Дисертації: "Molecular factor of recurrence"

1

Haller, Andrew Clayton. "The roles of the E26 transcription family member, SAM pointed domain-containing ETS transcription factor (SPDEF), in early stage prostate cancer and the development of castration recurrent disease." Thesis, State University of New York at Buffalo, 2013. http://pqdtopen.proquest.com/#viewpdf?dispub=3565756.

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One of the greatest problems in prostate cancer management today is accurate identification of patients who require treatment for aggressive disease versus those with indolent disease who are suitable for observational strategies. Histological appearance of the tumor, called Gleason score in the prostate cancer field, is the most predictive measure currently used. However, recent studies in multiple tumor types have shown that histological appearance does not always reflect the underlying molecular phenotype of the lesion. Therefore, in prostate cancer specifically, assessment of a molecular marker of androgen receptor driven epithelial differentiation may have clinical predicative capabilities. SAM pointed domain-containing Ets transcription factor (SPDEF) is a potential AR target gene that has shown to be necessary and sufficient for epithelial cell differentiation in many tissues. Although generally associated with good prognosis, SPDEF's role in cancer in unclear. This study demonstrates, through retrospective immunohistochemical analysis, the utility of SPDEF as a predictive biomarker for patients that have an extended benefit from androgen deprivation therapy (ADT). Furthermore, dual roles of SPDEF to inhibit the initiation and supporting the progression of castrate recurrent disease through novel androgen receptor expression regulation in castrate conditions are shown. In ADT naïve patients, SPDEF did not associate with metastatic disease or an induction of epithelial to mesenchymal transition. However, aggressive tumors tended to be larger, have greater SPDEF variability, and lack vimentin expression; a phenotype that could be explained by a partial EMT. In conclusion, SPDEF may be clinically useful to assess the epithelial phenotype of tumors, and could have utility identifying patients that will respond well to androgen deprivation therapy.

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2

RICCITELLI, RICCARDO. "L’utilizzo della Nadroparina Calcica in donne affette da abortività idiopatica ricorrente." Doctoral thesis, Università degli Studi di Roma "Tor Vergata", 2005. http://hdl.handle.net/2108/189.

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Inherited and acquired thrombophilias are associated with recurrent pregnancy loss(RPL) and venous thromboembolism(VTE). RPL is a major health problem affecting 1-2% of women at the reproductive age. While chromosomal aberration, endocrinologic dysfunctions and uterine abnormalities are etioloogic factors, a cause of RPL could not be identified. Inherited and acquired thrombophilias can be found in 50-65% of women with RPL of unknown cause, as well as in women with other vascular pathologies such peeclampsia, intrauterine restriction and placental abruption. Gestational outcame in woman with inherited thrombphilias who present with RPL is poor with less than 25% of prengnacies resulting in live birth. The purpose of this study is to verify the safe and efficacy of Nadroparin in RPL.

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3

Benzonana, Laura Lina. "Potential effects of anaesthetics on cancer recurrence following surgery : molecular mechanisms." Thesis, Imperial College London, 2012. http://hdl.handle.net/10044/1/39402.

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Patients with solid tumours are likely to undergo surgery during the course of their disease. Surgery and anaesthesia may influence the tumour's metastatic potential. Certain anaesthetics are shown to induce cellular phenotypic changes via cellular signalling pathways; including the hypoxia inducible factor (HIF) pathway. HIFs are heavily implicated in tumorigenesis and may play an important role in tumour cell proliferation, migration, invasion and angiogenesis (VEGF signalling). In the current thesis I aimed to investigate the potential impact of isoflurane on tumour cell progression and metastatic potential, as well as the potential anti-cancer effects of helium (a potential insuflation gas) in vitro. In a series of experiments, renal and prostate cancer cells were exposed to different anaesthetics and their effects on the metastatic potential of the cells were observed using different techniques such as western blotting, fluorescent immunocytochemistry, MTT assays, trypan blue assay and migration assays. The data derived from my PhD project show that isoflurane anaesthesia results in an increased metastatic potential of renal cell carcinoma and prostate cancer cells by increasing cell proliferation, migration, and invasion. The mechanism partially responsible for this effect has been shown to be the PI3K/HIF pathway. Furthermore helium was shown to have an anti-cancer effect on both cancer cell lines. These findings may have clinical implications for cancer patient care undergoing surgery under anaesthesia. Understanding the role of anaesthetics on growth and metastasis will defiantly shed light to better treatment options for cancer patients.

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4

Makino, Tomokazu. "Carbohydrate antigens as a risk factor for hematogenous recurrence of esophageal squamous cell carcinoma patients." Kyoto University, 2002. http://hdl.handle.net/2433/149342.

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5

TAKAHASHI, HIROSHI, CHISA HASHIZUME, TAKAHIKO TSUGAWA, YOSHIMASA MORI, and TATSUYA KOBAYASHI. "PROGNOSTIC FACTORS FOR TUMOR RECURRENCE AFTER GAMMA KNIFE RADIOSURGERY OF PARTIALLY RESECTED AND RECURRENT CRANIOPHARYNGIOMAS." Nagoya University School of Medicine, 2012. http://hdl.handle.net/2237/16031.

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6

Sideris, Michail. "The significance of molecular biomarkers in the recurrence of rectal tumors after Transanal Endoscopic Microsurgery." Thesis, King's College London (University of London), 2018. https://kclpure.kcl.ac.uk/portal/en/theses/the-significance-of-molecular-biomarkers-in-the-recurrence-of-rectal-tumors-after-transanal-endoscopic-microsurgery(e16b37d2-b2d4-47fb-9c30-563cd06be58d).html.

