Dissertations / Theses on the topic 'Tropomyosins'
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1
Au, Wing-han. "Brain-derived neurotrophic factor (BDNF)/tropomyosin-related kinase B (TRKB) signaling in ovarian cancer." Click to view the E-thesis via HKUTO, 2007. http://sunzi.lib.hku.hk/HKUTO/record/B39557947.
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Ebrahim, Seham. "Tropomyosins, N-terminal acetylation and their impact on yeast cytoskeletal function : a characterisation of novel tropomyosins from N. crassa and the N-terminal acetyltransferase, Nat3p." Thesis, Queen Mary, University of London, 2009. http://qmro.qmul.ac.uk/xmlui/handle/123456789/531.
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While the fundamental role of tropomyosins (Tms) in the maintenance of the actin cytoskeleton in yeast is established, details of their exact regulatory functions in lower eukaryotes remains to be deciphered. Here, two novel Tms have been identified from the filamentous yeast Neurospora crassa: a 161 residue protein spanning 4 actin monomers (crTm161p), and a 123 residue protein which spans 3 actin monomers (crTm123p). The latter isoform is the shortest naturally occurring Tm known. The isoforms are produced as a result of alternative splicing from a single gene- a phenomenon that has not previously been observed in yeast Tms. Both Tms were cloned, purified and crystallised. They were also characterised biochemically and biophysically, giving some insight into their role in fungal cytoskeleton regulation. The crystals produced provide the potential for future structural studies, as a high resolution structure of a complete Tm is still not available. The N-terminal acetylation of Tms is essential for their function, and is catalysed in Saccaromyces cerevisiae by the N-terminal N-acetyltransferase (NAT) Nat3p. Nat3p was expressed and purified. Its functionality was investigated via acetyl coenzyme A binding assays. The molecular structure of Nat3p was modelled using existing data from structural homologues. The closely related N-terminal NAT, Nat5p, was also expressed, purified and its structure modelled. Nat3p was largely insoluble while Nat5p was soluble and was successfully crystallised. Structural insights from molecular modelling were able to provide some justification for these differences. Finally the in vivo effects of genetic knockouts of the TPM1 and NAT3 genes in yeast were analysed quantitatively. Complementation of the defective knockout phenotypes by over-expression of various Tm and Nat3p constructs was also investigated. Quantifying the overlapping phenotypes of the NAT3Δ and TPM1Δ mutaunts has clarified their distinct impacts upon the cytoskeleton. The ability of the crTms to rescue TPM1Δ phenotypes implies they have roles similar to those of Tpm1p.
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Vlahovich, Nicole. "The role of cytoskeletal tropomyosins in skeletal muscle and muscle disease." Thesis, View thesis, 2007. http://handle.uws.edu.au:8081/1959.7/32176.
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Cells contain an elaborate cytoskeleton which plays a major role in a variety of cellular functions including: maintenance of cell shape and dimension, providing mechanical strength, cell motility, cytokinesis during mitosis and meiosis and intracellular transport. The cell cytoskeleton is made up of three types of protein filaments: the microtubules, the intermediate filaments and the actin cytoskeleton. These components interact with each other to allow the cell to function correctly. When functioning incorrectly, disruptions to many cellular pathway have been observed with mutations in various cytoskeletal proteins causing an assortment of human disease phenotypes. Characterization of these filament systems in different cell types is essential to the understanding of basic cellular processes and disease causation. The studies in this thesis are concerned with examining specific cytoskeletal tropomyosin-defined actin filament systems in skeletal muscle. The diversity of the actin filament system relies, in part, on the family of actin binding proteins, the tropomyosins (Tms). There are in excess of forty Tm isoforms found in mammals which are derived from four genes: α, β, γ and δTm. The role of the musclespecific Tms in striated muscle is well understood, with sarcomeric Tm isoforms functioning as part of the thin filament where it regulates actin-myosin interactions and hence muscle contraction. However, relatively little known about the roles of the many
cytoskeletal Tm isoforms. Cytoskeletal Tms have been shown to compartmentalise to form functionally distinct filaments in a range of cell types including neurons (Bryce et al., 2003), fibroblasts (Percival et al., 2000) and epithelial cells (Dalby-Payne et al., 2003). Recently it has been shown that cytoskeletal Tm, Tm5NM1 defines a cytoskeletal structure in skeletal muscle called the Z-line associated cytoskeleton (Z-LAC) (Kee et al., 2004).The disruption of this structure by over-expression of an exogenous Tm in transgenic mice results in a muscular dystrophy phenotype, indicating that the Z-LAC plays an important role in maintenance of muscle structure (Kee et al., 2004). In this study, specific cytoskeletal Tms are further investigated in the context of skeletal muscle. Here, we examine the expression, localisation and potential function of cytoskeletal Tm isoforms, focussing on Tm4 (derived from the δ- gene) and Tm5NM1 (derived from the γ-gene). By western blotting and immuno-staining mouse skeletal muscle, we show that cytoskeletal Tms are expressed in a range of muscles and define separate populations of filaments. These filaments are found in association with a number of muscle structures including the myotendinous junction, neuromuscular junction, the sarcolemma, the t-tubules and the sarcoplasmic reticulum. Of particular interest, Tm4 and Tm5NM1 define cytoskeletal elements in association with the
saroplasmic reticulum and T-tubules, respectively, with a separation of less than 90 nm between distinct filamentous populations. The segregation of Tm isoforms indicates a role for Tms in the specification of actin filament function at these cellular regions. Examination of muscle during development, regeneration and disease revealed that Tm4 defines a novel cytoskeletal filament system that is orientated perpendicular to the sarcomeric apparatus. Tm4 is up-regulated in both muscular dystrophy and nemaline myopathy and also during induced regeneration and focal repair in mouse muscle. Transition of the Tm4-defined filaments from a predominsnatly longitudinal to a predominantly Z-LAC orientation is observed during the course of muscle regeneration. This study shows that Tm4 is a marker of regeneration and repair, in response to disease, injury and stress in skeletal muscle.
Analysis of Tm5NM1 over-expressing (Tm5/52) and null (9d89) mice revealed that compensation between Tm genes does not occur in skeletal muscle. We found that the levels of cytoskeletal Tms derived from the δ-gene are not altered to compensate for the loss or gain of Tm5NM1 and that the localisation of Tm4 is unchanged in skeletal muscle of these mice. Also, excess Tm5NM1 is sorted correctly, localising to the ZLAC. This data correlates with evidence from previous investigations which indicates that Tm isoforms are not redundant and are functionally distinct (Gunning et al., 2005). Transgenic and null mice have also allowed the further elucidation of cytoskeletal Tm function in skeletal muscle. Analyses of these mice suggest a role for Tm5NM1 in glucose regulation in both skeletal muscle and adipose tissue. Tm5NM1 is found to colocalise with members of the glucose transport p fibres and analysis of both transgenic and null mice has shown an alteration to glucose uptake in adipose tissue. Taken together these data indicate that Tm5NM1 may play a role in the translocation of the glucose transport molecule GLUT4. In addition to this Tm5NM1 may play a role in adipose tissue regulation, since over-expressing mice found to have increased white adipose tissue and an up-regulation of a transcriptional regulator of fat-cell formation,
PPAR-γ.
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Vlahovich, Nicole. "The role of cytoskeletal tropomyosins in skeletal muscle and muscle disease." View thesis, 2007. http://handle.uws.edu.au:8081/1959.7/32176.
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Thesis (Ph.D.)--University of Western Sydney, 2007.A thesis presented to the University of Western Sydney, College of Health and Science, School of Natural Sciences, in fulfilment of the requirements for the degree of Doctor of Philosophy. Includes bibliographies.
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歐穎嫻 and Wing-han Au. "Brain-derived neurotrophic factor (BDNF)/tropomyosin-related kinaseB (TRKB) signaling in ovarian cancer." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2007. http://hub.hku.hk/bib/B39557947.
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Robinson, Paul John Robert. "The functional effect of disease causing mutations on thin filament regulatory proteins tropomyosin, troponin T troponin I and troponin C." Thesis, University of Oxford, 2007. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.670117.
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Patel, Dipesh A. Root Douglas. "Luminescence resonance energy transfer-based modeling of troponin in the presence of myosin and troponin/tropomyosin defining myosin binding target zones in the reconstituted thin filament." [Denton, Tex.] : University of North Texas, 2009. http://digital.library.unt.edu/permalink/meta-dc-9834.
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Kotadiya, Preeyal. "Regulation Of Osteoclast Function By Alpha Gene Tropomyosins, TM-2/3 And TM-5a/5b." The Ohio State University, 2009. http://rave.ohiolink.edu/etdc/view?acc_num=osu1250612152.
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McMichael, Brooke Kristin Trinrud. "Tropomyosin 4, myosin IIA, and myosin X enhance osteoclast function through regulation of cellular attachment structures." Columbus, Ohio : Ohio State University, 2008. http://rave.ohiolink.edu/etdc/view.cgi?acc%5Fnum=osu1206052974.
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McKay, Janet A. "A feasibility and exploratory study of cardiac rehabilitation in acute coronary syndrome." Thesis, University of Stirling, 2013. http://hdl.handle.net/1893/20346.
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Background: Cardiac Rehabilitation (CR) has been shown to be effective in reducing mortality and morbidity in Coronary Heart Disease (CHD). There is a limited amount of research that evaluates the impact of menu-based CR, in patients with Acute Coronary Syndrome with Low Troponin levels (ACSLT). Aim: This thesis contains a feasibility study and an exploratory study. The feasibility study aimed to examine the feasibility of a Randomised Controlled Trial (RCT) which would test the impact of a menu-based CR programme, on individuals diagnosed with ACSLT, against standard care. This feasibility study included staff views. The exploratory study aimed to explore the impact that ACSLT and CR can have on this client group. Method: The feasibility study was a repeated measures case-control trial of menu-based CR based on the theoretical framework of the Common Sense Model of Self-Regulation (CSM), using a range of health assessments. The areas assessed included misconceptions, symptoms, anxiety, depression and Health Related Quality of Life (HRQoL). In addition, focus groups were held with both ward and specialist CR staff to seek their views on the feasibility of a RCT of menu-based CR for ACSLT. The exploratory study consisted of description and analysis of the data that had been collected from the participants over the two year period as above. In addition it included qualitative data that had been collected during interviews with the participants. Findings: Participants (n=33) were recruited from cardiology wards following an admission with ACSLT. They were assessed at baseline (T1), nine months (T3) and 24 months (T4). Twenty-five participants completed the studies. The feasibility study was successful in its aim of testing the CR intervention and protocols for a further RCT. The intervention was acceptable to the participants and to the specialist staff, although the ward staff did not see the need for a RCT. The measures used, with the exception of the self-reporting measures, were suitable and provided a wide range of data that could be utilised in a RCT. However the changes to diagnostic categories meant that a RCT would no longer be feasible. The exploratory study found that both groups were similar on a range of baseline demographic and clinical factors. There was a tendency to benefit within the exploratory study which favoured the intervention. An additional finding from the exploratory study was the degree of uncertainty experienced by the participants, within the context of a changing political and clinical landscape. Discussion and conclusions: The studies presented in this thesis add to our knowledge by highlighting some of the difficulties in designing a RCT of menu-based CR in a specific subgroup of CHD and by presenting outcome data for a small group of participants that have not previously been studied within the literature. This data suggests that there was a tendency to benefit for the intervention that requires further study. Implications for practice: Patients with ACSLT are now being included in CR programmes due to the changes within the diagnostic criteria. Clinicians have little understanding of the impact of CR on this group of patients, or what type of interventions would work best. Large RCT’s will however be problematic and this thesis has highlighted that further work is required to explore how CR can best improve the well-being of individuals with ACSLT.
