central pattern generator (CPG) for locomotion. We used screening approaches to identify spinal neuronal subpopulations by their specific gene expression, potentially involved in CPG function. Amongst others we found paired-like homeodomain transcription factor 2 (Pitx2) as a cholinergic interneuron marker for partition cells, with a possible role in the spinal network for locomotion. In addition, we present two genes, Chondrolectin (Chodl) and Estrogen-related receptor beta (ERRβ) as novel markers for fast and slow motor neurons, respectively. The neuronal components of the CPG integrate three key functions; rhythm generation, ipsilateral flexors/extensors coordination and bilateral coordination over the midline. Commissural interneurons (CINs) are considered to participate in the latter. During development axons are guided to their targets by the help of axon guidance molecules. Netrin-1 and its receptor DCC (Deleted in Colorectal Cancer) have been shown to play an important role for spinal cord neurons in axon-pathfinding and migration towards the midline. We show that loss of netrin-1 functionally results in a switch from alternating to synchronous left-right locomotor activity and deletion of DCC surprisingly leads to a different phenotype, best described as uncoordination. Thus, during development, netrin-1 and DCC play a critical role for the establishment of a functional balanced CPG. Further we show a selective loss of CINs, predominantly from dorsally originating subtypes, not affecting the ventral-most V3 subtype in netrin-1 mutant mice, but a loss of CINs from all progenitor domains in Dcc mutant mice. Together, our data suggest a netrin-1-independent mechanism for DCC in axon guidance and a role of the most ventral originating CINs as part of the neuronal network controlling synchronous activities over the midline. Another pair of axon guidance molecules, EphA4 and ephrinB3, has been shown to cooperate in preventing ipsilateral interneurons from crossing the spinal midline and if either molecule is deleted in mice, this will result in a defect in left-right coordination of locomotion. We provide in vivo and in vitro evidence that the GTPase-activating protein α2-chimerin, as a downstream molecule of EphA4 signaling, is essential in axon guidance decisions involved in the correct formation of the spinal circuitry for locomotion.

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21

Ziyadeh, Azza. "Nouveaux mécanismes contribuant à la variabilité phénotypique de mutations N- et C-terminales du canal sodique cardiaque". Electronic Thesis or Diss., Paris 6, 2014. http://www.theses.fr/2014PA066070.

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Les mutations du gène SCN5A, codant la sous-unité ? du canal Na+ cardiaque Nav1.5, sont responsables d'arythmies cardiaques héréditaires. La pénétrance incomplète observée dans ces maladies suggère l'existence d'autres facteurs modulant le phénotype associé à ces mutations. Dans ce travail de thèse, nous avons caractérisé deux mutations identifiées dans SCN5A. Le mutant R104W, identifié chez un patient atteint du syndrome de Brugada, est retenu dans le réticulum endoplasmique (RE), dégradé par le protéasome et abolit le courant Na+. Co-exprimé avec le canal sauvage, R104W conduit à la rétention de celui-ci dans le RE, résultant en un effet dominant négatif sur les canaux sauvages. Nous avons démontré que ce nouveau mécanisme mettait en jeu une interaction entre les sous-unités ? de Nav1.5. La mutation R1860Gfs*12 a été identifiée dans une famille présentant des arythmies auriculaires. Dans un système d'expression hétérologue, ce mutant induit à la fois une perte et un gain de fonction de Nav1.5. La modélisation informatique nous a permis de montrer que la perte de fonction était plus prononcée dans les cellules auriculaires que ventriculaires. De plus, nous avons montré que la présence de polymorphismes en amont du gène PITX2 dans cette famille pouvait expliquer la variabilité des phénotypes observés. En conclusion, l'interaction entre les sous-unités ? de Nav1.5, les propriétés électriques différentes entre oreillette et ventricule et la présence de polymorphismes chez les patients porteurs de mutations SCN5A sont des facteurs importants dans l'interprétation des effets fonctionnels de ces mutations, contribuant à la variabilité phénotypique des canalopathies Na+
Mutations in the SCN5A gene, which encodes the α-subunit of the cardiac sodium channel Nav1.5, are implicated in different inherited cardiac arrhythmias. The incomplete penetrance observed in these diseases suggests the existence of other factors modulating the phenotype of these mutations. In this thesis work, we characterized two mutations identified in SCN5A. The R104W mutant identified in a patient with Brugada syndrome is retained in the endoplasmic reticulum (ER), degraded by the proteasome and abolishes the sodium current. Co-expressed with wild type (WT) channels, R104W leads to WT channels ER retention, causing a dominant-negative effect. We demonstrated that interaction between Nav1.5 α-subunits is responsible for the retention and the dominant-negative effect. The R1860Gfs*12 mutation was identified in a family with atrial arrhythmias. In a heterologous system, this mutant induces both loss- and gain-of-function effects on Nav1.5. Computer-model simulation showed that the loss-of-function was more pronounced in atrial than in ventricular cells. In addition, we showed that the presence of polymorphisms upstream of the PITX2 gene could explain the observed phenotypic variability in this family. In conclusion, the interaction between the α-subunits of Nav1.5, the different electrical properties between atria and ventricles and the presence of polymorphisms in patients with SCN5A mutations, are important factors in the interpretation of the functional effects of these mutations, which could explain the phenotypic variability of sodium channelopathies

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22

Gore, Panter Shamone Robinette. "Genetic and Functional Studies of LociAssociated with Atrial Fibrillation". Case Western Reserve University School of Graduate Studies / OhioLINK, 2014. http://rave.ohiolink.edu/etdc/view?acc_num=case1396521127.

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23

Larsson, Mats. "Human Iris Characteristics as Biomarkers for Personality". Doctoral thesis, Örebro University, Department of Behavioural, Social and Legal Sciences, 2007. http://urn.kb.se/resolve?urn=urn:nbn:se:oru:diva-1684.

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This dissertation explains why behavioral genetic research can be better informed by using characteristics in the human iris as biomarkers for personality, and is divided into five parts. Part I gives an introduction to the classical twin method and an overview of the findings that have led most developmental researchers to recognize that the normal variation of personality depends on a complex interplay between genetic and environmental factors. Part II highlights empirical findings that during the last twenty years have gradually moved genetic and environmental theory and research to evolve toward one another, and also presents the theory of genetics and experience that currently is used to explain how the interplay between genes and the environment works. Part III explains why, from a developmental perspective, it is of interest to identify candidate genes for personality, and gives a brief overview of genes that have been associated with personality. Problems associated with genetic research on the molecular level and how these apply to personality are also highlighted. Part IV examines molecular research on the iris and the brain, which suggests that genes expressed in the iris could be associated with personality, and explains how the use of iris characteristics can increase power to test candidate genes for personality by taking advantage of the self-organizing properties of the nervous system. The empirical foundation for the questions posed in this dissertation and also the empirical results are presented here. Part V discusses the associations found between iris characteristics and personality, and exemplifies how iris characteristics can be used within the theoretical frameworks presented in parts I, II, III and IV. In other words, Part V explains how iris characteristics – in addition to identify as well as test candidate genes for personality – can be used to investigate how people’s experiences in themselves are influenced by genetic factors.

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24

Hüttner, Johann [Verfasser]. "Die cis-regulatorische Landschaft von PITX1 / Johann Hüttner". Berlin : Medizinische Fakultät Charité - Universitätsmedizin Berlin, 2016. http://d-nb.info/1119803748/34.

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25

Chang, Wing Yean. "Role of the paired-like gene Pitx1 in Xenopus head development". Thesis, National Library of Canada = Bibliothèque nationale du Canada, 2000. http://www.collectionscanada.ca/obj/s4/f2/dsk1/tape2/PQDD_0023/MQ62198.pdf.

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26

Di, Giorgio Luciano. "Identification of critical amino acids in the N-terminus of Pitx2c". Thesis, McGill University, 2005. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=82224.

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Abstract (sommario):

Pitx2 is a homeodomain transcription factor. Three functional protein isoforms are encoded by the Pitx2 gene that differ only in their N-terminus. Functional studies have demonstrated that the Pitx2c isoform is the most downstream component of the left-right patterning cascade that is responsible for positioning the internal organs relative to the midline.
In all vertebrates, mRNA for the Pitx2c isoform is expressed asymmetrically in the left lateral plate mesoderm and in organs that become asymmetrically positioned. A Pitx2c isoform specific antibody was created and used to determine the pattern of Pitx2c protein expression in the chick embryo. Pitx2c protein expression appears to be identical to the pattern of Pitx2c mRNA expression.
I hypothesized that the Pitx2c N-terminus would be able to independently act as a dominant negative and interfere with the left-right patterning activity of endogenous Pitx2c protein. To test this hypothesis, the Pitx2c N-terminus was overexpressed in the left lateral plate mesoderm. The first morphological sign of left-right asymmetry is the rightward looping of the heart. The heart looped to the left side in 30% of the infected embryos suggesting that the Pitx2c N-terminus was acting as a dominant negative protein and interfering with the activity of the endogenous Pitx2c. Mutation analysis indicated that the amino acid residues 41 to 45, LAMAT, were required for the dominant negative activity of Pitx2cN.