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Local excision (LE) of rectal cancer has been practiced as a treatment for 30 years on a highly selected group of patients and tumours. A method of local excision which has recently gained wider acceptance in the treatment of low-grade CRC (T1) is transanal endoscopic microsurgery (TEMS). TEMS offers advantages for operative access and oncological clearance compared to those of transanal resection (TAR). A number of studies have shown that TEMS can have almost equal results to radical surgery for early rectal cancer. Radical surgery has the disadvantage of an approximate mortality rate of 5% and a complication rate of around 20%, and it impacts on quality of life due to stomas and of sexual dysfunction. However, TEMS has a higher local recurrence rate, and efforts have been made to classify risk with morphological and histological criteria. Molecular biomarkers in the evaluation of CRC prognosis and treatment stratification have been extensively discussed in the literature. Until now, 3 pathways have been identified to explain the background of CRC molecular pathogenesis. Microsatellite instability (MSI) refers to point mutations in the DNA mismatch repair genes. MSI is currently responsible for 15−20% of CRC cases, and there is enough evidence to support its association with prognosis. Chromosomal instability (CIN) is another pathway of carcinogenesis which affects 85% of CRC cases and has been flagged as a poor prognostic marker. CIN encompasses any structural chromosomal abnormality that results in aneuploidy or polyploidy. The third pathway is related to aberrant hypermethylation of suppressor promoter CpG islands, commonly known as CIMP. v-raf murine sarcoma viral oncogene homolog B (BRAF) encodes a serine-threonine protein kinase that acts as a downstream effector of Kirsten rat sarcoma viral oncogene homolog (KRAS) pathways. Various studies have revealed that v-raf murine sarcoma viral oncogene homolog B V600E (BRAF V600E) mutations appear to be valid prognostic indicators. KRAS is a proto-oncogene that encodes a GTPase, which is involved in facilitating cellular response to extracellular stimuli. KRAS point mutations appear in 40% of CRC cases, and their presence is associated with poor response to anti-epidermal growth factor receptor (anti-EGFR) chemotherapy. However, to date there has been no literature linking biomarkers with stratification of risk for the treatment of rectal cancer following TEMS. Aim: The aim of this dissertation is to assess the significance of these molecular biomarkers in the prediction of local recurrence and prognosis of early rectal cancer following TEMS. Materials and Methods: Initially, we performed a narrative review of the literature to consolidate the evidence available for molecular biomarkers in the evaluation of CRC. We then designed a retrospective pilot study to identify the molecular biomarker status of 41 confirmed CRC cases among 1,446 consecutive referrals for suspected cancer. As part of this study, we retrospectively analysed clinical, biochemical, and histopathological data. Gene profile analysis (KRAS, BRAF) of the specimens was also performed. Following this, we proceeded to analyse data from a series of patients who had undergone TEMS for Stage I rectal cancer at King’s College Hospital (KCH). Demographic, biochemical, histopathological, and follow-up data were prospectively collected. Molecular analysis was prospectively performed in the Advanced Diagnostics Laboratory of KCH to identify the status of BRAF, KRAS, p16 O6-methylguanine-DNA methyltransferase (MGMT), and β-catenin. Finally, we retrospectively collected equivalent data on a 4-year series of 135 confirmed Stage I−IV rectal cancer cases who underwent radical surgery +/- neoadjuvant chemoradiotherapy. Data on the status of the same molecular biomarkers were retrospectively collected. In both cohorts of rectal cancer cases (TEMS/radical surgery +/- additional treatment), the biomarker status was compared with the histopathology and follow-up outcomes, including recurrence and overall cancer-related survival. Results: In our pilot study (41 cases), there was no significant correlation of presenting haemoglobin (Hb) levels with eventual disease staging (p>0.05 for all associations). Patients with right-sided tumours were found to have a lower Hb level than patients with either left-sided or rectal tumours. Hb levels were also significantly lower in patients with the BRAF V600E mutation, although this may be because all 3 patients with the mutation had right-sided tumours. Neither KRAS status nor lymphovascular invasion (LVI) status had a specific correlation with Hb levels. Of 29 specimens of cases who underwent TEMS, there was a statistically significant association between KRAS mutant status and local recurrence (n=6, p=0.037). P16 expression > 5% (mean=10.8%, min=0, max=95) was associated with earlier recurrence within 11.70 months (n=7, p=0.004). Membranous β-catenin expression (n=12, 48%) was also related to KRAS mutant (mt) status (p=0.006) but not to survival (p>0.05). BRAF gene was found to be wild type in all cases tested (n=23). With regard to the specimens of rectal cancer cases who underwent radical excision, 28 cases were Stage I (20.9%), n=30; Stage II (22.4%), n=45; Stage III (33.6%) and n=31 Stage IV (23.1. KRAS mt status was associated with female gender (n=20, p=0.021) and older age (69.62 vs. 62.27, p=0.005). Stage I early cancer subgroup analysis showed that KRAS mt status was associated with distant recurrence of disease (n=4, p=0.045). Conclusions: BRAF V600E mutation seems to be associated with right-sided CRC and iron-deficiency anaemia. This could be used as an adjunct to diagnostic molecular tests for early diagnosis. KRAS, p16, and β-catenin could be used as biomarkers for prediction of local recurrence and stratification of the risk for further surgery in Stage I rectal cancer. Further to this, KRAS may be a predictor of distant recurrence in cases of early stage rectal cancer.

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7

Zanardelli, Sara. "ADAMTS13 : molecular recognition of Von Willebrand factor." Thesis, Imperial College London, 2006. http://hdl.handle.net/10044/1/8241.

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8

Fung, Marion R. "Molecular genetics of blood coagulation factor X." Thesis, University of British Columbia, 1988. http://hdl.handle.net/2429/28783.