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McConnell, Mark, and Mark McConnell. "Investigating the Effects of Tropomyosin D230N and cTnT R92L on the Tropomyosin Overlap Region." Diss., The University of Arizona, 2017. http://hdl.handle.net/10150/624576.
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The progression of genetically inherited cardiomyopathies from an altered protein structure to the clinical presentation of the disease is not well understood. One of the main roadblocks to mechanistic insight remains a lack of high-resolution structural information of multiprotein complexes within the cardiac sarcomere. One example is the tropomyosin (Tm) overlap region of the thin filament that is crucial for the function of the cardiac sarcomere. To address this central question, we devised coupled experimental and computational methods to characterize the baseline function and structure of the Tm overlap, as well as the effects that mutations causing divergent patterns of ventricular remodeling have on both structure and function.
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Clark, Ian David. "Coupled structural responses in tropomyosin." Thesis, University of British Columbia, 1990. http://hdl.handle.net/2429/30625.
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Fluorescence spectroscopy can be used to probe protein conformation and is recognized as a technique that provides very specific information. It has been applied/ in recent years/ to the study of tropomyosin (TM) and its role in regulation of contractile processes. In this thesis, two different approaches were used to further the understanding of the structure/function relationship in the two chain coiled coil of tropomyosin. The first involves a comparative study on TM and non-polymerizable TM (NPTM) (Mak, A.S., and Smillie, L.B. (1981) Biochim. Biophys. Res. Commun., 101, 208-214). Fluorescence involving pyrene (Py) and acrylodan (AD) bound at the only cysteine residue in the molecule (Cys-190), and circular dichroism (CD) studies led to the main conclusion that, while the two species, are very similar in stability, the COOH-terminus is required to hold the Cys-190 region in a specific conformation. This long-range structural effect may play a role in regulation of contraction.
A species having one intact COOH-terminus, made by hybridizing TM and NPTM, was found to be non-polymerizable suggesting that one intact COOH-terminus is insufficient to permit overlap with the NH₂-terminus of a neighbouring TM under polymerizing conditions. Unlike the TM/NPTM hybrid, the hybrid of TM and platelet TM (P-TM) was difficult to make due to the sequence mismatches in the terminal regions,
but small quantities could be detected by loss of excimer fluorescence from Py-P-TM on rapid cooling of a heated mixture of Py-P-TM and cardiac TM (C-TM).
The second approach was to investigate the effect of actin-binding proteins on the structure and function of tropomyosin. DNase I depolymerizes F-actin and is known to interfere with the end-to-end polymerizability of tropomyosin (Payne, M.R., Baydoyannis, H., and Rudnick, S.E. (1986) Biochim. Biophys. Acta 883, 454-459). Results presented here from fluorescence studies suggest that this effect is caused by a localized loss of structure in the tropomyosin at the sites of labelling upon binding of DNase I. This result is supported by CD studies on labelled and unlabelled tropomyosins.
Gelsolin is another actin-binding protein found in many cell types and in extracellular fluids. It is shown here to be able to depolymerize tropomyosin, but its mechanism of action is not the same as that of DNase I. The effect of interaction of gelsolin on the structure of tropomyosin, as determined from fluorescence studies, is negligible.Science, Faculty of
Chemistry, Department of
Graduate
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Had, Laurence. "Tropomyosines et développement du système nerveux." Montpellier 2, 1994. http://www.theses.fr/1994MON20051.
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Dans le systeme nerveux du rat, de nombreuses isoformes de tropomyosine, proteine qui stabilise les microfilaments d'actine, ont ete caracterisees au niveau de leur messager. Nous avons montre, par northern blot d'arn extraits de cultures pures, la presence d'un equipement en tropomyosine particulier dans chaque type de cellule du systeme nerveux, neurone (tm-4, tmbr-3), astrocyte (tm-4, tm-1, tm-2, tm-5a, tmbr-1 et tmbr-2) et oligodendrocyte (tm-4, tmbr-2). L'expression de ces isoformes est regulee de facon differentielle pendant la maturation. Dans les neurones, in vitro comme in vivo, tm-4 est surtout exprimee au cours des stades immatures, tmbr-3 est caracteristique des neurones differencies. Le dibutyryl-ampc et la cytochalasine d, qui induisent sur les astrocytes en culture une reorganisation du cytosquelette d'actine et un changement de la morphologie vers une forme etoilee, modifient fortement la concentration du messager de l'actine-beta et des 6 messagers de tropomyosine. Le dibutyryl-ampc reprime et la cytochalasine d stimule l'expression de l'ensemble de ces messagers. La coexpression des tropomyosines et de l'actine-beta semble indiquer une regulation parallele ou sequentielle de l'expression des proteines constitutives des microfilaments. Un antipeptide dirige contre tm-4 montre dans les neurones en culture un marquage des cones de croissance et dans le systeme nerveux du jeune rat un marquage transitoire des neurites. Chez l'animal mature, les terminaisons post-synaptiques sont marquees. Un antipeptide anti-tmbr-3 marque, au contraire, les terminaisons pre-synaptiques. Tm-4 interviendrait dans la poussee des prolongements neuronaux et, peut etre, dans les phenomenes de plasticite chez l'adulte. Tmbr-3 pourrait jouer un role dans le maintien des neurones dans leur forme mature et dans la liberation des neurotransmetteurs
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Kalyva, Athanasia. "Tropomyosin heterodimers in cardiac muscle regulation." Thesis, University of Kent, 2009. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.508567.
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Boussouf, Sabrina Eida. "Regulation of cardiac muscle contraction : effect of tropomyosin isoform expression and cardiomyopathy mutations in tropomyosin and troponin." Thesis, University of Kent, 2004. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.408903.
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Sheikh, Hajer Nisar. "Tropomyosin Phosphorylation in Cardiac Health and Disease." University of Cincinnati / OhioLINK, 2009. http://rave.ohiolink.edu/etdc/view?acc_num=ucin1242913472.
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Kreuz, Andrew Joseph. "Characterization of tropomyosin mutants in Drosophila melanogaster /." The Ohio State University, 1993. http://rave.ohiolink.edu/etdc/view?acc_num=osu1487843688958564.
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Lendner, Matthias. "Functional analysis of tropomyosin of parasitic nematodes." Doctoral thesis, Humboldt-Universität zu Berlin, Mathematisch-Naturwissenschaftliche Fakultät I, 2010. http://dx.doi.org/10.18452/16137.
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Parasitische Würmer gehören mit über 3,5 Milliarden Betroffenen zu den weltweit verbreitetesten Infektionskrankheiten. Der Erfolg dieser Parasiten beruht auf ihren ausgefeilten Mechanismen mit denen sie das Immunsystem ihrer Wirte manipulieren. Interessanter Weise gehen Wurminfektionen mit einer geringeren Wahrscheinlichkeit an Allergien zu erkranken einher. Wie genau die Parasiten das Immunsystem manipulieren ist weitgehend unbekannt. Um diese Mechanismen besser studieren zu können, wurde im Rahmen dieser Arbeit versucht RNA interference (RNAi), anhand des Modellmoleküls Tropomyosin zu etablieren. Wie sich am Beispiel des Strongyliden Heligmosomoides polygyrus bakeri zeigte, ist RNAi als Manipulationsmethode für Nematoden nicht oder nur in geringem Maße geeignet. Dies lässt sich auf das Fehlen von Aufnahme- und Verbreitungsmechanismen für Doppelstrang-RNA zurückführen. Desweiteren wurden die Auswirkungen von rekombinantem Tropomyosin der Filarie Acanthocheilonema viteae (rAv-TMY) auf die Entstehung allergischer Atemwegserkrankungen im Mausmodell untersucht. Eine viermalige Behandlung mit rAv-TMY in einem Zeitraum von vier Wochen führte zu verringerten entzündlichen Reaktionen in den Atemwegen. Die Analyse immunologischer Parameter ergab, dass rAv-TMY signifikant den Einstrom von Entzündungszellen in die Atemwege reduziert, allem voran den Einstrom von Eosinophilen. Dies lässt sich durch die verringerte Ausschüttung an IL-5, Eotaxin und MCP-5 zurückführen. Zudem wurde die Bildung von antigenspezifischen IgE verringert während sich die Produktion blockierender IgG1 Antikörper erhöhte. Diese Arbeit belegt somit die anti-allergischen Eigenschaften von rAv-TMY. Damit stellt rAv-TMY ein interessantes Kandidatenmolekül zur Behandlung allergischer Reaktionen dar. Desweiteren kann der Vergleich von allergenem, nicht allergenem und modulatorischem Tropomyosin wichtige Informationen über die allgemeinen Eigenschaften von Allergenen und ihrer molekularen Struktur geben.Parasitic worms are among the world''s most prevalent infectious diseases with more than 3.5 billion. The success of these parasites is based on their sophisticated ways to manipulate the immune system of their hosts. Interestingly, worm infections abate the risk to develop allergic disorders. How exactly parasitic worms modulate the immune system is so far largely unknown. In order to be able to investigate parasite induced modulation, this work aimed to establish RNA interference (RNAi), a method of genetic manipulation, using tropomyosin as target gene. As shown for the example of Heligmosomoides polygyrus RNAi is not or only to a small extent useful as method to genetically manipulate nematodes. This can be explained with the lack of uptake and spreading mechanisms for double stranded RNA. Furthermore, this work examined the impact of the recombinant muscle protein tropomyosin of Acanthocheilonema viteae (rAv-TMY) on the course of a rodent model of allergic airway inflammation. A four-time treatment with rAv-TMY over a period of four weeks resulted in decreased inflammatory responses in the airways. The analysis of immunological parameters showed that rAv-TMY significantly reduces the influx of inflammatory cells into the airways, especially eosinophils. The reduced eosinophil influx can be attributed to the decreased expression of IL-5, eotaxin and MCP-5 in the airways. In addition, the formation of antigen-specific IgE was impaired whereas the production of the blocking antibody IgG1 was increased. These results demonstrate the anti-allergic properties of rAv-TMY. For this reason rAv-TMY becomes an interesting model molecule for the treatment of allergic diseases. Furthermore, the comparison of allergenic, non-allergenic and modulatory tropomyosin might put some light on the nature of allergens and their molecular patterns.
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Janco, Miroslav. "Characterisation of tropomyosin heterodimers carrying single cardiomyopathy mutations." Thesis, University of Kent, 2013. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.655655.