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27

Wong, Shian Yea. "Identifying protein interaction partners of the Pitx2c N-terminus during embryogenesis". Thesis, McGill University, 2014. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=123305.

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The vertebrate body is patterned asymmetrically along the left-right axis. Left-right patterning is required to establish correct asymmetric organ formation and positioning, and ultimately it is essential for normal physiological functioning. Misregulation in this process can lead to severe physiological defects in multiple organs, including the heart and gut. Pitx2c is a paired-like homeodomain transcription factor and is crucial for translating left-right signals into asymmetric morphogenesis. It is asymmetrically expressed in the left lateral plate mesoderm and continues to be expressed on the left side of organ primordia that are destined to become asymmetrically formed or positioned organs. Previous work studying Pitx2c has demonstrated its evolutionarily conserved role in left-right patterning. While the homeodomain is important for Pitx2c's function, our lab identified a putative interaction domain in the N-terminus of Pitx2c that is required for its function in left-right patterning. To further characterize the role of Pitx2c, I used a yeast two-hybrid screen to identify candidate protein interaction partners of its N-terminus. Thirty-two candidates were identified from the screen. Candidates that were isolated at least twice were prioritized for further analysis. After eliminating non-coding clones, false positive interactions, and confirming interactions of the candidate putative interaction domains with the Pitx2c N-terminus in yeast, six candidates remained: Anti-silencing function 1 homolog B (S. cerevisiae; Asf1b), Eukaryotic translation initiation factor 3, subunit A (Eif3a), Eukaryotic translation initiation factor 3, subunit M (Eif3m), Niemann-Pick C type 2 (Npc2), Serine/arginine-rich splicing factor 2 (Srsf2), and Serine/arginine-rich splicing factor 15 (Srsf15). mRNA Expression patterns of these six Pitx2c candidate protein interaction partners was analyzed by whole mount in situ hybridization and compared to Pitx2c expression patterns. All candidates were expressed symmetrically in the embryo, including tissues where Pitx2c is asymmetrically expressed. In the chick embryo, Npc2 and Srsf2 were expressed in a tissue-specific manner from HH stage 10 to HH stage 26, while Eif3a and Eif3m were broadly expressed throughout the embryo from HH stage 10 and only became enriched in particular tissues at HH stage 24 to HH stage 26. All candidates were coexpressed with Pitx2c in the looped heart tube. Only Srsf2 was coexpressed with Pitx2c in the lateral plate mesoderm. The known functions of these candidates are consistent with the known role of Pitx2c in transcriptional regulation and suggests new roles for Pitx2c in mRNA processing and translational regulation during left-right patterning.
Le corps des vertébrés est structuré asymétriquement le long de l'axe gauche-droit. La latéralité est requise afin d'établir la formation des organes ainsi que leur positionnement qui, ultimement, sont essentielles pour un fonctionnement physiologique normal des vertébrés. Lors de ce processus, une mauvaise régulation peut causer des troubles physiologiques sévères dans plusieurs organes incluant le cœur et l'intestin.Le facteur de transcription homéodomaine Pitx2c joue un rôle essentiel lors de la traduction des signaux gauche-droit en morphogénèse asymétrique des organes. Pitx2c est exprimé de façon asymétrique dans le mésoderme de la plaque latérale gauche et continu d'être exprimé du côté gauche des futurs organes asymétriques. Quelques études ont démontré que Pitx2c a un rôle qui est conservé lors de l'évolution dans la latéralité. Le rôle de l'homéodomaine est important pour la fonction de Pitx2c et nous avons démontré dans notre laboratoire qu'un domaine d'interaction de la partie N-terminale de Pitx2c est aussi important pour sa fonction lors de la latéralité.Afin de caractériser d'avantage le rôle de Pitx2c, j'ai utilisé la méthode de double hybride afin d'identifier des protéines candidates d'interaction avec la partie N-terminale de Pitx2c. Trente-deux candidats ont été sélectionnés. De ce nombre, les faux positifs ainsi que les candidats qui représentent région la non-codante d'une protéine ont été supprimés ce qui a permis de réduire la liste à six candidats : Anti-silencing function 1 homolog B (S. cerevisiae; Asf1b), Eukaryotic translation initiation factor 3, subunit A (Eif3a), Eukaryotic translation initiation factor 3, subunit M (Eif3m), Niemann-Pick C type 2 (Npc2), Serine/arginine-rich splicing factor 2 (Srsf2), ainsi que Serine/arginine-rich splicing factor 15 (Srsf15).Les patrons d'expression des six protéines candidates potentielles d'interaction avec Pitx2c ont été examinés par hybridation in situ et ont été comparé avec le patron d'expression de Pitx2c. Chez tous les candidats, l'expression asymétrique dans les embryons de poulet a été observée incluant dans les tissus où Pitx2c est aussi exprimé asymétriquement. Npc2 et Srsf2 sont exprimés de façon spécifique dans certains tissus à partir du stade HH10 chez le poulet. Eif3a et Eif3m sont largement exprimés dans les embryons à partir du stade HH10 mais leur expression est enrichie dans certains tissus à partir du stade HH22. L'expression de tous les candidats coïncide avec celle de Pitx2c dans la courbure du tube cardiaque. Seulement l'expression de Srsf2 coïncide avec celle de Pitx2c dans le mésoderme de la plaque latérale gauche. Les fonctions des différents candidats suggèrent que ces partenaires potentiels d'interactions de la partie N-terminale de Pitx2c pourraient jouer un rôle important lors de la régulation de la transcription, la formation de l'ARN messager et la régulation de la traduction lors de la latéralité.

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Siontas, Dora. "The identification of proteins that interact with the N-terminal domain of Pitx2c". Thesis, McGill University, 2012. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=106609.

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Abstract (sommario):

During embryogenesis, a number of key events regulate the correct patterning of an organism, one of them being the correct positioning of internal organs. One of the factors important for this process is the homeodomain transcription factor Pitx2c,which is asymmetrically expressed in the left lateral plate mesoderm. Previous work has shown that the N-terminal domain of Pitx2c is important for left-right patterning, in that overexpression of the N-terminus randomizes the direction of heart tube looping. It is believed that the overexpressed N-terminus is antagonizing the activity of endogenous Pitx2c by competing for binding to one or more critical interaction partners. In order to better understand the role of the N-terminal domain, I have performed two different yeast two-hybrid library screens to identify proteins that interact with this domain. In the first, I used the chick Pitx2c N-terminal domain, cloned into the pGBKT7 vector in-frame with the DNA binding domain of the GAL4 transcription factor as bait, against a mouse adult cDNA library cloned in-frame with the GAL4 activation domain in the pGADT7 vector. In the second, I used a mouse Pitx2c N-terminal domain against an embryonic E11 mouse cDNA library. The resulting interacting candidates were analyzed and four were chosen based on the number of times they appeared in the screens and their function. The four chosen were Npc2, Ubc, Ube2n and Ube2e1. GST pull-down experiments were performed to confirm their interaction with the mouse and chick Pitx2c N-terminal domains, and proteins for the interacting candidates were generated using a coupled transcription translation system. Only Ube2n and Ube2e1 were confirmed as interacting candidates, suggesting that Pitx2 may be ubiquitinated and this may be a regulatory mechanism. Future experiments will provide more insight on their role in interacting to the N-terminal domain of Pitx2c.
Durant l'embryogenèse, un certain nombre d'événements clés réglementent la structuration correcte d'un organisme, l'un d'eux étant le positionnement correct des organes internes. Un des facteurs importants impliqué dans ce processus est le facteur de transcription homéodomaine Pitx2c, qui est asymétriquement exprimé dans le mésoderme de la plaque latérale gauche. Des études antérieures ont démontrées que le domaine N-terminal de Pitx2c est important pour la structuration gauche-droite, puisque la surexpression de l'extrémité N-terminale randomise la direction de la courbure du coeur. Nous croyons que la surexpression de la portion N-terminale de Pitx2c antagonise son activité endogène en faisant concurrence pour la liaison à un ou plusieurs partenaires d'interaction critique. Afin de mieux comprendre le rôle du domaine N-terminal de Pitx2c, j'ai effectué deux différentes sélections en utilisant la méthode de double hybride dans le but d'identifier les protéines qui interagissent avec ce domaine. Lors de la première sélection, j'ai utilisé le domaine N-terminal Pitx2c du poulet, cloné dans le vecteur pGBKT7 avec le domaine liant l'ADN du facteur de transcription GAL4 comme appât, contre une librairie adulte d'ADNc de souris avec le domaine d'activation GAL4 dans le vecteur pGADT7 comme proie. Dans la seconde, j'ai utilisé le domaine N-terminal Pitx2c de la souris contre une librairie d'ADNc d'embryon de souris E11. Les candidats qui interagissent ont été analysés et quatre ont été choisis en fonction du nombre de fois qu'ils apparaissent dans les sélections et selon leur function: Npc2, Ubc, Ube2n et Ube2e1. Des immunoprécipitations utilisant le glutathion-S-transférase comme étiquette ont été réalisées afin de confirmer l'interaction entre les partenaires potentiels et les domaines N-terminaux de Pitx2c de la souris et du poulet. Des protéines pour les candidats potentiels d'interactions avec Pitx2c ont été générées en utilisant un système de transcription-traduction. Ube2n et Ube2e1 sont les seuls candidats qui ont été confirmés comme protéines d'interaction, mettant en avant l'ubiquitination comme étant un mécanisme putatif par lequel Pitx2c agit dans la structuration gauche-droite durant le développement embryonnaire.