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Thirty thousand colonies of a bovine liver cDNA library were screened with a mixture of synthetic oligodeoxyribonucleotides coding for bovine factor X. Five positive colonies were identified, and plasmid DNA was isolated. Cleavage with restriction endonucleases showed that these plasmids (designated pBXl-5) contained inserts of 1530 bp, 770 bp, 700 bp, 1100 bp and 930 bp. DNA sequence analysis of the plasmid with the largest insert (pBXl) confirmed that bovine factor X cDNAs had been cloned. The cDNA sequence predicts that factor X is synthesized as a single chain precursor in which the light and heavy chains of plasma factor X are linked by the dipeptide Arg-Arg. The cDNA sequence also predicts that factor X is synthesized with a preproleader peptide. It is proposed that at least five specific proteolytic events occur during the conversion of preprofactor X to plasma factor Xa. A human liver cDNA library was screened by colony hybridization with a bovine factor X cDNA probe. Three of the positive plasmids contained overlapping DNA that coded for most of human factor X mRNA. A second human liver cDNA library was screened by in situ hybridization with 32P-labeled human factor X cDNA clones obtained from the first screen. Several clones were isolated that contained longer inserts. DNA sequence analysis of these clones allowed the prediction of the amino acid sequence of the precursor form of human plasma factor X. From these studies, it is predicted that human factor X is synthesized as a single polypeptide chain precursor in which the light and heavy chains of plasma factor X are linked by the tripeptide Arg-Lys-Arg. The cDNA sequence also predicts that human factor X is synthesized as a preproprotein having an aminoterminal leader peptide of 40 amino acid residues. A comparison of the amino acid sequences of human and bovine factor X shows high sequence identity around the calcium- binding regions and catalytic regions but low sequence identity around the nonfunctional regions. A human genomic phage library was screened with a human factor X cDNA as a hybridization probe. Thirty-two overlapping phage clones were isolated. Characterization of six of these clones indicates that over 32 Kbp of contiguous sequence is represented. DNA sequence and restriction map analysis shows that the factor X gene is comprised of at least 8 exons and 7 introns. No clones representing the 5' untranslated region and the prepeptide of the leader sequence were identified. Two further genomic phage libraries and two libraries specific for the 5' region of the factor X gene were screened, but no 5' end clones were obtained. Restriction enzyme mapping and Southern blot analysis indicate that thus far, the human factor X gene maps to 24 Kbp of the human genome. Comparison of the factor X gene with other vitamin K-dependent blood coagulation factor genes reveals homologous exon organization. Within the blood coagulation serine proteases factor X, factor IX, factor VII, and protein C form a closely related gene family.
Medicine, Faculty of
Biochemistry and Molecular Biology, Department of
Graduate

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9

Chen, Vivien Mun Yee Medical Sciences Faculty of Medicine UNSW. "The molecular mechanism of tissue factor activation." Awarded by:University of New South Wales. Medical Sciences, 2007. http://handle.unsw.edu.au/1959.4/40556.

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Tissue factor (TF) is the essential cofactor for FVIIa. Binding to transmembrane tissue factor increases the catalytic efficiency of FVIIa allowing activation of FX and FIX which initiates coagulation and propagates stable clot. Transmembrane TF resides in a cryptic configuration on the cell surface and in the circulation with low procoagulant activity. However TF can be rapidly switched to an active configuration in order to contribute to thrombus propagation. The precise nature of this switch is unknown, however it is known to be an extracellular event. The extracellular part of TF consists of 2 fibronectin type III domains. The disulfidebond in the membrane proximal domain (Cys186-Cys209) is a cross-strand bond which links adjacent strands in the same ?? sheet. It has the configuration, characteristic dihedral strain energy and bond length of an allosteric disulfide bond. This indicates that it has the potential to undergo thiol/disulfide exchange to change the function of the TF protein. We confirm that the integrity of the Cys186-Cys209 disulfide is required for coagulant function and that tissue factor contains free thiols in the cryptic state which are lost when TF becomes de-encrypted. Membrane based tissue factor procoagulant activity is blocked by the mono-thiol alkylators N-ethylmaleimide and methyl methanethiosulfonate; but increased by ECl/formation of the disulfide via the thiol oxidiser, HgCb or thiol cross-linkers, eimidohexane and bismalemidoethane. The increase in activity correlates with a conformation change in the TF protein adjacent to the disulfide. We show that redox active protein disulfide isomerase is associated with cryptic tissue factor and propose that the cryptic conformation of tissue factor is maintained through formation of an Snitrosylated complex with protein disulfide isomerase. Our results indicate that the activation of TF involves a change of conformation of the domain 2 of TF caused by formation of the cross-strand Cys186-Cys209 disulfide bond. We suggest that this is likely to be the physiological change that facilitates productive binding of FIX and FX in coagulation.

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10

Scime, Anthony. "Molecular mechanisms regulating the E2F4 transcription factor." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1999. http://www.collectionscanada.ca/obj/s4/f2/dsk1/tape7/PQDD_0031/NQ66234.pdf.

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11

Clarkson, W. D. "Molecular interactions of nuclear transport factor 2." Thesis, University of Cambridge, 1997. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.597746.

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In this Thesis, I describe a molecular dissection of the interaction of NTF2, with Ran and several nuclear pore proteins. To undertake these studies I first cloned and sequenced rat NTF2 cDNA, expressed the protein in bacteria, and purified it to homogeneity. NTF2 was then coupled to Sepharose beads, and used to characterise in detail the molecular interactions of NTF2 with other proteins. NTF2 specifically bound both Ran-GDP and also various repeat-containing nucleoporins, including mammalian p62, and yeast Nsp1p. These interactions were verified by a variety of alternative techniques including the yeast two hybrid screen. Competition experiments indicated that NTF2 has separate binding sites for Ran-GDP and nucleoporins. In addition, I used protein engineering to construct a range of targeted NTF2 mutants based on the known structure of NTF2 to identify the regions of NTF2 involved in these interactions. Although none of the engineered mutant proteins disturbed the nucleoporin binding site on NTF2, several mutants failed to bind Ran-GDP. Using the three dimensional crystal structure of wild-type and two of NTF2 mutants, the key features of the Ran-GDP binding site on NTF2 could be identified. Furthermore, the NTF2 mutants which were unable to bind Ran, were also unable to stimulate nuclear protein import in vitro, and additionally one mutant was found not to be viable in place of the wild type NTF2 gene in the yeast Saccharomyces cerevisiae. Taken together, these findings indicate that the NTF2-Ran interaction is essential for efficient nuclear protein import. Finally, the implications of these and other results for understanding NTF2 function during nuclear protein import are discussed.

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12

Campbell, Susan Christine. "Molecular characterisation of the transcription factor Pax4." Thesis, University of Aberdeen, 2000. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.302462.