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It is known that different point mutations in α-tropomyosin (Tm) can cause either hypertrophic (HCM) or dilated (DCM) cardiomyopathy. Both of these serious pathologies have a distinct phenotype with unknown mechanisms of development. Biochemical in vitro studies provide valuable information for exploring downstream consequences of cardiomyopathy mutations in sarcomeric proteins leading to cardiac remodelling and consequent heart failure. Tm is a linear a-helical coil-coiled dimer involved in calcium dependent regulation of muscle contraction. Prior to this work, the effects of mutations in Tm on regulation of muscle contraction have been made exclusively with Tm mutant homodimers. However, individuals with a heterozygous background may express mutant and WT proteins in a 1 : 1 ratio which can assembly into a mixture of αα, αα* and α*α* Tm dimers. We found that presence of mutation has little effect on dimer formation between the mutant and the WT monomers, therefore theoretical ratio of the Tm dimers in vivo may be 1 : 2 : 1, respectively. This assumption would make the heterodimer predominant. The properties of in vitro assembled Tm heterodimers carrying HCM (WT-D175N and WT-E180G) and DCM (WT-E40K, WT-E54K, and WT-D230N) causing mutations were examined including thermal stability, flexibility, actin affinity, calcium regulation of 51 biding, and calcium regulation of myofibril force. We showed that various properties of the heterodimers can be similar to those of the wild-type (thermal stability of reduced WTD175N; actin affinity of WT-E40K, and WT-D175N; ΔpCa of WT-E40K, WT-E54K, WT-E180G, WT-D230N), similar of those to the mutant homodimer (ΔpCa of WT-D175N, flexibility of WT-D175N and WT-E180G), intermediate between the two (actin affinity of WT-E180G), or different from both (thermal stability of reduced WT-E40K, WT-E54K, WT-E180G, and WTD230N; actin affinity ofWT-E40, WT-E54K, and WT-D230N; ΔpCa of D175N). The results demonstrate that the properties of Tm heterodimers cannot be predicted from the interpolation of the WT and mutant homodimer data. The distinct properties of heterodimers establish that it will be important to define if the pathogenic agent is a homodimer, a heterodimer or both for each known cardiomyopathy mutation in Tm.
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Le, Sommer Caroline. "Identification de facteurs régulant en trans la maturation différentielle de la région 3' terminale de l'ARN pré-messager tropomyosine α chez Xenopus laevis." Rennes 1, 2006. http://www.theses.fr/2006REN1S009.
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Nous utilisons le gène de la tropomyosine de xénope comme modèle moléculaire pour étudier les déterminants de la régulation tissulaire de l'épissage et de la polyadénylation. Ce gène contient dans sa région 3' terminale un exon alternatif, l'exon 9A9', dont l'utilisation, dans l'embryon de xénope, est dépendante de l'environnement tissulaire. Deux séquences, l'une inhibitrice l'autre activatrice, qui régulent l'épissage de cet exon, avait précédemment été identifiées. Afin de caractériser les mécanismes d'action de ces deux séquences, nous avons recherché les facteurs qui contrôlent l'utilisation de l'exon 9A9' via ces dernières. Nous avons identifié la protéine xPTB comme un facteur majeur de la répression de l'exon 9A9' et certains membres de la famille des protéines SR comme des facteurs capables d'activer cet exon. Nous avons par la suite démontré qu'il existe un antagonisme fonctionnel entre la protéine xPTB et la famille des protéines SR dans la régulation de l'exon 9A9'.
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Lynn, Melissa L., and Melissa L. Lynn. "D230N-Tm Induced Dilated Cardiomyopathy and the Role of Fetal cTnT Isoform Switching in Modulating Disease Severity." Diss., The University of Arizona, 2017. http://hdl.handle.net/10150/625579.
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In 1980, the World Health Organization task force first sought to define and classify cardiomyopathies. They defined cardiomyopathies as "heart muscle diseases of unknown cause" with three main classifications including: hypertrophic cardiomyopathy (HCM), dilated cardiomyopathy (DCM), and restrictive cardiomyopathy [1]. Over the next three decades it became patently obvious that this simple definition was not sufficient to describe the complex heterogeneity of diseases present in the patient population. More robust definitions were necessary for mechanistic links to be established and meaningful therapeutics to be developed. Since then the accepted definition of a cardiomyopathy has evolved and the classifications have greatly expanded. The most recent definition from the American Heart Association Council on Clinical Cardiology states: Cardiomyopathies are a heterogeneous group of diseases of the myocardium associated with mechanical and/or electrical dysfunction that usually (but not invariably) exhibit inappropriate ventricular hypertrophy or dilatation and are due to a variety of causes that frequently are genetic. Cardiomyopathies either are confined to the heart or are part of generalized systemic disorders, often leading to cardiovascular death or progressive heart failure–related disability [2]. This latest definition (2006) reflects the growing recognition of molecular genetics as a key factor in the development of cardiomyopathies and highlights the ever-growing complexity of disease classification. Today the genetic basis of HCM and DCM is widely recognized yet our understanding of the precise mechanisms underlying the disease remains unclear. To add to this disconnect, by the time patients become symptomatic, pathology has progressed past the initial phase, where meaningful treatment could occur, to advanced end-stage pathology. By this time often the only treatment options available become "blunt sword" therapeutics that are non-specific and used primarily for symptom management. In fact, over the last 3 decades there has been a marked decline in the innovation of cardiovascular pharmaceuticals owed partially to the vast complexity of disease presentation and progression [3].
In this dissertation, I will focus on a genetic sarcomeric DCM caused by a mutation in alpha-tropomyosin (Tm). Using novel accurate mouse models as a tool we will define the mechanism by which it leads to disease, investigate how disease severity due to the mutation is modified in an age-dependent manner, and examine what this mechanism could mean in the larger picture of cardiomyopathic disease progression. I hope to convince you that by using accurate models of this DCM at multiple levels of biological complexity to tease out the precise mechanisms of disease we can establish meaningful genotype-phenotype relationships that could lead to the development of specific novel therapeutics.
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Heydenreich, Monika. "Phänotypische Charakterisierung von Patienten mit hypertropher Kardiomyopathie und Varianten im [beta]-MHC-Gen [beta-MHC-Gen] und [alpha]-Tropomyosin-Gen [alpha-Tropomyosin-Gen]." [S.l.] : [s.n.], 2001. http://deposit.ddb.de/cgi-bin/dokserv?idn=965437485.
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Shanti, K. N. "Identification of Tropomyosin as the Major Cross-Reacting Crustacean Allergen." Thesis, Indian Institute of Science, 1994. http://hdl.handle.net/2005/103.
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Seafood including crustaceans, on ingestion, are known to provoke gastrointestinal as well as systemic allergic reactions. Crustaceans are aquatic arthropods with a chitinous exoskeleton and include shrimp, lobster, prawn and crab. Earlier studies in our laboratory have led to the identification and characterization of three allergens from shrimp, designated as Sa-I, Sa-I1 and Sa-III. The former two were shown to be heat stable proteins with a mol. wt. of 8.4 and 34 kDa respectively, while Sa-III was identified as tRNA Arg and TRNATyr ). Sa-II was found to be the major allergen contributing to more than 50% of the allergenic activity.
There are several reports on the existence of cross-reactivity among atopic allergens, in particular food allergens. It is well known that individuals with shrimp allergy often complain of adverse reactions following the ingestion of other re1ated crustaceans. Recognition of crustacea as a group causing adverse reactions in sensitive individuals has a basis in the close phylogenetic relationship of shrimp, lobster, crab and prawn. Thus, one could expect appreciable similarity in the IgE binding epitopes of the offending allergens from related crustaceans. The present study was, therefore, aimed towards the identification of the major cross-reacting crustacean allergen and localization of its IgE binding epitopes. Cross-reactivity among a1lergens from shrimp, prawn, crab and lobster was evaluated by immunochemical methods. Antigenic cross-reactivity was established by immunodiffusion using shrimp-specific rabbit IgG. Competitive ELlSA inhibition experiments using sera of shrimp sensitive patients revealed a high degree of allergenic cross-reactivity between different crustaceans. SDSPAGE and immunoblot analysis using the sera of shrimp sensitive patients have identified a 34 kDa protein as the cross-reacting crustacean allergen. Using shrimp as a model system and Sa-II as a representative crustacean allergen, further studies were carried out to get an insight into the structural and molecular basis of allergenic cross-reactivity. The strategies adopted were, (1) to raise allergen specific anti-idiotypic antibodies and explore the possibility of using these anti-idiotypic antibodies as surrogate allergens for diagnosis of crustacea allergy and (2) to identify the IgE binding epitopes on the major shrimp allergen Sa-II, which may be shared by the 34 kDa allergen from the related crustaceans.
In order to explore idiotypic, anti-idiotypic and anti-anti-idiotypic responses to Sa-II, Balb/c mice were immunized with affinity purified human idiotypic antibodies directed against the purified allergen. This resulted in the production of anti-idiotypic antibodies which were quantitated using rabbit idiotypic antibodies raised against the same allergen. The mouse anti-idiotypic antibodies recognized shrimp-specific human idiotypic antibodies of the IgE isotype from 18 of 20 individuals, and IgG antibodies from 14 of 20 shrimp sensitive patients. Immunization of Balb/ c mice with affinity purified, allergen-specific anti-idiotypic antibodies induced anti-allergen IgE and IgG responses in the absence of the allergen. The induction of anti-anti-idiotypic antibodies functionally identical to allergen-specific idiotypic antibodies confirmed that the anti-idiotypic antibodies generated, are indeed a mirror image of the allergen. The present study thus provides evidence that anti-idiotypic antibodies raised against allergen-specific idiotypic antibodies may substitute for the original allergen in the induction of allergen-specific idiotypic antibodies. The demonstration of shared idiotopes on IgG and IgE antibodies in the sera of shrimp sensitive patients supports the use of allergen-specific anti-idiotypic antibodies as surrogate allergens. These anti-anti-idiotypic antibodies not only recognized Sa-II, but also the 34 kDa allergen from prawn, lobster and crab.