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Boorman, Clive John. "Pitx in the evolution of chordate left- right asymmetry and the nasohypophysial placode". Thesis, University of Reading, 2002. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.250600.

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30

Müller, André Nicolai [Verfasser], e Gunther [Akademischer Betreuer] Döhlemann. "Funktionelle Charakterisierung des Ustilago maydis Effektorproteins Pit2 / André Nicolai Müller. Betreuer: Gunther Döhlemann". Marburg : Philipps-Universität Marburg, 2014. http://d-nb.info/1057000124/34.

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31

Maxwell, Sarah L. "The role of the homeodomain protein Pitx3 in the development and survival of midbrain dopaminergic neurons". Thesis, University of Edinburgh, 2006. http://hdl.handle.net/1842/1356.

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There is much interest in the study of midbrain dopaminergic (mDA) neurons as their functions include the regulation of motor function, emotion and reward pathways. Furthermore the dysfunction of these neurons is implicated in a number of human disorders such as Parkinson’s disease (PD), addiction and schizophrenia. PD is characterised by the degeneration of mDA neurons of the substantia nigra pars compacta (SNc), therefore, research into the specification and development of mDA neurons is of particular interest in relation to this disease. An understanding of the development of mDA neurons may lead to new methods of preventing their degeneration or potentially a human ES cell derived source of mDA neurons that could be used for transplantation in PD patients. Pitx3 is a bicoid-related homeodomain protein with an expression pattern restricted to the mDA neurons of the SNc and ventral tegmental area (VTA), within the central nervous system. To directly investigate a role for Pitx3 in mDA neuron development, I have analysed a line of transgenic mice with a green fluorescent protein (GFP) reporter under the control of the endogenous Pitx3 promoter. Use of the targeted GFP reporter as a midbrain dopaminergic lineage marker in the phenotypically normal heterozygous mice identified previously unrecognised ontogenetically distinct subpopulations of dopaminergic cells within the ventral midbrain. These subpopulations were detectable at E12.5 based on their temporal and topographical expression of Pitx3 and TH. Analysis of the Pitx3 null mice revealed that Pitx3 is required for the survival of a subset of nascent mDA neurons at the beginning of their terminal differentiation. The loss of mDA neurons via apoptosis continued throughout development resulting in a complete absence of SNc neurons whilst the VTA remained relatively intact in adult Pitx3 null mice. In addition, during embryonic development Pitx3 deficiency caused a loss of tyrosine hydroxylase (TH) expression specifically in the SNc dopaminergic neurons. Analysis of chimeric mice made with Pitx3 null and Pitx3 heterozygous ES cells revealed that Pitx3 acts in a cell autonomous manner. These findings point to two roles for Pitx3 in SNc mDA neurons, one in their survival and the other in regulation of TH expression. Taken together, these studies suggest that the ontogenetically distinct subpopulations may provide the molecular basis for the specific dependence of substantia nigra DA neurons on Pitx3. In addition, to establish whether the subpopulations identified at E12.5 do form the SNc and VTA, respectively, a strategy to track the fate of the earliest Pitx3- expressing cells has been initiated. In order to achieve this I have created transgenic mice in which a tamoxifen inducible form of Cre recombinase is under the control of the endogenous Pitx3 promoter. These mice can be crossed with existing mice which contain a ubiquitously expressed Cre-inducible reporter, such as LacZ or GFP, to give a temporally and spatially restricted reporter expression.

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Kragesteen, Bjørt Katrinardóttir [Verfasser]. "A Dynamic Chromatin Architecture Modulates Pitx1 Gene Regulation in Limb Development and Disease / Bjørt Katrinardóttir Kragesteen". Berlin : Freie Universität Berlin, 2018. http://d-nb.info/1152264303/34.

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33

O'Keeffe, Fiona. "The effects of Pitx3 and GDF-5 on the generation and survival of midbrain dopaminergic neurons". Thesis, University of Cambridge, 2012. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.610732.

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34

Christiaen, Lionel. "Origine ontogénétique et évolutive de l'hypophyse : étude cellulaire et moléculaire du développement du complexe neural de l'ascidie Ciona intestinalis". Paris 11, 2004. http://www.theses.fr/2004PA112117.

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J'ai initié une étude comparée du développement hypophysaire chez les Chordés, par l'analyse des aspects cellulaires et moléculaires du développement précoce du complexe neural chez l'ascidie, Ciona intestinalis. La spécification du champ hypophysaire présomptif est intimement liée au développement du stomodæum (ectoderme oral), dont dérive la placode adénohypophysaire. Les gènes de la famille pitx présentent une expression pan-stomodæale et pan-hypophysaire conservée chez les les Vertébrés et les Céphalochordés. Mes résultats montrent que l'expression pan-stomodæale de pitx est aussi conservée chez l'ascidie. Ces observations suggèrent que l'hypophyse des Vertébrés et le complexe neural des ascidies se développent à partir d'un primordium ancestral et conservé. L'analyse d'autres marqueurs hypophysaires et hypothalamiques suggère une diversification des étapes successives de l'organogenèse. L'étude des mécanismes d'activation de Ci-pitx a été entreprise par l'analyse de son système cis-régulateur. Mes observations montrent que la structure génomique et l'usage transcriptionnel des exons de pitx sont majoritairement conservés entre les ascidies et le Vertébrés. L'expression de Ci-pitx dans l'ANB et le stomodæum larvaire est contrôlée par des modules cis-régulateurs distincts dont les patrons spatio-temporels d'activité transcriptionnelle sont complémentaires. L'analyse détaillée du module D1, responsable de l'activation précoce dans la placode stomodæale (i. E. L'ANB), suggère que le code cis-régulateur intégrant une combinaison de signaux acivateurs et répresseurs puisse être conservé chez les chordés
I initiated a comparative study of pituitary development in chordates, by a molecular and cellular analysis of the neural complex development in the ascidian Ciona intestinalis. Pituitary primordium specification is intimately linked to early stomodaeal development (oral ectoderm), from which the adenohypophyseal placode arises. The pituitary homeobox (pitx) gene family constitutes conserved markers of the stomodaeal ectomere in vertebrates and cephalochordates. My data show that pitx genes also display pan-stomodaeal expression in ascidians, suggesting ancestry and homology of the pituitary and neural complex primordia. The analysis of additional pituitary and hypothalamic markers suggest that diversification occurred at later stages of organogenesis. The Ci-pitx cis-regulatory system was studied to investigate the developmental mechanisms that drive pitx expression. My observations show that the overall genomic structure and exon usage of pitx genes are essentially conserved between ascidians and vertebrates. Ci-pitx expression in the anterior neural boundary (ANB) and stomodaeum is driven by distinct cis-regulatory modules, the transcriptional activity of which display complementary spatio-temporal patterns. Detailed analysis of the main ANB/stomodaeal enhancer suggests that the cis-regulatory code that defines its activity is conserved among chordates

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Island, Marie-Laure. "Etude de la répression transcriptionnelle des gènes des interferons-A murins : rôles du facteur à homéodomaine Pitx1". Paris 5, 2001. http://www.theses.fr/2001PA05S015.