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Pax4 is a paired-domain-containing transcription factor that plays a crucial role in the development of pancreatic β- and δ-cells. In the absence of Pax4, no β- or δ-cells develop, but an increase in the number of α-cells is observed. To gain insight into Pax4 function, a rat insulinoma cDNA library was screened and two Pax4-related clones were isolated. One clone encoded a 349 amino acid protein with a molecular weight of 38K that corresponded to the full-length sequence of Pax4. The second cDNA, termed Pax4c, was identical to Pax4 but lacked the sequences encoding 117 amino acids at the COOH-terminus. Intracellular localisation studies indicated that Pax4 was sequestered specifically in the cytoplasm of β-cells. To determine the effect of Pax4 on islet cell gene expression, Pax4 was co-transfected with a series of human insulin and islet amyloid polypeptide (IAPP) promoter constructs into the β-cell line MIN6, and transcriptional activity was measured by reporter gene assay. Pax4 was found to have an inhibitory effect on the human insulin gene promoter, which was mapped, to the region -229 to -258. Electrophoretic mobility shift assay was used to show that Pax4 could bind to the C2 element located at -253 to -244 within this region. Pax4 was also found to have an inhibitory effect on the IAPP promoter, which was mapped to a sequence downstream of -138. Using a GAL4 reporter system, the repressive properties of Pax4 was further mapped to separate regions of the promoter between amino acids 2-230 and 231-349.

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13

Durkan, Garrett Christopher. "Matrix metalloproteinase-1 and -9 and tissue inhibitor of metalloproteinase-1 in bladder cancer : pathophysiological significance and relationship to epidermal growth factor receptor expression." Thesis, University of Newcastle Upon Tyne, 2001. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.369832.

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14

Wilson, William J. "Hypoxia inducible factor 1a : molecular mechanisms of regulation /." Stockholm, 2002. http://diss.kib.ki.se/2002/91-7349-223-X.

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15

Hawkins, Mark G. "Molecular characterization of the human actin depolymerizing factor." Thesis, University of Cambridge, 1992. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.240180.

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16

Green, Simon Richard. "Molecular analysis of eukaryotic initiation factor 2#alpha#." Thesis, University of Kent, 1989. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.330170.

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17

Brown, Andrew James. "Molecular genetic studies of the Prp8 splicing factor." Thesis, University of Edinburgh, 1995. http://hdl.handle.net/1842/15402.

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Prp8 is a component of the U5 and snRNP important for entry of U5 and other snRNPs into the spliceosome, and is present in the active site(s) at the times when the splicing reactions occur. Genes encoding this protein have been isolated from yeast and other eukaryotes and show that Prp8p is extraordinary well conserved, consistent with its multiple roles. This thesis focuses on the only region of yeast Prp8p not known to be common to other eukaryotes: a repetitive acidic and proline-rich domain at the N-terminus. Removal of part or all of this domain inhibits function, but I have found that cells lacking this domain are viable if the truncated protein is overproduced. A reconstruction approach suggests that proline is the most important feature of this domain, and thus function may be analogous to proline-rich regions of other proteins which in general complex assembly. The phenotype of truncation mutants is consistent with this. Spliceosome components are present in the yeast nucleus at much lower concentration than in other eukaryotes, suggesting a reason as to why this otherwise highly conserved protein possesses the extra domain. This thesis also describes the analysis of several mutants of yeast PRP8 which have the highly unexpected phenotype of a block to cell cycle progression. I show that these mutants also affect splicing. The growth defect of one of them (dbf3-1) is suppressed by a cDNA copy of the TUB1 gene. TUB1 contains an intron and encodes a microtubule monomer (α-tubulin) functional in M-phase. Suppression separates the cell cycle and splicing defects, as the splicing defect is unaffected by suppression. These data strongly support the hypothesis that the cell cycle defect is a secondary consequence of a splicing defect, the link being a gene (TUB1) which functions in the cell cycle and which contains an intron. The cell cycle block is observed because the splicing defect is mild: except for the TUB1 intron, splicing is sufficient to maintain growth.

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18

Yeh, Jennifer E. "Molecular Modulators of the Oncogenic Transcription Factor STAT3." Thesis, Harvard University, 2015. http://nrs.harvard.edu/urn-3:HUL.InstRepos:17463967.

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Since the neoplastic phenotype of a cell is largely driven by aberrant gene expression patterns, increasing attention has been focused on transcription factors that regulate critical mediators of tumorigenesis such as signal transducer and activator of transcription 3 (STAT3). Here we investigate how the inappropriate activation of STAT3 contributes to cancer pathogenesis and how it can be targeted therapeutically. As proteins that interact with STAT3 may be key in addressing these questions, we took three complementary approaches: a proteomics approach to identify novel STAT3-interacting proteins, a chemical biology approach to identify STAT3-interacting proteins critical for oncogenesis, and molecular analysis of a STAT3 structural domain known to mediate protein-protein interactions. First, we performed mass spectrometry on STAT3-containing complexes from breast cancer cells that have constitutively active STAT3 and are dependent on STAT3 function. We identified granulin as a novel STAT3-interacting protein that is critical to STAT3 transcriptional activity and STAT3-mediated tumorigenic phenotypes. Furthermore, granulin expression positively correlated with STAT3 gene expression signatures in breast cancer patients. We then applied this mass spectrometry approach to investigate the mechanism of two small molecules – ST3-01 and Pyrimethamine (Pyr) – identified by transcription-based reporter screens to inhibit STAT3 activity without altering its activation or nuclear localization. ST3-01 and Pyr reduced STAT3 interaction with the chromatin remodeler, BRG1, which we found to be necessary for STAT3 function. Next, we studied the role of the STAT3 N-terminal domain (NTD), which mediates interactions between two STAT3 dimers for cooperative DNA binding. We identified STAT3 target genes dependent on the NTD for transcriptional regulation. We then showed that NTD mutations which inhibit cooperative DNA binding reduced the induction of a subset of STAT3 target genes by decreasing STAT3 binding to their regulatory regions. These studies demonstrate that a proteomics approach can reveal critical modulators of transcription factor function. Moreover, our characterization of the impact of the STAT3 NTD on STAT3-dependent activity provides a deeper mechanistic understanding of STAT3 signaling as well as a structural template for drug design. Collectively, these insights into how STAT3 protein-protein interactions modulate its transcriptional function may guide future therapies that target this oncogenic signaling pathway.
Medical Sciences

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19

Schultheis, Anne Maria [Verfasser]. "Angiogenesis Gene Polymorphisms as Molecular Markers to Predict Recurrence in Stage III Colon Cancer / Anne Maria Schultheis." Köln : Deutsche Zentralbibliothek für Medizin, 2011. http://d-nb.info/1009933345/34.