Cross-reactivity studies using polyclonal sera of shrimp sensitive patients and Sa-II anti-anti-idiotypic antibodies have attributed the allergenic cross-reactivity observed among the related crustaceans to the presence of highly conserved IgE binding epitopes on the 34 kDa crossreacting allergen from shrimp, crab, lobster and prawn. In order to identify the igE binding epitopes on Sa-11, it was subjected to limited tryptic digestion and the peptides were separated by reverse phase HPLC. Amino acid sequence analysis of these peptides and several other peptides generated by Asp N and Lys C treatment revealed an 861 homology with the muscle protein tropomyosin from the fruit fly Drosophila melanogaster, suggesting that the major shrimp allergen is tropomyosin. To establish that Sa-II is indeed tropomyosin, the latter was isolated from shrimp and its physicochemical and immunochemical properties were compared with those of Sa-II. Both tropomyosin and Sa-II had the same molecular mass and focused in the isoelectric pH range of 4.8-5.4. In the presence of 6 M urea, the mobility of both Sa-I1 and shrimp tropomyosin shifted to give an apparent molecular mass of 50 kDa, which is a characteristic property of tropomyosins. Shrimp tropomyosin bound to specific IgE antibodies in the sera of shrimp sensitive patients as assessed by competitive ELISA inhibition and immunoblot analysis. Tropomyosin, similar to Sa-I1 was subjected to limited tryptic digestion and the tryptic maps of both Sa-II and tropomyosin as obtained by reverse phase HPLC were found to be super imposable. Dot blot immunoassay and competitive ELISA inhibition assay using the sera of shrimp sensitive patients identified two peptides, 6 and 9 that exhibited allergenic activity. Both the peptides were purified to homogeneity and sequenced. Peptide 6 is a nonapeptide corresponding to the amino adds 153-161 and peptide 9 has 17 amino acids corresponding to the aminoacid residues 50-66. The peptides individually blocked upto 50% the binding of allergen-specific IgE to hropomyosin. Sa-II specific mouse anti-anti-idiotypic antibodies recognized not only tropomyosin, but also the two allergenic peptides, thus confirming that these peptides represent the major IgE binding epitopes. The IgG binding activity was found to be associated with peptides 6 and 9 as assessed by dot blot immunoassay using the sera of shrimp sensitive patients. Thus, it was found that both IgG and IgE binding epitopes on shrimp tropomyosin are identical. Tropomyosins from both phylogenetically related and unrelated species were assessed for allergenic activity using the sera of shrimp sensitive patients. It was found that allergenic activity was associated with tropomyosins from related crustaceans and from Drosophila melanogaster which shares 86% homology with shrimp tropornyosin. However, tropomyosins from totally unrelated species like yeast, chicken, bovine, rat, rabbit and human did not exhibit allergenic activity. A comparison of the amino acid sequence of shrimp tropomyosin in the region of IgE binding epitopes with the corresponding regions of bopomyosins from different species confirmed lack of allergenic cross-reactivity. The allergenic peptides 6 and 9 were able to inhibit the binding of tropomyosins from related crustaceans to shrimp tropomyosin-specific IgE antibodies to the same extent, confirming the presence of highly conserved IgE binding epitopes. It has been established for the first time that the major crustacean allergen is the heat stable muscle protein, tropomyosin, and extensive cross-reactivity between different members of crustacea is due to the presence of highly conserved IgE binding epitopes on tropomyosins from these sources. Thus, from the present study, information with respect to the amino acid sequence of tropomyosin and localization of its 1gE binding epitopes, could be used to design synthetic peptides corresponding to the B cell and T cell epitopes which would find application in the diagnosis and desensitization of individuals allergic to crustacea.
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Mackenzie, Cassidy. "The properties and function of tropomyosin dimers in muscle regulation." Thesis, University of Kent, 2017. https://kar.kent.ac.uk/69463/.
Full textAbstract:
Myosin binding to actin, and thus muscle contraction, is regulated by Tropomyosin (Tpm), Troponin (Tn) and calcium (Ca2+). Tpm, is an α-helical coiled-coil dimer, which exists as a homo- or heterodimer. Two major isoforms of Tpm are found in striated muscle, α and β. Though it is known that different dimers exist, the mechanism by which they form and exchange is not fully understood. The thermal stability and exchange between dimers was explored with the use of circular dichroism and SDS PAGE densitometry analysis. Homodimers showed little exchange to form heterodimers at temperatures up to 20 °C . Dimer stability at these temperatures reduces the need for chemical cross-linking samples. While extensive exchange was seen at 37 °C . Reverse exchange of WT and mutant (E54K - dilated cardiomyopathy mutant) containing heterodimers to form homodimers did not show the same extent of exchange, suggesting a dimer preference. Results showed the ability to determine dimer content of a solution with the use of polyacrylamide gels and chemical cross-linking. The thermal melting curves of Tpm highlighted a significant destabilisation of β Tpm against the α isoform. Tpm heterodimer containing E54K mutant showed a decreased thermal stability. Notable differences were seen not only for isoforms and homo- and heterodimers, but also for buffer conditions and protein tags. Increased salt concentrations led to an increase in thermal stability. Crosslinking dimers increased thermal stability, whereas addition of His-tags led to a decrease in thermal stability. Changes in thermal stability highlighted the need for caution when tagging or cross-linking the protein. Dimer exchange on actin provided conflicting results between SDS PAGE cosedimentation assays and pyrene fluorescence cosedimendation assays, which highlighted limitations of cross-linking Tpm and using fluorescent labels. The stiffness of Tpm dimers was explored using atomic force microscopy (AFM) to image Tpm particles. Significant differences were seen between the relatively stiff α Tpm and less stiff β isoform. Changes in stiffness of Tpm affect its ability to cooperatively activate the thin filament, and provides insight into the assembly of dimers in vivo.
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Asiri, Saeed Ahmed. "Effects of myopathy-causing mutations on Tropomyosin structure and function." Thesis, University of Leicester, 2017. http://hdl.handle.net/2381/40307.
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Tropomyosin determinants for actin binding have not been identified completely and the nature and position of residues involved in thin filament dynamics has not been established. To date a number of Tropomyosin mutations have been linked to several muscle diseases including cardiomyopathies and skeletal muscle myopathies. In this thesis, we aimed to investigate the following tropomyosin mutations R90G, E163K, R167G, E240K, R244G and M2811 which have been shown to cause several severe skeletal muscle myopathies. We used various structural, biochemical and kinetic methods to assess the impact of these mutations on tropomyosin structure and biochemical properties. Fluorescence emission spectroscopy, and transient kinetics were used to assess the effect of these mutations on the equilibrium distribution and kinetics of transitions between different thin filament regulatory states. Overall the data demonstrated that: 1) all tropomyosin mutations except (E163K, E240K, M2811 and R90GR167G) affected the thermal stability of tropomyosin but not the a-helical coiled coil structure. 2) The size of the cooperative unit n was reduced by all tropomyosin mutations. 3) Tropomyosin mutations did not affect the proportion of thin filaments in the blocked state (at low Ca2+). 4) Tropomyosin mutations did affect the maximum observed rate constant of thin filament transition between the ON and OFF states. 5) Several tropomyosin mutations have affected tropomyosin-troponin binding affinity but none of the mutations had any effect on the actin binding affinity. Overall these results provide insight into the mechanism by which tropomyosin bind actin and troponin, tropomyosin related thin filament cooperativity and allosteric transitions.
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Pieples, Kathy. "THE FUNCTIONAL SIGNIFICANCE OF THE STRIATED ISOFORM OF TROPOMYOSIN 3 IN NORMAL AND PATHOLOGICAL STATES." University of Cincinnati / OhioLINK, 2001. http://rave.ohiolink.edu/etdc/view?acc_num=ucin997992638.
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Graham, Ian R. "Alternative splicing of tropomyosin pre-mRNA : control in non-muscle cells." Thesis, University of Leicester, 1992. http://hdl.handle.net/2381/35232.
Full textAbstract:
Alternative splicing of tropomyosin pre-mRNA: control in non-muscle cells. Ian R. Graham The human tropomyosin gene hTMnm contains a pair of mutually exclusive exons, NM and SK, which are used in non-muscle and skeletal muscle cells, respectively. I have undertaken an analysis of the factors affecting the splicing of these exons in the non-muscle cell line COS-1. I used a strategy involving mutation of the gene, followed by recloning of the appropriate region into a mammalian expression vector containing a tropomyosin cDNA clone. The wild-type and mutant mini-genes were transfected into the cell line, and the RNA produced after 48 hours' expression was isolated, then analysed by S1 nuclease protection mapping, and by a reverse transcriptase-polymerase chain reaction (RT-PCR) process. The results showed that exons NM and SK are not in competition in this non-muscle cell line; rather, I have shown that exon SK is not recognised as a splicing substrate when any other exons are present that can be used instead. Improvement, by mutation, of the branchpoint associated with exon SK restored use of that exon, as did replacement of the extreme 5' and 3' ends of the exon with the corresponding sequences from exon NM. The observation that exon SK is still overlooked by the cell's splicing apparatus, when it is placed in the exact context normally occupied by exon NM, strongly suggests that the exon itself is contributing to its deficiency. I have proposed a model in which the poor branchpoint sequence and elements within exon SK are responsible for preventing its recognition in the non-muscle cell, which is overcome, in skeletal muscle, by stimulation of the exon 4 to exon SK splice. Additionally, by studying the alternative splicing of the chick a-actinin gene, I have attempted to compare the regulation of splicing in smooth and skeletal muscle.
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Clayton, Joseph Emerson. "Barcoding the actin track: Differential regulation of myosin motors by tropomyosin." ScholarWorks @ UVM, 2016. http://scholarworks.uvm.edu/graddis/638.
Full textAbstract:
Myosins and tropomyosins represent two types of actin filament-associated proteins that often work together in contractile and motile processes in the cell. While the role of thin filament troponin-tropomyosin complexes in regulating striated muscle myosin II is well characterized, the role of tropomyosins in non-muscle myosin regulation is not well understood. Fission yeast has recently proved to be a useful model with which to study regulation of myosin motors by tropomyosin owing to its tractable genetics, well-defined actin cytoskeleton, and established actin biochemistry.
A hallmark of type V myosins is their processivity -- the ability to take multiple steps along actin filaments without dissociating. However, the fission yeast type V myosin (Myo52) is a nonprocessive motor whose activity is enhanced by the sole fission yeast tropomyosin (Cdc8). The molecular mechanism and physiological relevance of tropomyosin-mediated regulation of Myo52 transport was investigated using a combination of in vitro and in vivo approaches. Single molecules of Myo52, visualized by total internal reflection fluorescence microscopy, moved processively only when Cdc8 was present on actin filaments. Small ensembles of Myo52 bound to a quantum dot, mimicking the number of motors bound to physiological cargo, also required Cdc8 for continuous motion. Although a truncated form of Myo52 that lacked a cargo-binding domain failed to support function in vivo, it still underwent actin-dependent movement to polarized growth sites. This result suggests that truncated Myo52 lacking cargo, or single molecules of wild-type Myo52 with small cargoes, can undergo processive movement along actin-Cdc8 cables in vivo. These findings outline a mechanism by which tropomyosin facilitates sorting of transport to specific actin tracks within the cell by switching on myosin processivity.
To understand the broader implications of actomyosin regulation by tropomyosin we examined the role of two mammalian tropomyosins (Tpm3.1 and Tpm4.2) recently implicated in cancer cell proliferation and metastasis. As previously observed with Cdc8, Tpm3.1 and Tpm4.2 isoforms significantly enhance non-muscle myosin II (Myo2). Additionally, the mammalian tropomyosins enable Myo52 processive movement along actin tracks. In contrast to the positive regulation of Myo2 and Myo52, Cdc8 and the mammalian tropomyosins potently inhibit skeletal muscle myosin II, while having negligible effects on the highly processive mammalian myosin-Va. Thus, different motor outputs favoring functional specification within the same myosin class are possible in the presence of certain tropomyosins. In support of a conserved role for certain tropomyosins in regulating non-muscle actomyosin structures, Tpm3.1 rescued normal contractile ring dynamics, cytokinesis, and fission yeast cell growth in the absence of functional Cdc8. This work has broad implications with regard to regulation of non-muscle and muscle actomyosin function in complex cellular environments such as developing muscle tissue and metastatic cancer cells.