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L'étude de la régulation transcriptionnelle des différents membres de la famille multigénique des interférons-A (IFN-A) constitue un bon modèle pour comprendre les mécanismes d'activation et de répression d'un gène eucaryote. Certains de ces gènes sont hautement inductibles par les virus alors que d'autres ne le sont pas. Dans les cellules L929 murines après infection par le virus NDV, l'INF-A4 est hautement inductible tandis que l'IFN-A11 est faiblement inductible. L'expression différentielle des gènes des IFN-A murins s'explique par des substitutions dans les éléments proximaux activateurs de réponse aux virus (VRE), mais également par la présence dans la partie distale du promoteur IFN-A11 d'un domaine de régulation négative. Le même élément est présent dans le promoteur de l'IFN-A4 hautement inductible cependant, l'effet silenceur du DNRE est rendu inopérant par la présence d'un domaine antisilenceur central situé entre l'élément distal DNRE et l'élément proximal activateur VRE. L'élément DNRE est capable de fixer les protéines High Mobility Group I (Y) qui ne semblent pas être impliquées dans l'effet répresseur et une protéine qui pourrait être responsable de l'activité silenceur. Le clonage d'expression "in vivo" chez "Saccharomyces cerevisiae" (clonage simple-hybride) a permis d'isoler une protéine à homéodomaine : Pitx1 (Pituitary homeo box 1). Nous avons montré que Pitx1 joue un rôle important sur la répression transcriptionnelle des gènes IFN-A et qu'elle est impliquée dans l'expression différentielle de ces gènes. Nous avons recherché le mécanisme de répression de ce facteur par l'étude de sa structure et de sa fonction dans le cadre de la régulation transcriptionnelle des gènes IFN-A après induction virale. Cette étude a permis de mettre en évidence de nouveaux domaines fonctionnels de la protéine Pitx1 impliqués dans la trans-répression et dans l'interaction avec de nouveaux partenaires de ce facteur présents sur les éléments VRE-A
The regulation of the expression of type I interferon (IFN-A and IFN-B) multigene family is a model for understanding the positive and negative transcriptional mechanisms of eukaryotic gene regulation. The interferon (IFN)-A4 gene is highly inducible upon virus infection, whereas the IFN-A11 is not. In mouse L929 cells, the weak response of this promoter to viral induction is due to a combination of both point mutations in the virus responsive element (VRE) and the presence of negatively regulating sequences surrounding the VRE that can negatively modulate the transcriptional response to viral induction. This minimal 20-mer distal negative regularoty element (DNRE) in both promoters is necessarey and sufficient for the silencing and a region in the highly inducible IFN-A4 promoter located between the silencer and the virus responsive element overrides the silencer activity. (. . . )

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36

Mechakra, Asma. "Étude cellulaire et moléculaire de quelques aspects de la fibrillation atriale et du syndrome du QT long : rôle des connexines 40 et 43, du facteur de transcription PITX2c et du canal potassique codé par KCNH2". Thesis, Lyon 1, 2014. http://www.theses.fr/2014LYO10005.

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La fibrillation atriale (FA) est l'arythmie cardiaque soutenue la plus fréquente chez les adultes. Elle est associée à une augmentation du risque d'accident vasculaire cérébral, d'insuffisance cardiaque et de la mortalité. Un mécanisme de torsades auriculaires a été décrit chez des patients atteints du syndrome du QT long congénital (SQTL). Malgré une littérature très riche sur le sujet, les mécanismes impliqués dans la genèse et le maintien de ces arythmies restent mal connus et constituent un obstacle dans le diagnostic et la prise en charge de ces maladies. Dans la première partie de ce travail, nous avons abordé l'histologie et la distribution des connexines (Cxs) chez deux groupes de patients avec et sans FA par une approche de microscopie confocale. Nous avons ainsi décrit d'une part un réseau entre fibroblastes et myocytes communiquant via les Cx 40 et 43 et d'autre part la présence de myofibroblastes, d'une forte fibrose et d'un remodelage des Cx 40 et 43 dans le tissu de patients FA. Par ailleurs, pour identifier de nouvelles mutations impliquées dans ces arythmies, nous avons étudié une cohorte de 60 patients atteints de FA. Les recherches génétiques et l'étude fonctionnelle nous ont permis d'associer 5 nouvelles mutations: P41S et M207V (PITX2), G277E (Cx 40) A253V (Cx 43) et P1034H (KCNH2) à la FA. Celles-ci semblent jouer un rôle clé dans la constitution du substrat arythmogène. Enfin, dans la dernière partie, nous avons exploré l'impact électrophysiologique d'un variant de KCNH2, R148W, trouvé tout d'abord chez un enfant décédé de mort subite pendant le sommeil, puis chez plusieurs membres de la famille, dont certains présentent un intervalle QT allongé. Ce variant, exprimé dans les ovocytes de Xénope et étudié en voltage-clamp, réduit le courant de 29% et pourrait alors prédisposer à la survenue de torsades de pointes et expliquer en partie l'allongement du QTc. Outre les nouveaux variants géniques découverts, ce travail est le premier à associer un gain de fonction du facteur de croissance PITX2c en relation avec la FA. Le remodelage histologique des Cx et les variants nucléotidiques touchant les gènes GJA1, GJA5, PITX2 et KCNH2 pourraient ainsi participer à l'étiologie de la FA et du QT long
Atrial fibrillation (AF) is the most common sustained arrhythmia in adults. It is associated with an increased risk of stroke, heart failure and mortality. A mechanism of atrial torsades has been described in patients with congenital long QT syndrome (LQTS). Despite the already existing body of literature, the mechanisms involved in the genesis and maintenance of these arrhythmias remain poorly understood and constitute an obstacle in diagnosis and management of these diseases. In the first part of this work, we discussed the histology of connexins (Cxs) and their distribution in two groups of patients (with and without FA), by confocal microscopy approach. We have described a network of fibroblasts and myocytes communicating across Cx 40 and 43 and the presence of myofibroblasts, of a strong fibrosis and of a remodeling of Cx 40 and 43 distribution in the tissue of AF patients. In addition, to identify new mutations involved in these arrhythmias, we studied a cohort of 60 patients with AF. Genetic investigations and functional study enabled us to associate five novel mutations with AF: M207V and P41S (PITX2), G277E (Cx 40) A253V (Cx 43) and P1034H (KCNH2). These mutations likely play a key role in the formation of the arrhythmogenic substrate. Finally, we explored the electrophysiological impact of a KCNH2 variant, R148W, initially found in a child who died suddenly during sleep and subsequently disclosed in several family members, some with a long QT interval. When expressed in Xenopus oocytes and studied in voltage-clamp, this variant reduces the current by 29%, which might predispose to torsades de pointes and partly explain the QTc prolongation. In addition to these newly discovered gene variants, this work is the first to report a gain-of-function mutation of the transcription factor PITX2c in AF. Histological remodeling of Cxs and the nucleotide variants affecting GJA1, GJA5, PITX2 and KCNH2 might thus participate in the etiology of AF and LQTS

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Ho, Hsin-Yi. "Pitx3 : its role in lens development and application as a midbrain dopaminergic neuron reporter in embryonic stem cell differentiation". Thesis, University of Edinburgh, 2007. http://hdl.handle.net/1842/1483.

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The homeobox gene Pitx3 has been implicated as a key regulator for lens development because homozygous mutant aphakia mice, which are hypomorph for Pitx3, fail to develop lenses. One aim of my thesis is to investigate the underlying cellular and molecular mechanism of Pitx3 mediated lens defect by studying knockout mice lacking Pitx3. Chimeric embryos, generated by aggregating the wild type embryos with Pitx3 heterozygous or Pitx3 homozygous mutant ES cells, have been used to analyse lens development. Pitx3 null cells failed to colonise the lens epithelium in Pitx3 null wild type chimeric lens, suggesting that Pitx3 is cell-autonomously required for lens epithelial cells. Further study of Pitx3 null mice revealed an earlier downregulation of the lens epithelial markers PDGFR-alpha and E-cadherin in E11.5 lens epithelium, suggesting the loss of lens epithelial identity in Pitx3 deficient mice. Furthermore, cell cycle inhibitors p27KIP1 and p57KIP2 were ectopically expressed throughout the morphologically normal Pitx3 mutant lens vesicle, suggesting that inactivation of Pitx3 leads to cell cycle exit of epithelial lens cells. In addition, precocious activation of the fibre cell-specific proteins beta- and gamma-crystallins was observed in Pitx3 null lens. Beta-crystallin expression could be observed as early as E10.5 throughout the entire Pitx3 null lens vesicle and gamma-crystallin was detected in the malformed Pitx3 deficient lens at E11.5. RNA in situ hybridisation study revealed that the expression of the transcription factor Foxe3 was lost in Pitx3 null lens at E10.5, suggesting that Pitx3 maintains the lens epithelial cells partly via the regulation of transcription factor Foxe3 during lens development. Accordingly, this study provides the cellular and molecular basis for the lens defect observed in Pitx3 null and Pitx3 hypomorph aphakia mice. Pitx3 is a key transcription factor for the maintenance of lens epithelium and its absence leads to premature activation of fibre cell differentiation programme of lens epithelial cells. In the other part of my PhD, I have further developed the Pitx3-GFP knockin ES cell system with a goal to use this tool for the identification of determinants of midbrain dopaminergic (mDA) neurons, the type of cells lost in Parkinson’s disease (PD) patients. Experimental cell therapy and clinical trials have shown that foetal midbrain tissues, but not tissues from other DA neuron containing regions, can functionally restore the lost mDA neurons when transplanted in Parkinson’s disease patients. Therefore, it is essential to coax mDA properties on stem cell-derived neurons when considering therapeutic development. Within the central nervous system, Pitx3 is expressed exclusively in mDA neurons. Using a Pitx3-GFP knockin mouse line previously generated in the laboratory I have derived heterozygous and homozygous Pitx3-GFP ES cells from mouse blastocysts. In keeping with previous findings in our laboratory, the heterozygous Pitx3-GFP (Pitx3GFP/+) ES cell-derived GFP positive cells of neuronal morphology can be detected after in vitro differentiation using the PA6 coculture system. Furthermore, I have shown that these cells express tyrosine hydroxylase and midbrain markers Engrailed-1 and Nurr-1, demonstrating their midbrain characteristics. I have also generated supertransfectable Pitx3GFP/+ ES cells to offer a rapid and efficient way to express a transgene episomally. The Cre-mediated inducible system of Pitx3-GFP reporter ES cells has also been developed in our laboratory and I have shown that they have high induction efficiency thus allows transgene activation in a temporally controlled manner. The Pitx3 null ES cells showed impaired potential to differentiate into mDA neurons thus they may be used to evaluate candidate Pitx3 downstream target by gain-of-function test. In summary, I have developed a Pitx3-GFP reporter ES cell system to identify mDA regulators functionally by in vitro differentiation.