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20

Van, Antwerp Daniel J. "Molecular and biological studies on the regulation of transcription factor nuclear factor-kappa B /." Diss., Connect to a 24 p. preview or request complete full text in PDF format. Access restricted to UC campuses, 1999. http://wwwlib.umi.com/cr/ucsd/fullcit?p9932588.

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21

Portman, Jonathan Lewis. "Virulence Factor Regulation in Listeria monocytogenes." Thesis, University of California, Berkeley, 2018. http://pqdtopen.proquest.com/#viewpdf?dispub=10620349.

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Listeria monocytogenes is a Gram-positive intracellular pathogen that is readily amenable to genetic manipulation and for which there are excellent in vitro and in vivo virulence models. These attributes have allowed a thorough examination of the molecular underpinnings of L. monocytogenes pathogenesis, however, there are still a number of major unresolved questions that remain to be answered. For example, it has been known for many years that L. monocytogenes rapidly changes its transcriptional profile upon access to the host cytosol, however the host cues and bacterial components that are involved in driving this change have remained continually unanswered. One large piece of evidence came when the long-sought co-factor for the primary virulence regulator, PrfA, was discovered to be the antioxidant tripeptide, glutathione. Glutathione was demonstrated to play a crucial role in the activation of PrfA in vivo— a finding that has since led to two important discoveries that are described herein. First, the activation of PrfA in vitro requires both exogenous glutathione and a metabolic licensing step that can be recapitulated by a chemically defined synthetic media. Second, glutathione also functions as a post-translational regulator of the pore-forming virulence factor, Listeriolysin O (LLO), by reversibly binding via an S-glutathionylation reaction and preventing membrane association of the LLO monomers. These discoveries elucidate numerous regulatory roles for glutathione during infection and describe how L. monocytogenes is able to sense and respond to critical host compartments to mount a successful infection.

Upon entry to the host cell cytosol, the facultative intracellular pathogen Listeria monocytogenes coordinates the expression of numerous essential virulence factors by allosteric binding of glutathione (GSH) to the Crp-Fnr family transcriptional regulator, PrfA. Here we report that robust virulence gene expression can be recapitulated by growing bacteria in a synthetic medium (iLSM) containing GSH or other chemical reducing agents. Bacteria grown under these conditions were 45-fold more virulent in an acute murine infection model and conferred greater immunity to a subsequent lethal challenge compared to bacteria grown in conventional media. During cultivation in vitro , PrfA activation was completely dependent on intracellular levels of GSH, as a glutathione synthase mutant (ΔgshF) was activated by exogenous GSH but not reducing agents. PrfA activation was repressed in iLSM supplemented with oligopeptides, but suppression was relieved by stimulation of the stringent response. These data suggest that cytosolic L. monocytogenes interpret a combination of metabolic and redox cues as a signal to initiate robust virulence gene expression in vivo.

Cholesterol-dependent cytolysins (CDCs) represent a family of homologous pore-forming proteins secreted by many Gram-positive bacterial pathogens. CDCs mediate membrane binding partly through a conserved C-terminal undecapeptide, which contains a single cysteine residue. While mutational changes to other residues in the undecapeptide typically have severe effects, mutating the cysteine residue to alanine has minor effects on overall protein function. Thus, the function of this highly conserved reactive cysteine residue remains largely unknown. We report here that the CDC Listeriolysin O (LLO), secreted by the facultative intracellular pathogen Listeria monocytogenes, was post-translationally modified by a S-glutathionylation at this conserved cysteine residue, and that either endogenously synthesized or exogenously added glutathione was sufficient to form this modification. When recapitulated with purified protein in vitro, this modification completely ablated the activity of LLO, and this inhibitory effect was fully reversible by treatment with reducing agents. A cysteine-to-alanine mutation in LLO rendered the protein completely resistant to inactivation by S-glutathionylation and retained full hemolytic activity. A mutant strain of L. monocytogenes expressing the cysteine-to-alanine variant of LLO was able to infect and replicate within bone marrow-derived macrophages indistinguishably from wild-type in vitro, yet was attenuated 4-6 fold in a competitive murine infection model in vivo. This study suggests that S-glutathionylation may represent a mechanism by which CDC family proteins are post-translationally modified and regulated, and help explain an evolutionary pressure behind the highly conserved undecapeptide cysteine.

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22

Youseff, Brian. "The Role of Tumor Necrosis Factor Receptor-Associated Factor 6 in Tick-Borne Flavivirus Infection." University of Toledo Health Science Campus / OhioLINK, 2019. http://rave.ohiolink.edu/etdc/view?acc_num=mco155691388498993.

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23

Bonin, Fanny. "Cytoprotective effects of intracellular platelet activating factor acetylhydrolases." Thesis, University of Ottawa (Canada), 2003. http://hdl.handle.net/10393/26529.

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Platelet activating factor (PAF) is a biologically active phospholipid implicated in the developmental brain disorder Miller-Dieker Syndrome (MDS) and purported to be a primary mediator of cell death in HIV-dementia, ischemia, and epilepsy. As part of my honour's thesis, I demonstrated that PAF can elicit cell death independently of its G-protein coupled receptor (PAFR) in PC12 cells. In my M.Sc. research, I have sought to identify how PAF-mediated cell death is regulated in PC12 cells. PAF is inactivated in brain by two intracellular PAF-acetylhydrolases (PAF-AHs): PAF-AH I and PAF-AH II. PAF-AH I is a trimeric complex composed of two catalytic subunits (alpha1 and alpha2) and one regulatory subunit (beta). Mutations in the Lis1 gene, coding for the beta subunit of PAF-AH I, are the genetic determinant of MDS. However, it is not clear whether these mutations impact on PAF-AH I enzymatic activity in MDS. Furthermore, it is not known whether cytosolic PAF-AH activity regulates the kinetics of neuronal loss following pathophysiological challenge. To begin to address these questions, I sought to identify an in vitro model system suitable for study of PAF-AH activity.* (Abstract shortened by UMI.) *This dissertation is a compound document (contains both a paper copy and a CD as part of the dissertation). The CD requires the following system requirements: QuickTime.