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Sereda, Michal Janusz. "Characterization of the molecular and immunological properties of Acanthocheilonema viteae tropomyosin." Doctoral thesis, Humboldt-Universität zu Berlin, Mathematisch-Naturwissenschaftliche Fakultät I, 2009. http://dx.doi.org/10.18452/15882.
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Diese Arbeit beschreibt die immunologischen Eigenschaften von Acanthochilonema viteae Tropomyosin, einem Muskel-assoziierten Protein. A. viteae ist ein zu den Filarien gehörender Parasit von Gerbilen, ähnlich dem humanpathogenen Filarie Onchocercha volvulus. Diese Arbeit hatte die funktionelle Charakterisierung von A. viteae Tropomyosin im Kontext der natürlichen Infektion und experimentellen Vakzinierung zum Ziel. Das allergene Potential des Tropomyosins und die Produktion von spezifischen IgE-Antikörpern wurden untersucht. Außerdem wurden Tropomyosin-spezifische monoklonale Antikörpern (mAk) entwicklet. Es konnte gezeigt werden, dass Tropomyosin als vielversprechender Kandidat zur Vakzinentwicklung gegen Filariosen angesehen werden kann, wobei berücksichtigt werden muss, dass deutliche Effekte nur unter Th1-Bedingungen auftreten. Die Vakzinierung mit Protein oder DNA reduzierte Adultwurmzahlen um 30 bzw. 45%. Während einer Infektion fungiert Tropomyosin als funktionelles Allergen und führt zur Produktion von hohen Mengen spezifischen IgEs. Ein Screening synthetischer Peptid-Bibliotheken zeigte gemeinsame 13 IgG- und 11 IgE-Epitope. Weiters konnte Kreuzreaktivität mit anderen Tropomyosinen und diesen gemeinsamen IgE-Epitopen nachgewiesen werden. Mit Hilfe von mAk konnte gezeigt werden, dass Tropomyosin auf der Oberfläche der Kutikula der Larvenstadien L3 und microfilariae des Parasiten vorkommt. Durch die Deglykosylierung des nativen Proteins wurde deutlich, dass einige Epitope von posttranslationellen Modifikationen gebildet werden. Weitere Immunisierungen mit Tropomyosin führten zu einem ähnlichen Profil der Zellaktivierung und Antikörperproduktion wie der Adjuvant Aluminiumhydroxid. Jedoch führte die Behandlung zur IL-10 Produktion und zur Zunahme von Gr1+/ CD11b+ Zellpopulationen, welche natürliche Surpressors darstellen. Zusammenfassend kann gesagt werden, dass A. viteae Tropomyosin immunmodulierende Eigenschaften aufweist und als Komponente eines zu entwickelnden Vakzins in Frage kommt.This study describes the immunological properties of Acanthocheilonema viteae muscle-associated protein tropomyosin. A. viteae is a filarial parasite of jirds that resembles the important human parasite Onchocerca volvulus. Focus of experiments is on unraveling the functional properties of tropomyosin in the context of an infection and experimental vaccination. Additionally, allergenic potential of tropomyosin was investigated and the ability to induce high levels of specific IgE. A part of the study was also aimed at the development of anti-tropomyosin monoclonal antibodies (mAb). This study revealed that tropomyosin is a promising antigen for vaccines against filarial nematodes, however, effective only in a Th1 biased environment. Vaccination with protein or DNA resulted in 30% - 45% protection that was not associated with specific IgG or IgE. During infection tropomyosin is an allergen and leads to the production of high levels of specific IgE. Screening of synthetic peptide libraries showed 13 IgG and 11 IgE co-located epitopes and revealed cross-reactivity with other tropomyosins and sharing of IgE epitopes. mAb were raised against A. viteae tropomyosin and showed that tropomyosin is abundant on the cuticle of L3 and microfilariae of the parasite. Deglycosylation of the native protein showed that some epitopes were formed by the posttranslational modifications. Additionally, immunization shows that tropomyosin induces a similar pattern of cell activation and antibody production as aluminium hydroxide adjuvant, but leads to the induction of IL-10 and the increase of population of GR1+/CD11b+ cells. These cells are regarded as natural suppressors. Taken together, results show that A. viteae tropomyosin has immunomodulating properties and can be considered as a component of an efficient vaccine.
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Clark, Ian David. "A fluorescence study of the COOH-terminus region of equine platelet tropomyosin." Thesis, University of British Columbia, 1987. http://hdl.handle.net/2429/26190.
Full textAbstract:
The use of fluorescent molecules as probes of protein conformation is recognized as a technique which provides very specific information and has been applied, in recent years, to the study of the role of tropomyosin (TM) in the regulation of contractile processes. The isolation and sequencing of TM from horse blood platelets (P-TM) has shown it to be different from muscle TM, especially near the NH₂-and COOH-termini. These differences have been suggested to weaken end-to-end interaction of P-TM molecules. TM's are two chain coiled coils and P-TM has cysteine residues at the penultimate COOH-terminus position of adjacent chains. These can be labelled with
sulfhydryl-specific fluorescent probes that reflect conformational changes in that region of the molecule via changes in their emission characteristics.
The results of experiments on both pyrene (Py) (40) and acrylodan (AD) labelled P-TM show that there is a preferred interaction of the COOH-terminus of P-TM with the NH₂-terminus of cardiac TM over that with the NH₂-terminus of P-TM. This indicates that the altered NH₂-terminus of P-TM, with respect to muscle TM, is responsible for the relative loss of polymerizability of P-TM at low salt concentration. Addition of actin to the Py-P-TM (40) and AD-P-TM species showed changes in emission characteristics indicative of binding to the F-actin filaments, suggesting
that the presence of the probes had not affected the function of the P-TM adversely. However, the presence of pyrenes at the COOH-terminus seemed to reduce further the ability of P-TM to self-polymerize. Thermal denaturation of AD-P-TM, AD-C-TM and AD-labelled truncated P-TM followed by fluorescence polarization suggested that, contrary to the theory of Skolnick and Holtzer on the stability of two chain coiled coils, the region towards the COOH-terminus is among the last to lose its helical character.Science, Faculty of
Chemistry, Department of
Graduate
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Coles, J. L. "The regulation of a novel exon in the rat α-tropomyosin gene." Thesis, University of Cambridge, 2007. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.597846.
Full textAbstract:
A conserved novel exon has been identified downstream of exon 3 of the rat α-tropomyosin gene. Inclusion of this exon is predicted to result in nonsense mediated decay of the α-tropomyosin transcript due to the presence of premature termination codons. Previous work showed that this nonsense exon is readily activated by a number of mutations in the 3’splice site and the 5’ end of the exon. Furthermore, there is evidence that the nonsense exon is activated in kidney, heart and lung tissue, indicating that its inclusion may be regulated in a tissue-specific manner. In this study, several computational data-sets were used to predict the locations of auxiliary cis-acting regulatory elements along the length of the nonsense exon. This information was used to design mutants in the putative regulatory elements, which revealed the presence of at least five exons splicing silencers (ESSs) and four exon splicing enhancers (ESEs). A number of trans-acting factors were subsequently identified as regulating the inclusion of the nonsense exon. These include hnRNP H and hnRNP F, the binding of which to a G-rich ESS at the 5’ end of the nonsense exon is required for repression. hnRNP A1 and CELF4 activate nonsense exon inclusion possibly by acting as anti-repressors of hnRNP H and hnRNP F. In addition SRp30c was identified as an activator of the nonsense exon, whilst SRp20 and hnRNP I (PTB) were identified as additional repressors, though the binding sites for SRp30c and SRp20 have yet to be elucidated. Inclusion of the nonsense exon was shown to result in NMD of the transcript in an α-tropomyosin-based reporter construct.
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Fairhead, Giles. "The interactions of Troponin T with Troponin C, Troponin I and Tropomyosin." Thesis, University of Birmingham, 2004. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.422010.
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Taylor, Claire Frances. "The regulation of splicing of the human alpha sigma-tropomyosin pre-mRNA." Thesis, University of Leicester, 1996. http://hdl.handle.net/2381/35167.
Full textAbstract:
The human alpha-s-tropomyosin pre-mRNA, encoded by the hTMnm gene, is alternatively spliced. In skeletal muscle cells, amino acids 189-213 are encoded by a skeletal-muscle specific exon, called SK. In all other cell types, amino acids 189-213 are encoded by an alternative exon called NM. The pattern of splicing of SK and NM is mutually exclusive. Previous work had shown that in non-muscle cells, the selection of NM was determined by the intrinsic inactivity of the SK exon. SK inactivity was believed to be due to non-muscle-specific repression of the SK exon. The principal cis-acting sequence mediating this repression was believed to lie within the first fifteen nucleotides of the SK exon [Graham et al., 1992]. The aim of this thesis was to identify and characterise trans-acting factors which bound to the first fifteen nucleotides of the SK exon. It was hoped that this approach would identify the putative trans-acting repressor of the SK exon. Two factors which bound with apparent specificity were identified. The first of these was a 56 kDa protein, believed to be a member of the Y-box family of transcription factors. Irreproducibility of certain experiments, coupled with failure to successfully perform necessary confirmatory experiments mean that the status of p56 as a putative SK repressor remains uncertain. Data is presented to show that U1 snRNP also binds within the first fifteen nucleotides of SK, at a site which partially overlaps the 3' splice site. A hypothesis is proposed in which U1 snRNP is directly involved in the repression of SK in non-muscle cells. This finding is discussed in the context of related systems of alternative splicing.
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Pasquet, Stéphanie. "Etude de la régulation transcriptionnelle du gène alpha-tropomyosine dans les cellules musculaires." Bordeaux 2, 2003. http://www.theses.fr/2003BOR21036.
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Nous avons étudié la régulation de la transcription du gène alpha-tropomyosine (TM) dans les cellules musculaires lisses (CML), squelettiques et cardiaques en culture. Les séquences régulatrices C-rich et MCAT activent la transcription dans les trois types cellulaires. Le trans-facteur TEF-1, qui lie la séquence MCAT, joue un rôle majeur dans la régulation de la transcription dans les trois types de muscles. Le facteur SRF interviendrait de façon indirecte, dans les CML et les cardiomyoctes, en augmentant la liaison de TEF-1 sur la boîte MCAT, et directement dans les myoblastes squelettiques en se liant à la boîte CArG. La fonction activatrice de TEF-1 et SRF semble corrélée à leur localisation sub-cellulaire dans les CMLWe have studied the transcriptional regulation of alpha-tropomyosin (α-TM) gene, in smooth, skeletal and cardiac muscle cells in culture. The regulatory sequences, C-rich and MCAT enhance transcription of the α-TM gene in the three muscle types. TEF-1 trans-factor plays a major role in the transcriptional regulation in the three muscle types, by binding to MCAT sequence. SRF factor seems to be involved, directly and indirectly, in the transcription activation of the gene. SRF could act indirectly in smooth and cardiac muscle cells, by enhancing TEF-1 binding, and directly in skeletal muscle cells, by binding to a CarG box. The role of TEF-1 and SRF factors could be related to their subcellular localization in smooth muscle cells
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Marknell, DeWitt Åsa. "Use of recombinant allergens for component-resolved diagnostics (CRD) in IgE-mediated allergy /." Uppsala : Acta Universitatis Upsaliensis : Univ.-bibl. [distributör], 2007. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-7813.