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Santos, Tatiana Nayara Libório dos. "Transcritos do gene PITX1 em carcinomas epidermóides de boca: amplificação por RT-PCR e localização por hibridização in situ". Universidade de São Paulo, 2004. http://www.teses.usp.br/teses/disponiveis/23/23141/tde-02022005-123358/.

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Os genes homeobox atuam na morfogênese e na diferenciação celular e vêm sendo relacionados a cânceres em humanos. O projeto Genoma Câncer de Cabeça e Pescoço encontrou aproximadamente 20 desses genes possivelmente envolvidos com esse tipo de neoplasia e o PITX1 foi um dos genes encontrados. Sabe-se que o gene PITX1 está relacionado ao desenvolvimento das estruturas anteriores do embrião, porém sua participação em neoplasias não está bem estabelecida até o momento. Nesse estudo nos propusemos a verificar a presença de transcritos do gene PITX1 em carcinomas epidermóides de boca e tecidos não tumorais adjacentes, analisando sua possível relação com a morfologia das células neoplásicas. Para tanto, os transcritos do gene foram amplificados por RT-PCR e sua localização celular determinada por hibridização in situ com sondas de mRNA específicas. Os resultados mostraram que o transcrito sofreu amplificação em 86,2% dos casos, sendo que 28% foram somente nos tumores, 16% somente nos tecidos não tumorais e 56% em ambos os tecidos. A análise estatística através do Z-test (teste de diferença de proporção entre duas populações) mostrou que não houve diferença significativa entre a amplificação e o tipo de tecido analisado. Reações de hibridização in situ mostraram marcação predominante no componente epitelial de maneira heterogênea ora intensa ora branda em populações celulares diversas tanto nos tecidos tumorais quantos nos tecidos não tumorais adjacentes à lesão, sem relação aparente com a morfologia celular. De maneira geral, o sinal foi mais intenso no epitélio do tecido não tumoral quando comparado ao neoplásico. Frente aos resultados obtidos sugere-se que o gene PITX1 pode estar envolvido na carcinogênese de boca, sendo que a sua expressão está reduzida na maioria das células do carcinoma epidermóide de boca.
Homeobox genes have functions on morphogenesis and cell differentiation and have been related with cancer in humans. PITX1 was one of the genes found in the Head and Neck Cancer Genome Project. It is known that PITX1 gene is related with anterior structures of the developing embryo, although its relation with neoplasm is not we ll established. In this study we proposed to verify the presence of PITX1 gene transcripts in oral squamous cell carcinoma and adjacent non-tumoral tissues, analyzing its possible relation with the morphology of neoplastic cells. For such study, gene transcripts were amplified by RT -PCR and its cellular localization determined by in situ hybridization with specific riboprobes. Results showed that the transcript was amplified in 86,2% of the cases; 28% were only in the tumors, 16% non-tumoral tissues and 56% were amplified in both tissues. Statistics analysis by Z-test showed there was no significant difference between amplification and the type of tissue analyzed. In situ hybridization reactions showed transcripts mostly expressed in the epithelium component in a heterogenic manner sometimes intense and others discrete in several cells populations in both tumoral and non-tumoral adjacent tissues, with no apparent relationship to cell morphology. In general, signal was more intense in the non-tumoral epithelium when compared to the neoplasia. These results suggest that PITX1 gene can be involved with oral carcinogenesis and that its expression is reduced in most of the oral squamous cell carcinoma.

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Peng, Changgeng [Verfasser], Angelika [Akademischer Betreuer] Schnieke e Wolfgang [Akademischer Betreuer] Wurst. "Function of Pitx3 and miRNAs in midbrain dopaminergic neurons / Changgeng Peng. Gutachter: Wolfgang Wurst ; Angelika Schnieke. Betreuer: Wolfgang Wurst". München : Universitätsbibliothek der TU München, 2012. http://d-nb.info/1031075453/34.

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40

Jaszczyszyn, Yan. "Etude des séquences cis-régulatrices de gènes du système nerveux antérieur et de la placode antérieure chez l'ascidie Ciona intestinalis". Paris 11, 2009. http://www.theses.fr/2009PA112347.

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La biologie comparée du développement a mis en évidence la conservation de l'expression de certains gènes même entre des groupes évolutivement éloignés. Les séquences régulant des expressions similaire sont souvent, elles, trop divergentes pour être comparées facilement. Mon travail a été réalisé principalement chez l'ascidie Ciona intestinalis, un chordé invertébré qui occupe la position de groupe frère des vertébrés. L'analyse de la région cis-régulatrice du gène pitx, exprimé à la frontière antérieure neurale a permis de mettre en évidence un élément régulateur D1. Son analyse a montré qu'il contenait des sites de liaison pour des facteurs de transcription, conservés évolutivement, essentiels à son activité et présents en deux exemplaires. Un de ces sites lie l'homéoprotéine OTX qui joue un rôle majeur dans la formation du système nerveux central (SNC) antérieur. Une analyse bioinformatique basée sur la recherche de sites conservés et dupliqués dans le génome a permis de montrer que le motifliant OTX (GATTA) était surreprésenté dans les éléments conservés flanquant les gènes du système nerveux antérieur. Ces éléments ont été testés par transgénèse pour confirmer que la grammaire de sites GATTA dupliqués permettait de prédire des séquences régulatrices actives dans le SNC antérieur. Nous avons montré que les éléments non-codants conservés contenant la grammaire 2xGATTA présents autour des gènes pitx chez le médaka, un poisson téléostéen, conduisaient l'expression dans les régions antérieures du SNC. Cette logique cis-régulatrice est donc conservée à longue distance évolutive, entre des gènes à l'expression conservés dont les séquences régulatrices sont très différentes
Comparative Developmental Biology has shown that gene expression is conserved among evolutionary distant species. However cis-regulatory regions that drive the expression of those genes are often not conserved. My work was done on an ascidian, Ciona intestinalis, an invertebrate marine animal closely related to vertebrates. An analysis of the promoter region of pitx, a gene expressed at the anterior neural boundary, reveals the presence of a regulatory element named D1. We show that D1 likely contains binding sites for transcription factors that are evolutionarily conserved and essential for its activity. Furthermore we show that these motifs need to be duplicated to be functional. One of these motifs is a binding site for Otx, a transcription factor involved in the formation of the anterior central nervous system (CNS). A bioinformatics analysis was performed on the genome sequence of C. Intestinalis and showed that the OTX binding motifs (GATTA) are overrepresented in conserved elements flanking genes that are specificaIly expressed in the anterior CNS. A number of such conserved elements were tested through transient transgenesis and confirm that the duplicated GATTA sites grammar was sufficient to identify regulatory elements active in the anterior CNS. We also show that conserved non-coding elements containing the 2xGATTA grammar are found around pitx genes in the teleost fish Orizias latipes and drive the expression in anterior regions of the CNS. The cis-regulatory logic, predictive in ascidians for anterior CNS enhancers can also be found in some vertebrates genes

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Poulin, Gino. "Formation d'un complexe transcriptionnel spécifique aux cellules corticotropes de l'hypophyse entre facteurs bHLH (NeuroD1/E47) et Pitx1, un facteur à homéodomaine". Thesis, National Library of Canada = Bibliothèque nationale du Canada, 2001. http://www.collectionscanada.ca/obj/s4/f2/dsk3/ftp05/NQ65327.pdf.

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42

Bon, Nina. "Etude du rôle des protéines PiT1/Slc20a1 et PiT2/Slc20a2 dans la détection du phosphate extracellulaire dans le squelette des mammifères". Thesis, Nantes, 2017. http://www.theses.fr/2017NANT1034/document.