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24

Fung, Hiu Leong. "Human C7orf30 is a novel mitochondrial translation factor." Thesis, McGill University, 2011. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=103744.

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Mitochondria generate the majority of cellular energy through oxidative phosphorlyation. The machinery of oxidative phosphroylation consists of five enzyme complexes that are located in the inner mitochondrial membrane. A small number of essential subunits in these complexes are encoded by mtDNA and synthesized on a dedicated mitochondrial translation apparatus. Defects in mitochondrial translation system cause many mitochondrial diseases, but the mechanisms that regulate mitochondrial translation remain largely unknown. We have identified an unnamed human protein C7orf30, as a possible mitochondrial translation factor. The orthologue of C7orf30 in maize is thought to be a chloroplast ribosome assembly factor. We identified human C7orf30 to be a mitochondrial protein through bioinformatics analysis and immunocytochemistry. We knocked down the expression of C7orf30 in human fibroblast using shRNA and observed a reduction in cytochrome c oxidase activity. Using a translation assay, we observed a global reduction in the synthesis of mitochondrially encoded proteins when C7orf30 was knocked down, while the transcript levels were not affected. The assembly of Complex I, III, IV and V also demonstrated defects. Sucrose density gradient analysis suggests C7orf30 interacts with 39S subunit of the mitoribosome. The assembly of the mitoribosome and the levels of 12S and 16S MT-rRNA were not affected by C7orf30 knockdown, suggesting C7orf30 is not necessary for mitochondrial ribosome assembly. We hypothesize that C7orf30 interacts with the mitoribosome and is a regulator of mitochondrial translation.
Les mitochondries génèrent la majorité de l'énergie cellulaire grâce à l'oxydation phosphorylative. La chaîne respiratoire responsable de ce phénomène est composée de cinq complexes enzymatiques localisés dans la membrane interne de la mitochondrie. Certaines des sous-unités essentielles de ces complexes sont codées par l'ADN mitochondrial. Leur synthèse est assurée par la mitochondrie qui possède son propre système de traduction des protéines. Les déficiences de la traduction mitochondriale sont à l'origine de nombreuses maladies et les mécanismes qui régulent le processus de traduction restent à ce jour peu élucidés. Dans cette étude, nous avons identifié chez l'homme, C7orf30, une protéine probablement impliquée dans la régulation de la traduction mitochondriale. Il existe un homologue de cette protéine chez le maïs. Une étude suggère son rôle en tant que facteur d'assemblage des ribosomes des chloroplastes. Des programmes informatiques prédisent la localisation de la protéine C7orf30 humaine dans la mitochondrie ce que nous avons confirmé par des expériences d'immunocytochimie. L'utilisation de shRNA dirigés contre C7orf30 dans des fibroblastes humains révéle d'abord une réduction de l'activité cytochrome c oxydase (complexe IV). Des expériences de traduction ex vivo montrent ensuite une réduction globale de la synthèse des protéines codées par la mitochondrie dans les cellules déficitaires en C7orf30, la transcription étant normale. L'assemblage des complexes I, III, IV et V de la chaîne respiratoire est également affecté. La séparation des protéines par gradient de sucrose suggère que C7orf30 interagit avec la sous unité 39S des ribosomes mitochondriaux. Cependant, l'assemblage et les niveaux d'expression des rRNA 12S et 16S ne sont pas affectés par la diminution de la protéine ce qui suggère qu'elle n'est pas indispensable à l'assemblage des ribosomes mitochondriaux en soit. Dans cette étude, nous émettons l'hypothèse que C7orf30 est un composant du ribosome et agit comme un régulateur de la traduction mitochondriale.

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25

Uprichard, William James Nicolas. "Molecular and structural analysis of human Factor X deficiency." Thesis, University College London (University of London), 2005. http://discovery.ucl.ac.uk/1446783/.

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Factor X (FX) is one of the vitamin K-dependent serine proteases, and is g of a catalytic serine protease domain, two EGF domains and a Gla domain. It is crucial for coagulation, however its role in disease has not been fully characterised. This thesis describes the phenotypic and genotypic analysis of kindred with FX deficiency. Because FX deficiency is rare in the general population, patient samples were drawn from across the world. Laboratory assays for FX and direct sequencing of the gene were performed on samples from ten kindred. A total of ten mutations were identified, nine of which were novel and summarised as follows. One mutation was identified in two unrelated families (Asp373Asn). Compound heterozygous mutations were identified in two kindred (Glul9Val and IVS5+3 A-G in one kindred; Cysl32Stop and Arg273Trp in the other). Two other mutations were identified in the catalytic domain (Gly222Asp and Pro382Leu), one in the EGF-1 domain (Glu51Lys), and one in the signal peptide (Ala-26Asp). No mutation was identified in one kindred. Molecular modelling of mutations in terms of the available crystal structure of factor Xa was performed in order to correlate genotype with phenotype. Explanations were proposed in terms of the perturbation of the structure itself or its biochemical function. For Glu51Lys and Arg273Trp, it was predicted that ligand binding would be disrupted. In the case of Asp373Asn, it was proposed that perturbation of the FX sodium-binding site would impair its enzymatic function. It was predicted that the Gly222Asp and Pro382Leu mutations would disrupt protein folding. In order to show that selected known mutations are causative for FX deficiency, recombinant FX with the Arg-1Thr, Glu51Lys, Arg273Trp and Asp373Asn substitutions was expressed. Their expression was successful. In preliminary work with these, it was not possible to recapitulate the in vivo data for these expressed proteins.

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26

Tonissen, Kathryn Fay. "A molecular study of the early pregnancy factor phenomenon /." Title page, table of contents and summary only, 1990. http://web4.library.adelaide.edu.au/theses/09PH/09pht6651.pdf.