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Schulz, Emily M. "To Phosphorylate or Not to Phosphorylate: The Role of Tropomyosin Phosphorylation in Cardiac Function and Disease." University of Cincinnati / OhioLINK, 2012. http://rave.ohiolink.edu/etdc/view?acc_num=ucin1355156757.
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Halsall, D. J. "The effects of troponin and tropomyosin on rabbit skeletal actomyosin subfragment 1 interactions." Thesis, University of Bristol, 1987. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.378788.
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Gaillard, Catherine. "Le gene alpha-tropomyosine chez xenopus laevis : organisation et etude de son controle transcriptionnel." Rennes 1, 1996. http://www.theses.fr/1996REN10152.
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L'objectif de ce travail a ete de caracteriser la structure du gene alpha-tm de xenope et d'etudier les modalites de son controle transcriptionnel au cours du developpement. L'analyse de deux clones genomiques nous a permis de montrer que la structure de la region 5' du gene etait conservee par rapport aux genes aviaires et mammiferes. Cette region possede deux promoteurs et le couple d'exons alternatifs 2a/2b. Nous avons montre que l'exon 2a etait specifiquement utilise dans les cellules musculaires lisse. Par une approche utilisant la pcr, nous avons etablit la structure partielle du gene et localise les exons 3 a 7 et 9a, 9b et 9d par homologie avec les genes tm des autres especes. Afin de determiner si le gene pouvait generer des isoformes specifiques du cerveau homologues a celles decrites chez le rat et le poulet, nous avons crible une banque d'adnc de cerveau. Aucun des clones isoles ne correspond aux isoformes decrites. La sequence de la region comprise entre les exons 9b et 9d ne revele pas la presence de l'exon 9c decrit chez les mammiferes et les aviaires. Mais un des clone isole code pour une nouvelle isoforme de tm. Nous avons montre que les deux promoteurs du gene etaient actives de maniere sequentielle au cours du developpement. Le promoteur interne qui genere les isoformes non musculaires est actif des le debut de l'ovogenese alors que le promoteur distal est silencieux. Le promoteur distal qui genere les isoformes musculaires est active des le stade 15 de l'embryogenese. Les arnm musculaires squelettiques sont localises dans les somites et le cur embryonnaire indiquant que le gene est un marqueur precoce des lignages musculaires squelettiques et cardiaques. Les arnm codant pour les isoformes de muscle lisse apparaissent plus tardivement au cours de l'embryogenese vers le stade 40. Par la technique de surexpression de genes dans des explants embryonnaires de blastula, nous avons montre que le gene pouvait etre active par les facteurs myogeniques de la famille myod et que les facteurs mef2 pouvaient moduler cette expression. Afin d'etudier le controle transcriptionnel du gene dans les cellules musculaires, nous avons construit un ensemble de genes chimeriques et analyse leur activite transcriptionnelle dans l'embryon et les cellules myogeniques de caille en culture. Nous avons defini un promoteur minimal musculaire qui correspond a la region de 285 nt en amont du site d'initiation de la transcription. Une analyse mutationnelle nous a permis de caracteriser quatre elements importants pour la transcription du gene dans les cellules musculaires, a savoir les boites e, c, m-cat, carg et une region riche en a/t
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Hamon, Gouault Sandra. "Etude de la maturation différentielle du gène alpha-fast tropomyosine chez xénopus Laevis : identifications d'éléments en cis régulant l'utilisation d'un exon composite interne/3' terminal." Rennes 1, 2002. http://www.theses.fr/2002REN10046.
Full textAbstract:
L'exon 9A/9' du gène de l'alpha-tropomyosine de Xenopus laevis est utilisé en tant qu'exon 3' terminal dans les cellules musculaires embryonnaires ou en tant qu'exon interne dans les muscles striés embryonnaires et adultes. Il est par contre exclu dans les cellules non musculaires. L'exon 9A/9' se caractérise par des sites d'épissage et un signal de polyadénylation suboptimaux ainsi que par un point de branchement éloigné situé à 274 pb en amont du site d'épissage 3'. Une approche par transgenèse transitoire dans l'embryon a permis de mettre en évidence deux éléments régulateurs chevauchants en amont de l'exon 9A/9'. Un élément répresseur correspondant à des motifs consensuels de liaison à la PTB réprime l'utilisation de cet exon dans les cellules non musculaires. Un élément activateur correspondant à un motif conservé chez les oiseaux et les mammifères, jouxté de plusieurs pseudo points de branchement favorise la sélection de l'exon 9A/9' en tant qu'exon terminal dans les cellules musculaires embryonnaires.
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Grellscheid, S. N. "The role of cis-acting sequences in the regulation of α-tropomyosin alternative splicing." Thesis, University of Cambridge, 2003. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.599698.
Full textAbstract:
The main investigation was a study of the possible role of a zero length exon (ZLE) observed 234 nucleotides downstream of exon 3, in α-TM splicing. The ZLE overlaps with the DRE, and consists of potential branchpoints, a polypyrimidine tract and a 3’ splice site (3’ss) GAG, followed immediately by a 5’ splice site (5’ss). In addition, a pseudo 5’ss is located 107 nucleotides downstream of the 3’ss. All of these sequences are conserved between rat mouse and human α-TM genes. While the initial aim was to study the role of the ZLE, results from this study suggest that the ZLE is part of a 107 nucleotide pseudoexon, formed by the ZLE 3’ss and the downstream pseudo 5’ss. RT-PCR analysis of RNA from various rat tissues allowed identification of splicing intermediates involving the 3’ss of the pseudoexon and exons 2 or 3. Transfection studies showed that activation of the ZLE 3’ss or repression of the ZLE 5’ss resulted in inclusion of the pseudoexon, together with exon 3. Blocking the ZLE in the endogenous gene using an anti-sense approach resulted in an increase in levels of exon 2 products. Exon 3 transcripts including the pseudoexon contain a premature termination codon, and therefore, are expected to be degraded by nonsense mediated decay. This may represent a mechanism of negative regulation of α-TM exon 3. In a study of the optimum distance required between the branch point and the 3’ splice site in the α-TM system, inhibition of splicing of an alternative exon may require sub-optimal recognition of some key sequences, creating a capacity for regulation. The typical distance between the branchpoint and 3’ss AG is 18-40 nucleotides. In α-TM exons 2 and 3, this distance is unusually long. The resulting sub-optimal recognition of the polypyrimidine tract by U2AF, may be essential to allow competing regulatory factors to bind.
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Holeman, Teryn A., and Teryn A. Holeman. "Effects of Three Cardiomyopathic-Causing Mutations (D230N, D84N, and E62Q) on the Structure and Flexibility of α-Tropomyosin." Thesis, The University of Arizona, 2017. http://hdl.handle.net/10150/624101.
Full textAbstract:
Cardiac contraction at the level of the sarcomere is regulated by the thin filament (TF) composed of actin, alpha tropomyosin (TPM), and the troponin (Tn) complex (cTnT: cTnC: cTnI). The "gate-keeper" protein, α-TPM, is a highly conserved α-helical, coiled-coil dimer that spans actin and regulates myosin-actin interactions. The N-terminus of one α-TPM dimer inter-digitates with the C-terminus of the adjacent dimer in a head-to-tail fashion forming the flexible and cooperative TPM-overlap that is necessary for myofilament activation. Two dilated cardiomyopathy (DCM) causing mutations in TPM (D84N and D230N) and one hypertrophic cardiomyopathy (HCM) causing mutation (E62Q), all identified in large, unrelated, multigenerational families, were utilized to study how primary alterations in protein structure cause functional deficits. We hypothesize that structural changes from a single point mutation propagate along the -helical coiled-coil of TPM, thus affecting its regulatory function. Structural effects of the mutations studied via differential scanning calorimetry (DSC) on TPM alone revealed significant changes in the thermal unfolding temperatures of both the C- and N-termini for all mutants compared to WT, indicating that mutational effects propagate to both ends of TPM, thus affecting the overlap region. Although, of note, the proximal termini to the mutation has shown more significant structural changes compared to WT. DSC analysis on fully reconstituted TF’s (Tn:TPM:Actin) revealed effects on the TPM-Actin cooperativity of activation, affecting interaction strength (thermal stability), and the rigidity of TPM moving along actin (FWHM). To characterize the resultant functional effect of these discrete changes in thermal stability and TPM rigidity, ATPase assays were used to measure actomyosin activation in the presence and absence of Ca2+. Together, these data will provide a molecular level understanding of the structural and functional deficits caused by these mutations to help elucidate the mechanisms leading to disease.
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Nadal, Magriñà Laura. "Muscarinic, adenosine and tropomyosin-related kinase B receptors modulate the neuromuscular developmental synapse elimination process." Doctoral thesis, Universitat Rovira i Virgili, 2017. http://hdl.handle.net/10803/441749.
Full textAbstract:
El desenvolupament del sistema nerviós perifèric implica una inicial exuberant producció de neurones i, una posterior reducció dependent de l'activitat del nombre de sinapsis de les unions neuromusculars (NMJ). Aquest procés s’anomena eliminació sinàptica. Al final de la primera setmana postnatal, cada fibra muscular està innervada per una sola motoneurona. Els receptors muscarínics d’acetilcolina (mAChR), els receptors d’adenosina (AR) i el receptor cinasa de tropomiosina B (TrkB) podrien permetre la competició entre terminals nerviosos durant el procés d’eliminació sinàptica mitjançant la modulació de l’alliberament d’acetilcolina. En aquesta tesi s’ha investigat, mitjançant microscòpia confocal i un anàlisi morfològic quantitatiu, el paper dels receptors mAChRs (M1, M2 i M4), dels AR (A1 i A2A) i del receptor TrkB en el procés d’eliminació en el desenvolupament de la NMJ.
Els resultats mostren que els receptors mAChRs, AR i el receptor TrkB promouen una desconnexió axonal al principi de la segona setmana postnatal independentment de la maduració dels receptors d’acetilcolina postsinàptics. En resum, els receptors mAChRs, AR i el receptor TrkB endarrereixen el procés d’eliminació sinàptica a P7 però l’acceleren a P9. Pel que fa la cooperació d’aquests receptors, M4 produeix un efecte oclusiu sobre M1 i un efecte additiu sobre a P7. La cooperació entre els receptors M1, A1 i A2A promou la pèrdua axonal a P9, mentre que, l’efecte de M2 és independent dels altres receptors. El M1 i TrkB treballen junts per incrementar la pèrdua axonal a P9 independentment dels receptors M2 i TrkB.