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Le phosphate (Pi) est vital pour de nombreux processus physiologiques. Un déficit prolongé en Pi provoque une hypophosphatémie conduisant à une déminéralisation osseuse, alors qu'une hyperphosphatémie entraîne des calcifications vasculaires morbides. Un contrôle adéquat de la concentration sérique de Pi est ainsi essentiel à la qualité et l'espérance de vie. La première étape de ce contrôle est la capacité de la cellule ou de l'organisme à détecter les changements des concentrations extracellulaires de Pi. Contrairement aux levures et aux bactéries, chez lesquelles les transporteurs de Pi membranaires jouent un rôle clé dans cette détection, le mécanisme reste inconnu chez les mammifères. Dans ce travail, nous avons émis l’hypothèse que les cotransporteurs Na-Pi de haute affinité, PiT1/Slc20a1 et PiT2/Slc20a2, étaient impliqués dans ce mécanisme. Dans un premier temps, nous avons montré que la délétion de PiT1 ou de PiT2 dans des ostéoblastes et des chondrocytes in vitro empêchait l’activation de la voie MAPK/ERK1-2 par le Pi. En utilisant une approche de BRET, nous avons ensuite montré que PiT1 et PiT2 pouvaient former des hétéro-dimères modulés par les variations de Pi extracellulaire, sans lien avec le transport de Pi au travers de la membrane. En utilisant des souris invalidées pour le gène PiT2, nous avons montré que PiT2 était indispensable à la régulation par le Pi de la sécrétion de FGF23, l’hormone principalement responsable du maintien de l’homéostasie du Pi. En conclusion, nos données représentent un argument fort en faveur d’un rôle clé des protéines PiTs dans la détection du Pi chez les mammifères, indépendamment de leur fonction de transport
Phosphate (Pi) is a vital ion involved in numerous biological processes. Prolonged deficiency of Pi results in hypophosphatemia leading to serious consequences, including impaired bone mineralization. On the other hand, hyperphosphatemia can lead to life-threatening situations such as vascular calcifications. Controlling serum Pi concentration is therefore critical to life quality and expectancy. The first necessary step of this control is the ability to detect changes in extracellular Pi levels, which implies the existence of a Pi-sensing mechanism that would inform the body or the individual cell. Contrary to yeasts and bacteria for which Pi membrane transporters play a key role in Pi signaling, the underlying mechanism in mammals remains unknown. In this work, we hypothesized that the high affinity Na-Pi cotransporters PiT1/Slc20a1 and PiT2/Slc20a2 could be involved in Pi-sensing in mammals. As a first step, we showed in vitro that deleting either PiT1 or PiT2 blunted the Pi-dependent activation of MAPK/ERK1-2 both in osteoblasts and chondrocytes. This suggested that both PiTs were necessary to Pi signaling. Using a BRET approach, we then demonstrated that PiT1 and PiT2 could form a hetero-dimeric complex that was modulated by variations of extracellular Pi, but not by Pi transport across the membrane. Using PiT2 knockout mice, we showed that PiT2 was also necessary for the Pi-dependent regulation of FGF23 secretion, the main hormone responsible for Pi homeostasis. Taken together, our data propose that the PiT proteins could play a pivotal role in the Pi-sensing mechanism in mammals, which may be uncoupled from their Pi transport function

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Garric, Laurence. "Rôle de la voie Nodal et du facteur de transcription pitx2c dans la mise en place des asymétries neurales dans l'épithalamus du poisson zèbre". Toulouse 3, 2013. http://thesesups.ups-tlse.fr/2312/.

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Malgré notre apparente symétrie, il existe de nombreuses différences entre nos côtés gauche et droit tant au niveau de nos viscères qu'au niveau de notre cerveau. Ces asymétries neuroanatomiques sous-tendent des asymétries fonctionnelles et comportementales : le langage, par exemple, est un processus principalement réalisé dans l'hémisphère gauche. Des défauts dans la latéralisation (ou orientation droite ou gauche des asymétries) ont été corrélés à des désordres neuropathologiques comme la schizophrénie, l'autisme ou des maladies neurodégénératives. Il semble donc nécessaire d'étudier la mise en place de ces asymétries dans un modèle plus simple pour pouvoir en disséquer les étapes. Le poisson zèbre émerge depuis plusieurs années comme un modèle de choix pour étudier la mise en place de ces asymétries grâce à une structure asymétrique, l'épithalamus, qui appartient au diencéphale. L'épithalamus est constitué des noyaux gauche et droit des habenulae situés de part et d'autre de la ligne médiane et du complexe pinéal, lui même composé de la glande pinéale médiane et de la glande parapinéale. La localisation de la parapinéale pointe une asymétrie flagrante car cette glande est située dans 95% des cas à gauche de l'axe médian. Mais en plus de cette asymétrie neuroanatomique, l'épithalamus comporte une asymétrie moléculaire au niveau des noyaux de l'habenula : l'expression préférentielle dans le noyau droit ou dans le noyau gauche de certains marqueurs comme kctd12. 1/leftover vont définir des sous domaines au sein de l'habenula. Il s'avère qu'une corrélation parfaite entre la latéralité de la parapinéale et la distribution des sous domaines existe grâce à une communication bidirectionnelle entre parapinéale et habenulae. Par ailleurs, il existe également une asymétrie de neurogenèse entre les noyaux habénulaires. En effet, des neurones apparaissent plus précocement au sein du noyau gauche de l'habenulae. La voie de signalisation TGF-ß/Nodal a été décrite comme coordonnant la mise en place des différentes asymétries présentes dans l'épithalamus du poisson zèbre : l'activité de cette voie dans le territoire gauche de l'épithalamus biaise la migration de la parapinéale vers la gauche et impose l'asymétrie de la neurogenèse. La façon dont Nodal régule latéralité et asymétries dans l'épithalamus reste néanmoins inconnue. Le projet dans lequel s'inscrit ce projet de thèse vise à révéler les gènes cibles de Nodal qui jouent un rôle dans la mise en place des différentes asymétries de l'épithalamus. Pitx2c, facteur de transcription appartenant à la famille des gènes à homéodomaines, connu pour être une cible directe de la voie Nodal, est un candidat de choix pour faire le lien entre la voie de signalisation Nodal, neurogenèse asymétrique et latéralité de l'épithalamus. De nombreuses études montrent le rôle essentiel de pitx2c dans la latéralité et l'asymétrie de structures viscérales comme le cœur ou les poumons. De plus, il est exprimé dans le territoire gauche de l'épithalamus. Dans ce contexte, mes travaux de thèse ont pu mettre à jour l'implication de pitx2c dans la mise en place des asymétries de l'épithalamus ; Pitx2c permet d'imposer l'asymétrie de neurogenèse des habenulae. De façon inattendue, pitx2c régule également la taille de la glande parapinéale qui est essentielle à l'établissement correct de l'identité des neurones habénulaires
Although we consider ourselves to be symmetric along the left-right axis, our bodies display obvious asymmetries in structure and position of organs; our heart, for instance, is on the left side. Structural differences can also be found in the brain between the left and right hemispheres. These neuroanatomical asymmetries underlie functional and behavioural asymmetries, an example being language that is processed predominantly by the left hemisphere of our brain. Intriguingly, defects in brain lateralisation (the left or right orientation of asymmetries) have been recently correlated to neuropathological disorders such as schizophrenia, autism or neurodegenerative diseases. It seems, thus, that the correct establishment of neuroanatomical asymmetries is essential for a functional nervous system. In the zebrafish, Danio rerio, the establishment of left-right brain asymmetries is being addressed thanks to an asymmetrical brain structure, the epithalamus, which is located in the dorsal diencephalon. The epithalamus is made up of right and left habenular nuclei and the pineal complex, which contains the epiphysis (or pineal gland) and the parapineal gland. Parapineal location is an obvious neuroanatomical asymmetry because in 95% of cases it is located on the left side. Habenular nuclei also display asymmetries at neuroanatomic, cytoarchitectural and molecular levels which are conserved through vertebrate evolution. In the zebrafish, habenula asymmetries can be detected by differential expression of molecular markers, which define left and right nuclei identity. For instance, the kctd12. 1/leftover gene is preferentially expressed in the left nucleus. An asymmetry in habenular neurogenesis is also present: a pool of neurons appears earlier in the left habenular nuclei. A perfect correlation exists between the laterality (left or right orientation) of the parapineal migration and the distribution of the subdomains markers of habenulae. Studies have showed that this association is established through a stepwise series of interactions between the habenulae and parapineal during development. The TGF-ß/Nodal signaling pathway is a key factor in the establishment of epithalamic asymmetries: its activity in the left epithalamic territory biases the parapineal migration to the left and imposes asymmetry on neurogenesis. How Nodal impose laterality and asymmetries in the epithalamus remains unknown. My PhD project aims to identify Nodal targets genes that are involved in the establishment of asymmetry in the epithalamus. Pitx2c, a transcription factor, known to be a Nodal direct target gene, is a good candidate to bridge the gap between Nodal signaling, asymmetrical neurogenesis and laterality. Moreover, pitx2c has been shown to be involved in the positioning of organs relative to the midline and in asymmetrical organogenesis. In this context, my PhD work revealed the implication of pitx2c in building an asymmetrical epithalamus: pitx2c imposes asymmetric neurogenesis upon the impulsion of Nodal signalling. Unexpectedly, in parallel to its role in habenular neurogenesis, pitx2c also controls parapineal size which is crucial for correct acquisition of habenular neuronal identities

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44

Laplante-Campbell, Marie-Pier. "Caractérisation de modèles murins de la maladie de Parkinson (MPTP, PITX3) et modification des cellules souches hématopoïétiques pour stimuler la production du « Brain-derived neurotrophic factor » (BDNF)". Thesis, Université Laval, 2008. http://www.theses.ulaval.ca/2008/25874/25874.pdf.