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27

Gile, Gillian Heather. "Molecular evolution of the eukaryotic translation elongation factor, EFL." Thesis, University of British Columbia, 2009. http://hdl.handle.net/2429/11588.

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The eukaryotic translation elongation factor EFL (for EF-Like) is a paralogue of the better-known elongation factor 1-alpha (EF-1α), which brings aminoacyl-tRNAs to the ribosome during translation. This essential protein was thought to be ubiquitous in eukaryotes until the recent discovery of EFL in a small number of diverse, mainly unicellular, eukaryotic organisms that were found to lack EF-1α. Because of the great evolutionary distances between EFL-encoding lineages and the near mutual exclusivity of the two proteins, the observed complex distribution of EFL was initially attributed entirely to multiple lateral gene transfers. In the enclosed chapters, the distribution of EFL was characterized in more detail in four distantly related eukaryotic lineages at both fine and broad taxonomic scales in order to better understand the effects that endosymbiotic gene transfer, differential loss, and lateral gene transfer have had on the molecular evolution of EFL. Endosymbiotic transfer of EFL was detected in the chlorarachniophytes, a group of algae whose secondary plastids retain a vestigial nucleus, known as a nucleomorph, in their reduced eukaryotic cytoplasm, known as the periplastid compartment (PPC). The endosymbiotically transferred EFL carries a bipartite targeting sequence similar to those of plastid-targeted proteins in this group and to plastid- and PPC-targeting sequences in cryptomonads to direct it to the PPC, suggesting similarities in the way these two lineages have solved their shared challenge of targeting to complex plastids with nucleomorphs. No clear phylogenetic evidence for lateral transfer of EFL has yet emerged; rather, differential loss of EFL and EF-1α from an ancestral state of co-occurrence was characterized in euglenozoans and detected in publicly available data from heterokonts and opisthokonts, unexpectedly revealing a significant role for this process in shaping the complex distribution of EFL and EF-1α. This finding serves as a cautionary reminder that adequate taxon sampling and a robust organismal phylogenetic hypothesis are crucial in order to correctly infer lateral gene transfer.

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28

Smurthwaite, Lyn. "Molecular studies on a putative human mitochondrial elongation factor." Thesis, Imperial College London, 1999. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.321998.

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29

McCullough, Peter W. "Hepatocyte growth factor : Molecular Mechanisms of Colorectal Cancer Metastasis." Thesis, Imperial College London, 2009. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.508939.

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30

Barhoover, Melissa. "Molecular Mechanism of Incorporation of Factor Va into Prothrombinase." Cleveland State University / OhioLINK, 2007. http://rave.ohiolink.edu/etdc/view?acc_num=csu1197570167.

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31

CARUSO, Pierpaolo. "Molecular bases of the modulation of coagulation factor levels." Doctoral thesis, Università degli studi di Ferrara, 2009. http://hdl.handle.net/11392/2389194.

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The maintenance of blood uidity within the vascular system is ensured by the balance between procoagulant and anticoagulant components. Hemostasis and blood coagulation are processes regulated by a complex network of interactions, involving plasma/tissue proteins and cells. Numerous epidemiologic studies indicate that deciencies in blood coagulation proteins cause serious bleeding problems, whereas the deciencies in coagulation inhibitors and certain mutations in coagulation factors may lead to thrombotic disorders. The coagulation process has been conceptualized as being dependent primarily on adequate levels of the coagulation proteins. From clinical studies it is assumed that the normal concentration range of proteins involved in blood coagulation and regulation of this process may vary in the average blood sample from 50% to 150% of their mean plasma values. This suggests that a wide range of factor levels is compatible with normal hemostatic function; however, even within the normal range, variations can aect the rate and extent of thrombin generation. As a consequence, the stimulus/response which follows tissue factor presentation to blood and the subsequent expression of thrombin activity is highly variable even in the normal population. In addition, the qualitative performance of these proteins is governed by many molecular events which are in uenced both by genetic background, and by environmental processes that alter coagulation proteins during circulation and may produce very dierent phenotypic responses. All these considerations emphasize the importance in measuring and monitoring variations of coagulation factor levels, providing potential correlations with non physiologic conditions. Also, the evidence of how the coagulation proteins are involved in the acute phase of in ammation and several other human diseases, confirm that analysis of their level changes and modulation, represent the way required for appropriate diagnosis and therapeutic intervention. Aims A number of molecular mechanisms intragenic, extragenic and environmental, acting at different levels of the gene information ow, could produce variations in the expression of coagulation protein or activity levels. To get inside this complex regulative network we report four different studies aimed at elucidate aspects of clotting factor modulation. Study of oscillations in coagulation factor levels, due to inherited or environmental factor, could establish novel relationships between coagulation and in ammatory components, or with determinant of other human pathologies. Methods Methods are included in articles published in 2007/2008; they are focused on thrombin generation assay, an overall assay measuring the amount of thrombin produced in plasma samples in standard condition. This method evidences subjects coagulation-phenotype, allowing the correlation with other clotting factors levels. Main Results The combined effect of ten common prothrombotic polymorphisms as a determinant of MI was tested in a population of 804 subject, half of whom affected by severe Coronary Artery Disease (CAD). The number of procoagulant alleles was signicantly associated with the Endogenous Thrombin Potential (ETP), similarly, subjects with a high number of procoagulant alleles had significantly higher ETP values as compared to subjects with fewer alleles. Similar approach was used to investigate a rare bleeding disorder, vitamin K-dependent clotting factor deficiency type 2 (VKCFD2), which offered several quantitative parameters providing us information on vitamin K-dependent regulation of plasma factor levels; a markedly variation in Thrombin Generation parameters and others clotting factors measurements improved the identification of a clear coagulation phenotype. We also probed the effects of an integrated healthy diet on a wide panel of hemostatic and in ammatory variables. The changes observed in coagulation initiation and amplification phases, body composition and lipid profile could translate into a remarkable decrease in the risk for cardiovascular disease, suggesting novel relationship between coagulation and in ammation components. Finally we investigate how Tissue Factor-bearing Microparticles was generated by monocyte-derived Dendritic Cells stimulated with the P2X7R ag- onist benzoyl ATP (BzATP); our observations identify a novel pathway for tissue factor release, providing new insights into the mechanisms underlying the generation and spreading of procoagulant activity. The activity of this potential circulating trigger of coagulation and in ammation stimuli could be involved in progression of several pathology and in ammation process. Conclusions Our studies provide quantitative evidence for genetic and environmental determinants of coagulation factor levels in plasma, and for their integrated eects on generation of thrombin. The consequent variation of coagulation phenotype produces ample temporal and individual variations in coagulation function. These ndings have implications on the diagnosis, prevention/prophylaxis and therapy of coagulation diseases, and encourage further investigations to better understand the clinical signicance of the molecular heterogeneity of the human hemostatic proteome.