En conclusió, l’eliminació sinàptica postnatal és regulada per un mecanisme que depèn de varis receptors, involucrant la cooperació dels diferents subtipus de receptors muscarínics, d’adenosina i el receptor TrkB, els quals garanteixen la monoinnervació de les sinapsis neuromusculars al final del procés.El desarrollo del sistema nervioso periférico implica una inicial exuberante producción de neuronas y, una posterior reducción dependiente de actividad del número de sinapsis en las uniones neuromusculares (NMJ). Este proceso se denomina eliminación sináptica. Al final de la segunda semana postnatal, cada fibra muscular esta inervadas por una solo motoneurona. Los receptores muscarínicos de acetilcolina (mAChR), los receptores de adenosina (AR) y el receptor quinasa de tropomiosina B (TrkB) podrían permitir la competición entre los terminales nerviosos durante el proceso de eliminación sináptica mediante la modulación en la liberación de acetilcolina. En esta tesis se ha investigado, mediante microscopía confocal y un análisis morfológico cuantitativo, el papel de los receptores mAChRs (M1, M2 y M4), de los receptores de adenosina (A1 y A2A) y del receptor TrkB en el del proceso de eliminación en el desarrollo de la NMJ. Los resultados muestran que los receptores mAChRs, AR y el receptor TrkB promueven una desconexión axonal al inicio de la segunda semana postnatal independientemente de la maduración de los receptores de acetilcolina postsinápticos. En resumen, los receptores mAChRs, AR y el receptor TrkB retrasan el proceso de eliminación sináptica en P7 pero lo aceleran en P9. En la cooperación de estos receptores, se ha demostrado que M4 produce un efecto oclusivo sobre M1 y aditivo sobre A1 en P7. La cooperación entre M1, A1 y A2A promueve la pérdida axonal en P9, mientras que M2 es independiente de los otros receptores. M1 y TrkB cooperan para incrementar la pérdida axonal en P9 independientemente de M2 y TrkB. En conclusión, la eliminación sináptica postnatal está regulada por un mecanismo que depende de varios receptores, involucrando la cooperación de diferentes subtipos de receptores muscarínicos, de adenosina y el receptor TrkB, los cuales garantizan la monoinnervación de las sinapsis neuromusculares al final del proceso. saludable.
The development of the peripheral nervous system involves an initially exuberant production of neurons and a subsequent activity-dependent reduction in the number of synapses at the neuromuscular junctions (NMJ). This process is called synaptic elimination. At the end of the first postnatal week, each muscle fiber is innervated by a single motoneuron. Muscarinic acetylcholine receptors (mAChR), adenosine receptors (AR) and the tropomyosin-related kinase B (TrkB) receptor may allow the direct competition between nerve endings during synapse elimination through the modulation of acetylcholine release. Here, it has been investigated by confocal microscopy and quantitative morphological analysis the involvement of the individual and synergic or oclusive effect of M1-, M2- and M4-subtypes of mAChRs, A1 and A2A of ARs and TrkB in the control of the axonal elimination in developing NMJ. The results show that mAChRs, ARs and TrkB promote axonal disconnection at the beginning of the second postnatal week without affecting the postsynaptic maturation of the nicotinic receptor cluster. In summary, mAChRs, ARs and TrkB delay axonal loss at P7 but accelerate it at P9. In terms of receptor cooperation, M4 produces some occlusion of the M1 pathway and some addition to the A1 pathway at P7. The cooperation between M1, A1 and A2A receptors promotes axonal loss at P9, whereas the effect of M2 is independent of the other receptors. M1 and TrkB receptors work together to increase axonal loss rate at P9 but the effect of M2 is largely independent of the TrkB receptor. In conclusion, postnatal synapse elimination is a regulated multireceptor mechanism involving the cooperation of several muscarinic, adenosine and TrkB receptor subtypes that guarantees the monoinnervation of the neuromuscular synapses in the end of the process.
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Ly, Thu, Natalia Moroz, Christopher T. Pappas, Stefanie M. Novak, Dmitri Tolkatchev, Dayton Wooldridge, Rachel M. Mayfield, Gregory Helms, Carol C. Gregorio, and Alla S. Kostyukova. "The N-terminal tropomyosin- and actin-binding sites are important for leiomodin 2's function." AMER SOC CELL BIOLOGY, 2016. http://hdl.handle.net/10150/621526.
Full textAbstract:
Leiomodin is a potent actin nucleator related to tropomodulin, a capping protein localized at the pointed end of the thin filaments. Mutations in leiomodin-3 are associated with lethal nemaline myopathy in humans, and leiomodin-2-knockout mice present with dilated cardiomyopathy. The arrangement of the N-terminal actin- and tropomyosin-binding sites in leiomodin is contradictory and functionally not well understood. Using one-dimensional nuclear magnetic resonance and the pointed-end actin polymerization assay, we find that leiomodin-2, a major cardiac isoform, has an N-terminal actin-binding site located within residues 43-90. Moreover, for the first time, we obtain evidence that there are additional interactions with actin within residues 124-201. Here we establish that leiomodin interacts with only one tropomyosin molecule, and this is the only site of interaction between leiomodin and tropomyosin. Introduction of mutations in both actin- and tropomyosin-binding sites of leiomodin affected its localization at the pointed ends of the thin filaments in cardiomyocytes. On the basis of our new findings, we propose a model in which leiomodin regulates actin poly-merization dynamics in myocytes by acting as a leaky cap at thin filament pointed ends.
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BALVAY, LAURENT. "Etude de la regulation de l'epissage des exons alternatifs 6a et 6b des genes de tropomyosine." Paris 7, 1994. http://www.theses.fr/1994PA077007.
Full textAbstract:
La plupart des genes codant pour des proteines eucaryotes sont divises en introns et en exons. L'epissage permet de passer de la structure morcelee de l'information codante sur l'adn (exons) a la sequence des arns messagers. Ce processus se complique quand un meme gene comporte plusieurs exons potentiels pouvant engendrer plusieurs types de transcrits. On parle alors d'epissage alternatif. La diversite des isoformes de tropomyosine chez les vertebres est essentiellement generee par des phenomenes d'epissage alternatifs. Quatre genes sont ainsi a l'origine de plus d'une vingtaine d'isoformes chez les oiseaux. Le gene de la beta tropomyosine contient deux exons internes mutuellement exclusifs: l'exon 6a et l'exon 6b qui sont utilises respectivement dans les myoblastes puis dans les myotubes au cours de la differenciation d'un muscle squelettique. Le travail presente concerne d'une part la caracterisation des transcrits specifiques du cerveau du gene de la tropomyosine alpha-tm1 aviaire, et d'autre part l'etude des mecanismes d'epissage alternatifs par lesquels le gene beta genere plusieurs types de transcrits au cours du developpement. Nous avons montre qu'une region riche en pyrimidines situee dans la partie 5' de l'intron entre les deux exons alternatifs est necessaire a la repression de l'exon 6b et a l'activation de l'exon 6a dans les cellules non musculaires et les cellules de muscle lisse. La caracterisation de l'ensemble des elements en cis montre que la machinerie d'epissage est controlee dans les myoblastes par une repression de l'utilisation de l'exon 6b et dans les myotubes par une modulation du consensus des sites d'epissages permettant une competition en faveur de cet exon. Des comparaisons entre les differents genes beta de vertebres ont recemment permis de mettre en evidence des divergences dans le controle de l'epissage des exons alternatifs entre les classes aviaires, amphibiennes et mammaliennes. Elles pourraient montrer une evolution differente des processus de controle des epissages alternatifs
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Hardy, Serge. "Caracterisation d'adn complementaires tropomyosines chez le xenope. Expression et modeles d'organisation des genes alpha et beta." Rennes 1, 1991. http://www.theses.fr/1991REN10014.
Full textAbstract:
Les tropomyosines existent sous de nombreuses isoformes specifiques d'un tissu. Cette diversite est due a l'existence de plusieurs genes mais aussi a la synthese d'isoformes multiples a partir d'un meme gene par l'utilisation de promoteurs multiples et d'exons alternatifs. Nous avons entrepris une etude sur l'organisation des genes tropomyosines chez le xenopedans l'objectif d'utiliser l'ovocyte et l'embryon de xenope pour une etude in vivo des regulations transcriptionnelles et post-transcriptionnelles des genes tropomyosines. En criblant des banques d'adnc d'embryons et d'ovocytes de xenope nous avons isole six adnc. L'analyse des sequences de ces adnc et leurs comparaisons avec celles de genes tropomyosines d'autres especes nous ont permis de definir trois genes chez le xenope: 1) un gene alpha qui code pour une isoforme alpha squelettique en utilisant un promoteur distal et pour une isoforme non musculaire a partir d'un promoteur interne. Ce gene possede trois groupes d'exons epissesalternativement; 2) un gene beta qui code pour une isoforme beta squelettique et une isoforme beta de muscle lisse a partir du meme promoteur. Ce gene possede deux groupes d'exons episses alternativement; 3) le gene xtmp1 qui code pour une isoforme non musculaire. L'expression de ces trois genes a ete etudiee au cours du developpement precoce et dans differents tissus adultes
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Heydenreich, Monika. "Phänotypische Charakterisierung von Patienten mit hypertropher Kardiomyopathie und Varianten im Beta-MHC-Gen und Alpha-Tropomyosin-Gen." Doctoral thesis, Humboldt-Universität zu Berlin, Medizinische Fakultät - Universitätsklinikum Charité, 2002. http://dx.doi.org/10.18452/14777.