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La maladie de Parkinson est une maladie neurodégénérative caractérisée par des dysfonctions locomotrices causées, en grande partie, par la perte de neurones dopaminergiques de la substance noire. Les patients atteints de la maladie de Parkinson présentent un déficit en « brain-derived neurotrophic factors » (BDNF). Cette neurotrophine est nécessaire pour le développement, le maintien et la survie des neurones dopaminergiques. Nous avons utilisé la capacité naturelle des cellules souches hématopoïétiques à infiltrer les régions lésées du cerveau pour libérer ce facteur neurotrophique. Nous avons démontré que la modification des cellules de la moelle osseuse pour favoriser la production du BDNF permet d’améliorer les déficits locomoteurs des souris parkinsoniennes. De plus, la modification des cellules hématopoïétiques permet d’augmenter les niveaux de BDNF dans la substance noire, le cortex et le thalamus. La surproduction de BDNF permet également de stimuler la production de la dopamine au niveau de la substance noire.
Parkinson’s disease is a common neurodegenerative disorder characterized by locomotor dysfunctions. These motor symptoms are due to a severe loss of dopaminergic neurons in the substantia nigra pars compacta. Parkinson’s patients also have a deficit in the expression of the brain-derived neurotrophic factor (BDNF). This neurotrophin plays an important role in the development, survival and neurotransmission of dopaminergic neurons. Considering that hematopoietic stem cells can infiltrate damaged brain regions, we have modified these cells to deliver the neurotrophic factor in Parkinson’s disease mouse models. We have demonstrated that modification of bone marrow cells attenuates the locomotor dysfonctions in Parkinson’s mice. In addition, overproduction of BDNF by hematopoietic cells increases BDNF levels in the substantia nigra, cortex and thalamus. Overproduction of BDNF also stimulates biosynthesis of dopamine in the substantia nigra.

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45

Laplante-Campbell, Marie-Pier, e Marie-Pier Laplante-Campbell. "Caractérisation de modèles murins de la maladie de Parkinson (MPTP, PITX3) et modification des cellules souches hématopoïétiques pour stimuler la production du " Brain-derived neurotrophic factor " (BDNF)". Master's thesis, Université Laval, 2008. http://hdl.handle.net/20.500.11794/20555.

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La maladie de Parkinson est une maladie neurodégénérative caractérisée par des dysfonctions locomotrices causées, en grande partie, par la perte de neurones dopaminergiques de la substance noire. Les patients atteints de la maladie de Parkinson présentent un déficit en « brain-derived neurotrophic factors » (BDNF). Cette neurotrophine est nécessaire pour le développement, le maintien et la survie des neurones dopaminergiques. Nous avons utilisé la capacité naturelle des cellules souches hématopoïétiques à infiltrer les régions lésées du cerveau pour libérer ce facteur neurotrophique. Nous avons démontré que la modification des cellules de la moelle osseuse pour favoriser la production du BDNF permet d’améliorer les déficits locomoteurs des souris parkinsoniennes. De plus, la modification des cellules hématopoïétiques permet d’augmenter les niveaux de BDNF dans la substance noire, le cortex et le thalamus. La surproduction de BDNF permet également de stimuler la production de la dopamine au niveau de la substance noire.
La maladie de Parkinson est une maladie neurodégénérative caractérisée par des dysfonctions locomotrices causées, en grande partie, par la perte de neurones dopaminergiques de la substance noire. Les patients atteints de la maladie de Parkinson présentent un déficit en « brain-derived neurotrophic factors » (BDNF). Cette neurotrophine est nécessaire pour le développement, le maintien et la survie des neurones dopaminergiques. Nous avons utilisé la capacité naturelle des cellules souches hématopoïétiques à infiltrer les régions lésées du cerveau pour libérer ce facteur neurotrophique. Nous avons démontré que la modification des cellules de la moelle osseuse pour favoriser la production du BDNF permet d’améliorer les déficits locomoteurs des souris parkinsoniennes. De plus, la modification des cellules hématopoïétiques permet d’augmenter les niveaux de BDNF dans la substance noire, le cortex et le thalamus. La surproduction de BDNF permet également de stimuler la production de la dopamine au niveau de la substance noire.
Parkinson’s disease is a common neurodegenerative disorder characterized by locomotor dysfunctions. These motor symptoms are due to a severe loss of dopaminergic neurons in the substantia nigra pars compacta. Parkinson’s patients also have a deficit in the expression of the brain-derived neurotrophic factor (BDNF). This neurotrophin plays an important role in the development, survival and neurotransmission of dopaminergic neurons. Considering that hematopoietic stem cells can infiltrate damaged brain regions, we have modified these cells to deliver the neurotrophic factor in Parkinson’s disease mouse models. We have demonstrated that modification of bone marrow cells attenuates the locomotor dysfonctions in Parkinson’s mice. In addition, overproduction of BDNF by hematopoietic cells increases BDNF levels in the substantia nigra, cortex and thalamus. Overproduction of BDNF also stimulates biosynthesis of dopamine in the substantia nigra.
Parkinson’s disease is a common neurodegenerative disorder characterized by locomotor dysfunctions. These motor symptoms are due to a severe loss of dopaminergic neurons in the substantia nigra pars compacta. Parkinson’s patients also have a deficit in the expression of the brain-derived neurotrophic factor (BDNF). This neurotrophin plays an important role in the development, survival and neurotransmission of dopaminergic neurons. Considering that hematopoietic stem cells can infiltrate damaged brain regions, we have modified these cells to deliver the neurotrophic factor in Parkinson’s disease mouse models. We have demonstrated that modification of bone marrow cells attenuates the locomotor dysfonctions in Parkinson’s mice. In addition, overproduction of BDNF by hematopoietic cells increases BDNF levels in the substantia nigra, cortex and thalamus. Overproduction of BDNF also stimulates biosynthesis of dopamine in the substantia nigra.

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46

Lopez, Sanchez Uriel. "Dual functions of the XPR1/SLC53A1 phosphate exporter and other transporters as nutrient transporters and receptors of gammaretrovirus envelope-like glycoproteins". Thesis, Montpellier, 2018. http://www.theses.fr/2018MONTT042.