Full textAbstract:
Die hypertrophe Kardiomyopathie ist eine autosomal dominant vererbte Herzmuskelerkrankung. Es kommt zu einer asymmetrischen Hypertrophie insbesondere des Herzmuskels. Symptome sind unspezifisch und reichen von Dyspnoe bis hin zu Synkopen. Gelegentlich ist der plötzliche Herztod die erste Manifestation der Erkrankung. Molekulargenetische Untersuchung des Genomes dieser Patienten zeigten, dass diese Patienten Mutationen im Proteinen des Sarkomers aufwiesen. Hierunter fällt das beta-MHC-Gen und alpha-Tropomysin-Gen. Wir untersuchten 45 nicht miteinanderverwandte Patienten auf Mutationen im beta-MHC-Gen und im alpha-Tropomysin-Gen. Folgende molekulargenetische Untersuchungen wurden angewendet: Polymerase-Ketten-Reaktion, Single-Strand-Polymorphismus-Anlyse und Sequenzierung. Bei 6 Patienten (13%) fanden wir eine Mutation im beta-Myosin-Gen. Kein Patient hatte eine Mutation im alpha-Tropomyosin-Gen. In der Literatur wird eine Mutation im beta-MHC-Gen in bis zu 35% und im alpha-Tropomyosin-Gen in bis zu 3% der Fälle angenommen. Unsere Patienten hatten die Mutationen: Exon 13 Arg403Trp und Val411Ile, Exon 19 Arg719Trp, Exon 20 Ile736Thr, Exon 21 Leu796Phe und Exon 23 Cys905Phe. Die Mutationen Arg403Trp und Arg719Trp waren vorher bereits bekannt. Die Mutationen, Ile736Thr, Val411Ile, Leu796Phe und Cys905Phe wurden in der Form von uns erstmals ermittelt. Offensichtlich besitzen unsere Patienten überdurchschnittlich häufiger Mutationen in anderen Genen, die in dieser Studie nicht untersucht wurden. Der Phänotyp der Krankheit der HCM war bei unserem Patientenkollektiv sehr heterogen, und es ließen sich keine signifikanten Unterscheidungen eststellen. So haben wir uns darauf beschränkt, bei den 6 Patienten mit Mutationen nur nach den von Burn et al. (1997) aufgestellten Risikofaktoren zu suchen, die einen plötzlichen Herztod herbeiführen können, und haben sie danach in Patienten mit einem höheren oder niedrigeren Risiko eingestuft. Kriterien für ein erhöhtes Risiko, einen plötzlichen Herztod zu erleiden, wurden von Patienten mit den Mutationen: Exon 13 Arg403Trp, Exon 13 Val411Ile, Exon 19 Arg719Trp, Exon 20 Ile736Thr und Exon 21 Leu796Phe erfüllt. Mutationen an diesen Positionen sind auch in der Literatur mit einem erhöhten Risiko für einen plötzlichen Herztod assoziiert worden. Der Patient mit der Mutation im Exon 23 Cys905Phe wies wenige Risikofaktoren auf und unterscheidet sich somit nicht von den in der Literatur beschriebenen Patienten. Wir konnten dies mit unseren Ergebnissen bestätigen.Hypertrophic Cardiomyopathy is disease of the cardiac muscle which results in an asymmetric hypertrophy especially of the interventricularseptum of the heart. It is transmitted in an autosomal dominant way. The symptoms are unspecific reaching from dyspnoe to syncopes. Sometimes the sudden death is the first manifestation of the disease. Molekular genetic researches showed that in the patients genes Mutations in proteins of the sarkomer were detectable. Two of them are alpha-Tropomyosin and beta-Myosin Heavy Chain. We examined 45 unrelated Patient of the existence of Mutations in alpha-Tropomyosin and beta-Myosin Heavy Chain. We used following Examinations: PCR, SSCP, Sequencing. A mutation in the beta-Myosin Heavy Chain were found in 6 Patients (13%), non in alpha-Tropomyosin. Generally mutations are expected in 35% in beta-Myosin Heavy Chain and 3% in alpha-Tropomyosin. Our patients seem to have mutations in genes we did not examine in this study. We detected Mutations in: Exon 13 Arg403Trp and Val411Ile, Exon 19 Arg719Trp, Exon 20 Ile736Thr, Exon 21 Leu796Phe und Exon 23 Cys905Phe. Mutation Arg 403Trp and Arg719Trp have been known in this form before, the others were new. As the phenotypes of our patients were heterogenous and not significantly to be distinguished we looked for risk factors for sudden death as described by Burn et al. 1997 within our group of patients with mutations. Five Persons showed risk factors as discribed: Exon 13 Arg403Trp and Val411Ile, Exon 19 Arg719Trp, Exon 20 Ile736Thr, Exon 21 Leu796Phe. The person with the mutation Exon 23 Cys905Phe showed no risk factors for sudden death. Our results correlate with those of earlier studies. The patient with the mutation Exon 23 Cys905Phe was classified as a low risk patient while the other mutations correlate with a further high risk.
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Harder, Megan Michelle. "Morphological Changes Associated with Severe Early Onset Dilated Cardiomyopathy Caused by a Mutation in Alpha Tropomyosin." Thesis, The University of Arizona, 2015. http://hdl.handle.net/10150/579427.
Full textAbstract:
A point mutation in alpha tropomyosin (Tm) Asp230Asn (D230N) has been found in two unrelated multigenerational families to be causative for dilated cardiomyopathy (DCM). In these families, a distinct "bimodal" distribution of severity was observed whereby children have a severe DCM and adults have a mild to moderate phenotype. If children harboring this mutation survive the initial presentation of DCM, they often regain some systolic function. This "bimodal" presentation led us to ask what changes occur in the heart between fetal and postnatal life that could account for this; a potential candidate was the switch from fetal to adult cardiac troponin T (cTnT). Therefore, we hypothesize that temporal isoform switching by cTnT results in the "bimodal" phenotype observed in patients with D230N. Whole hearts, obtained from transgenic littermates of a murine model showed overall enlargement of the ventricles. H&E staining showed normal fiber size, distribution and no evidence of inflammation indicating that there is no myocarditis. Additionally, there was no histopathological evidence of collagen deposition in trichrome stained hearts. These results suggest a primary DCM in which the D230N mutation causes a structural DCM that is worsened in the presence of fetal TnT possibly due to further alterations in structure.
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Naye, François. "Caractérisation et rôle de TEF-1 au cours du développement embryonnaire de Xenopus Laevis." Bordeaux 2, 2007. http://www.theses.fr/2007BOR21467.
Full textAbstract:
Caractérisation et rôle de TRF-1 au cours du développement embryonnaire de Xenopus Levis. L'étude de la régulation du gène α-tropomyosine (α-TM)de xénope a permis d'identifier une région régulatrice de 30 pb dans la région promotrice du gène. Cette région est très conservée entre les différents gènes α-TM de vertébrés et possède une séquence MCAT (5' -CATTCCT-3') essentielle pour l'activité du gène dabs les trois lignages musculaires. Cette séquence fixe le facteur de transcription TEF-1. L'analyse par transgenèse a montré que l'expression spatiotemporelle correcte du gène α-TM dans l'embryon dépendait de la séquence MCAT intacte. L'étude du rôle de TEF-1, au cours du développement embryonnaire de xénope, a été entreprise. Deux gènes codant pour XNTEF-1 et le gène codant pour XVgl-2, co-facteur de TEF-1, ont été identifiés. Le profil d'expression spatiotemporelle de ces gènes, ainsi que leur régulation par les voies d'inductionmésodermique et les facteurs myogéniques ont été établis. La fonction de TEF-1 dans l'embryon a été analysée par une approche gain de fonction et perte de fonction. A fortes doses, l'injectiond'ARNm TEF-1 sauvage ou codant pour une protéine à effet dominant négatif (EnR-TEA) induit une apoptose. Pour de faibles doses l'ARNm TEF-1 induit une ventralisation de l'embryon. L'injection de l'ARNm ENR-TEA induit la formation d'un second axe dorsal et lève l'inhibition de la différenciation neurale par BMP. TEF-1 serait impliqué dans la ventralisation du mésoderme via la voie BMPCharacterization and role of TEF-1 during Xenopus laevis embryonic development. The analysis of the transcriptional control of the Xenopus laevis α-tropomyosin (α-TM) gene has led to the identification of a 30bp regulatory region. This region is conserved between all known vertebrate α-TM genes promoter ans contains an MCAT (5'-CATTCCT-3') sequence that is critical for the expression of the gene in all muscle cell types. This sequence can be bound by the transcription factor TEF-1 and is essential for a correct temporal and spatial expression of an α-TM transgene in the embryo. Studies about the role of TEF-1 in the embryonic development have been undertaken. Two genes, namely XNTEF-1, and the Xenopus Vestigial-2 gene, that encodes a TEF-1 cofactor, have been identified. The developmental expression pattern of these genes has been investigated together with their regulation by growth factors and myogenic factors. The role of TEF-1 in the developing embryo has been investigated by gain and loss of function strategies. The microinjection of high doses of either TEF-1 encoding mRNA or encoding a dominant negative protein (EnR-TEA) induced apoptosis. When low doses of TEF-1 mRNA are injected, embryos showed a ventralized phenotype. The microinjection of EnR-TEA mRNA induces the formation of a secondary dorsal axis in the embryo and blocks the neural inhibition due to BMP. In conclusion, TEF-1 could be implicated in the BMP signalling pathway which induces ventral mesoderm in the embryo
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LIBRI, DOMENICO. "Caracterisation du gene codant pour l'isoforme beta de la tropomyosine chez le poulet. Etude de l'epissage alternatif." Paris 11, 1990. http://www.theses.fr/1990PA112069.
Full textAbstract:
Le gene codant pour l'isoforme beta de la tropomyosine a ete isole et caracterise. Ce gene est soumis a l'epissage alternatif. Deux exons (6a et 6b) sont utilises de facon mutuellement exclusive et sont specifiques respectivement des cellules non musculaires/muscle lisse et du muscle squelettique. Des minigenes contenant les deux exons alternatifs entoures par deux exons constitutifs ont ete utilises pour transfecter des cellules myogeniques. L'etude du controle de l'epissage a ete poursuivi en introduisant des mutations dans la region de l'exon 6b. Cela nous a permis d'identifier un certain nombre d'elements impliques in cis dans cette regulation, parmis lesquels les sequences exoniques et introniques, la position anormale du point de branchement en amont de l'exon 6b et l'implication probable de structures secondaires du transcrit primaire. Des facteurs agissant in trans, probablement specifiques du muscle squelettique, sont egalement impliques dans la regulation
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Zhao, Rathje Li-Sophie. "Tropomyosin in Normal and Malignant Cells and the Action of Picropodophyllin on the Microfilament and Microtubule Systems." Doctoral thesis, Stockholms universitet, Wenner-Grens institut, 2009. http://urn.kb.se/resolve?urn=urn:nbn:se:su:diva-27767.
Full textAbstract:
Cell motility is a fundamental process, enabling cells to migrate, for instance during embryogenesis, tissue repair and defense. Force is generated by two protein systems, which also participate in cell proliferation, control macromolecular and organelle distribution and determine the fine structure of the cell interior. The major components of these are actin and tubulin, respectively, and they are referred to as the microfilament and the microtubule systems. This thesis focuses on tropomyosin, one of many microfilament associated proteins coupled to actin dynamics and organization and expressed in several isoform variants. Altered distribution and isoform expression of tropomyosin are signatures of malignant cells and are dealt with in the current thesis. The presence of tropomyosin isoforms in protruding lamellipodia of migrating cells is demonstrated, and a method to fractionate tropomyosin depending on its organization in an easily extractable, and a more tightly bound cytoplasmic form is presented. Analysis of the loosely associated tropomyosin fraction by gel filtration chromatography revealed that most of the tropomyosins in this fraction exist in a multimeric form. It was also observed that the distribution of tropomyosin varied between non-transformed and transformed cells with most of the isoforms enriched in the loosely bound fraction in the latter category of cells. Possibly this reflects the extensive reorganization of the microfilament system observed in cancer cells and which, depending on the context, can be normalized by introduction of certain tropomyosin isoforms. Many anti-cancer drugs target the microtubule system, inhibit cell division and promote apoptosis. Here it is shown that picropodophyllin, which has promising anticancer properties has a destabilizing effect on microtubules and via the microfilament system causes cells to detach from their substratum. Furthermore, picropodophyllin interferes with stimulation of the insulin-like growth factor receptor, which is involved in growth stimulation, differentiation and survival and whose expression is up-regulated in cancer cells.