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Le phosphate inorganique (Pi) est un minéral essentiel de notre organisme, qui intervient dans la composition des acides nucléiques et des phospholipides, dans la minéralisation des os et des dents, dans la production d’énergie, et dans la régulation des voies de signalisation. L’homéostasie du Pi est étroitement régulée par différents transporteurs, et des anomalies du transport de Pi peuvent avoir des conséquences cliniques sévères. Chez l’homme, ils existent 3 transporteurs de Pi distincts de type SLC (solute carrier) avec une distribution tissulaire large et initialement identifiés en tant que récepteurs de rétrovirus : PiT1/SLC20A1, PiT2/SLC20A2, et XPR1/SLC53A1.Des mutations dans PiT2 sont associées à une maladie rare, la calcification cérébrale primaire familiale (PFBC), caractérisée par des dépôts de phosphate de calcium dans les noyaux gris centraux et par l’expression de troubles neuropsychiatriques. Alors que PiT1 ne semble pas être impliqué dans cette maladie, nous avons découvert que des mutations dans XPR1 étaient présentes chez des patients PFBC, renforçant le lien entre la maladie PFBC et les désordres de l’homéostasie du phosphate.Dans ce travail, nous avons cherché à comprendre comment PiT2 et XPR1, deux transporteurs de Pi mais de flux opposés peuvent conduire à une même maladie. Pour cela, nous avons étudié le lien entre PiT2 et XPR1 dans la régulation du Pi ainsi que les domaines de XPR1 impliqués dans le transport. Nous avons d’abord identifié de nouveaux variants PFBC dans PiT2 et XPR1 et confirmé l'effet délétère de ces mutations sur l’import et l’export. Nous avons pu distinguer des mutations qui abolissaient l’expression en surface de XPR1, et donc indirectement l’export de Pi, alors que d’autres avaient un impact fonctionnel direct sur les transporteurs pourtant présents à la membrane plasmique.L’inactivation de XPR1 dans des cellules haploïdes humaines induit une altération profonde de l’export de Pi sans effet notable sur l’import. De manière surprenante, l’inactivation de PiT2 entraine un effet modéré sur l’import, probablement dû à l'activité complémentaire de PiT1, avec une chute de l’export dépendant de XPR1. Cet effet identifie une boucle de régulation que nous avons montrée être essentielle au maintien des niveaux de phosphate et d’ATP. Ces résultats révèlent que le défaut d’export de phosphate par inactivation de PiT2 et XPR1 est susceptible d’être l’étape-clé qui conduit à une maladie commune, la PFBC.Nous nous sommes concentrés sur cette étape d‘export régulée en étudiant le domaine SPX de XPR1 dans lequel la plupart des mutations PFBC ont été retrouvées. Nous avons identifié la tankyrase (TNK) comme interactant cellulaire, et localisé son site d’interaction à la bordure carboxyle de SPX. Nous avons observé que la délétion de SPX entrainait un défaut d’export de Pi, et que la perte d’interaction de TNK à XPR1 par mutagenèse ponctuelle avait le même effet, suggérant que TNK et SPX sont 2 composants essentiels à l’export de phosphate. Enfin, nous avons identifié de nouvelles mutations de XPR1 à l'extrémité C-terminale qui abolissaient l’export de Pi, et montré que la délétion de ce domaine entrainait un défaut d’expression de XPR1 à la membrane plasmique. Nos résultats indiquent donc que les domaines N- et C-terminaux jouent un rôle clé dans l’export, et donc dans l’homéostasie du phosphate, avec le domaine C-terminal jouant plutôt un rôle dans le trafic en surface de XPR1.L’ensemble de ce travail a permis de documenter de nouvelles mutations PFBC dans les gènes PiT2 et XPR1, de démontrer que ces transporteurs étaient impliqués dans l’homéostasie intracellulaire du phosphate, en dévoilant que l’export de phosphate est vraisemblablement l’étape clé de la PFBC, ouvrant ainsi de nouvelles pistes dans la compréhension de cette maladie. Nous avons également identifié des domaines de XPR1 et un partenaire cellulaire, essentiels à l’export de Pi et/ou au trafic membranaire
Phosphate (Pi) is a key mineral that participates directly in the synthesis of nucleic acids and membranes, bone and tooth mineralization, energy production, and signal transduction. Pi homeostasis is tightly regulated by transporter-mediated fluxes that adjust Pi concentration in real time, and defect in Pi transport has been associated with several pathologies. In humans, three Pi transporters, which belong to the solute carrier (SLC) superfamily, are widely expressed: PiT1/SLC20A1, PiT2/SLC20A2, and XPR1/SLC53A1. Interestingly, all three were initially identified as receptors for mammalian gammaretroviruses.Mutations in PiT2/SLC20A2 are responsible for a rare neurodegenerative disorder, the primary familial brain calcification (PFBC), characterized by deposits of calcium Pi in the basal ganglia and other regions of the brain, and associated with diverse neuropsychiatric clinical manifestations. While PiT1/SLC20A1 has not been involved in PFBC, we recently identified mutations in XPR1/SLC53A1 as causative for PFBC, thus linking further the disease with cellular Pi homeostasis dysfunction.In this work, we aimed to understand how defects of opposite Pi transport functions lead to PFBC, investigated the relationship between PiT2 and XPR1 in cellular Pi regulation, and studied XPR1 domains in Pi transport. We first identified several PFBC mutations in PiT2/SLC20A2 and XPR1/SLC53A1, and confirmed their impact on Pi import or export, respectively. Some of the mutations altered transporter cell surface expression, resulting in Pi transport impairment, while others did neither alter cell surface expression, nor retroviral receptor functions, confirming that Pi transport function and viral envelope glycoprotein binding can be structurally distinguished.Using single gene knock-out human haploid cells, we showed that depletion of XPR1/SLC53A1 resulted in a dramatic Pi export alteration, with no detectable effect on Pi import, in agreement with Pi exporter function of XPR1. Interestingly, depletion of PiT2/SLC20A2 had little impact on Pi uptake, most likely due to compensatory function of PiT1/SLC20A1, with, however, a surprising impact on Pi export mediated by XPR1. This effect is reminiscent to a regulation loop that we found to maintain both Pi and ATP constant. This results unveil for the first time that Pi export alteration, and not Pi import, is likely to be the common pathophysiological impact of mutations in both PiT2 and XPR1. This would explain the synonymous pathological effects of two transporters that have opposite transport activity.We further explored this regulated phosphate export by characterizing the SPX N-terminal cytoplasmic domain of XPR1, which harbors most of the PFBC mutations. We identified a cellular tankyrase (TNK) as a binding partner and mapped the TNK-binding site to the carboxyl border of SPX; furthermore, we found that mutations that abolished TNK binding resulted in loss of Pi export. Full deletion of SPX domain maintained cell surface expression but altered export, suggesting that both TNK and SPX are essential components for Pi export. Finally, during this work, we identified mutations in the XPR1 C-terminal domain as responsible for PFBC that also impaired Pi export, and showed that deletion of this domain prevented XPR1 cell surface expression. Our results therefore indicate that N- and C-terminal domains of XPR1 play a key role in phosphate homeostasis, the latter domain appearing to exert a more prominent role in XPR1 membrane trafficking and/or folding

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47

Ahmad, Nafees [Verfasser], Jochen Akademischer Betreuer] Graw e Heinrich H. D. [Akademischer Betreuer] [Meyer. "Functional and regulatory network analysis of Pitx3 in aphakia – a mouse model for microphthalmia and Parkinson’s disease / Nafees Ahmad. Gutachter: Jochen Graw ; Heinrich H. D. Meyer. Betreuer: Jochen Graw". München : Universitätsbibliothek der TU München, 2011. http://d-nb.info/1016035047/34.

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48

Krug, Christian [Verfasser], Ahmed [Akademischer Betreuer] Mansouri, Gregor [Akademischer Betreuer] Bucher, Ralf [Akademischer Betreuer] Heinrich, Reinhard [Akademischer Betreuer] Schuh, Ernst A. [Akademischer Betreuer] Wimmer e Andreas [Akademischer Betreuer] Wodarz. "Funktionelle Analyse des Transkriptionsfaktors Pitx3 während der Entwicklung dopaminerger Neuronen im murinen Mittelhirn / Christian Krug. Gutachter: Ahmed Mansouri ; Gregor Bucher ; Ralf Heinrich ; Reinhard Schuh ; Ernst A. Wimmer ; Andreas Wodarz. Betreuer: Ahmed Mansouri". Göttingen : Niedersächsische Staats- und Universitätsbibliothek Göttingen, 2013. http://d-nb.info/1044768630/34.

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49

Mesplede, Thibault. "Etude de la régulation de la transcription des gènes interférons A murins : étude du rôle des facteurs de transcription Pitx1 et Oct-1 dans la régulation de la transcription des gènes IFN-A murins". Paris 6, 2006. http://www.theses.fr/2006PA066642.

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50

Strungaru, Marcela Hermina. "Investigation of the role of PITX2 in ocular expression pathways and human disease". Phd thesis, 2010. http://hdl.handle.net/10048/1183.

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The overall goal of my work has been to gain a better understanding of Axenfeld-Rieger Syndrome (ARS), a human autosomal dominantly inherited mal-development of the anterior segment of the eye that is associated with glaucoma. By studying rare genetic causes of this complex disease we are gaining insight into the initial steps that ultimately lead to blindness. To achieve the goal of better understanding ARS, my research project had two parts. In the first part, I performed a retrospective clinical study in which I analyzed the glaucoma-related clinical presentation of ARS patients with FOXC1 and PITX2 defects. This study showed a good genotype-phenotype correlation which may be important for the physician in dealing with ARS patients. Patients with FOXC1 mutations had the mildest prognosis in glaucoma development, while patients with PITX2 defects and patients with FOXC1 duplication had a more severe prognosis in glaucoma development than patients with FOXC1 mutations. I tried to determine the best treatment for glaucoma in these patients. Unfortunately, in this study, current medical therapies did not successfully lower intraocular pressure or prevent progression of glaucoma in ARS patients with FOXC1 or PITX2 alterations. This clinical study also provided useful diagnostic criteria to identify the gene responsible for ARS. The second part of the project was to study the gene regulatory pathways of the PITX2 gene, mutations of which cause ARS. PITX2 is a transcription factor that regulates the expression of genes in the eye. The discovery of direct downstream targets of PITX2 is necessary for understanding the genetic mechanisms underlying complex, highly regulated processes such as development and underlying heritable human disorders. To find direct target genes of PITX2, I have used a recently developed method: the hormone receptor (HR)-inducible expression system for transcription factors coupled microarray analysis. The results obtained using this method have involved PITX2 in control of cellular stress. Recent investigations have suggested significant roles for cellular stress in glaucoma pathology. Understanding the control of these key aspects of cell function will have profound implications for understanding and treating the glaucoma that is the most clinically serious consequence of mutations of PITX2.

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