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32

Ito, Masaaki. "P-21 activated kinase 1 : a new molecular marker for intravesical recurrence after transurethral resection of bladder cancer." Kyoto University, 2008. http://hdl.handle.net/2433/135792.

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33

Kabbani, Nazir. "Chemical-genetic profiling of platelet-activating factor in yeast." Thesis, University of Ottawa (Canada), 2009. http://hdl.handle.net/10393/28189.

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The basic biological processes between the yeast Saccharomyces cerevisiae and mammals are highly conserved. Yeast posses many genes that are implicated in human diseases and have been successfully used as a model for the study of neurodegeneration. Platelet-Activating Factor (C16:0 PAF) causes neuronal cell death independent of its receptor and has been implicated in Alzheimer's disease. I hypothesized that yeast could be used as a model system for deciphering PAF receptor-independent signalling and have utilized genome-wide chemical genomic screening in yeast to further characterize the molecular mechanism of PAF toxicity. Two complementary screens implicate PAF in many cellular processes, some of which parallel results obtained in mammalian studies. I have found that PAF challenge is cytotoxic, delays cell cycle progression, and affects actin stability leading to spindle misorientation and bi-nucleate mother cells.

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34

Burnette, Ethan Williams. "Endothelium-derived hyperpolarizing factor (EDHF) in rat mesenteric artery." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 2001. http://www.collectionscanada.ca/obj/s4/f2/dsk3/ftp05/NQ66130.pdf.

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35

Moutinho, Cátia Alexandra Martins de Freitas. "Epidermal growth factor receptor in gastric cancer. Receptor do factor de crescimento epidérmico no cancro do estômago." Master's thesis, Faculdade de Medicina da Universidade do Porto, 2007. http://hdl.handle.net/10216/22117.

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36

Moutinho, Cátia Alexandra Martins de Freitas. "Epidermal growth factor receptor in gastric cancer. Receptor do factor de crescimento epidérmico no cancro do estômago." Dissertação, Faculdade de Medicina da Universidade do Porto, 2007. http://hdl.handle.net/10216/22117.

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37

Subang, Maria Cristina. "The regulation of ciliary neurotrophic factor, leukemia inhibitory factor and monocyte chemoattractant protein-1 in injured peripheral nervous tissue." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 2000. http://www.collectionscanada.ca/obj/s4/f2/dsk1/tape2/PQDD_0034/NQ64675.pdf.

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38

Hedlund, Eva-Maria. "Molecular mechanisms of angiogenic synergism between Fibroblast Growth Factor-2 and Platelet Derived Growth Factor-BB." Thesis, Södertörn University College, School of Life Sciences, 2006. http://urn.kb.se/resolve?urn=urn:nbn:se:sh:diva-932.

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39

Österlund, Maria. "Molecular probing of local protein-protein interactions : studies of the tissue factor, factor VIIa complex formation /." Linköping : Univ, 2001. http://www.bibl.liu.se/liupubl/disp/disp2001/tek670s.pdf.

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40

Pacheco, Ryan John. "Characterication of aggregate gland silk factor 1." Scholarly Commons, 2014. https://scholarlycommons.pacific.edu/uop_etds/186.

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Spider silk is a high performance fiber with extraordinary mechanical properties, including high tensile strength and toughness. Due to these outstanding material properties, scientists are rapidly pursuing the production of synthetic spider silks for a variety of different applications. In these studies, we characterize the aggregate gland specific factor 1 (AgSF1) from the black widow spider, Latrodectus hesperus. After the development of an anti-AgSF1 polyclonal antiserum, we demonstrate by western blot analyses that the AgSF1 protein is highly expressed in the aggregate gland and the protein is localized to the connection joints of cobweaver webs. We also overexpress and purify two different recombinant AgSF1 fusion proteins, named AgSF1G and AgSF1G+GXPXP. These recombinant proteins encompass different regions within the AgSF1 amino acid sequence. Using wet-spinning methodology we also demonstrate that these proteins can be spun into synthetic silk fibers. Mechanical studies and ultrastructure analyses of the synthetic fibers reveal tensile strengths and toughness values that are below natural dragline silk fibers. Secondary structural analyses of the AgSF1 recombinant proteins in solution using circular dichoism reveal the N-terminal region of AgSF1 is alpha helical in nature. Collectively, these studies advance our understanding of silk proteins that are expressed in the aggregate gland and support that these proteins play an important role in prey capture in cobweavers.

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41

McGuffie, Bryan A. "A sigma factor and anti-sigma factor that control swarming motility and biofilm formation in Pseudomonas aeruginosa." Thesis, Harvard University, 2015. http://nrs.harvard.edu/urn-3:HUL.InstRepos:17467530.

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Pseudomonas aeruginosa is an environmental bacterium and opportunistic human pathogen of major clinical significance. It is the principal cause of morbidity and mortality in patients with cystic fibrosis (CF) and a leading cause of nosocomial infections. Although the organism is unicellular, P. aeruginosa exhibits two forms of multicellular behaviors when associated with a surface under the right conditions: swarming motility and biofilm formation. Swarming motility is a multicellular cooperative form of flagella-dependent surface motility, while biofilm formation produces a sessile community of bacteria enclosed by a self-produced extracellular polymeric matrix. P. aeruginosa is thought to grow as a biofilm in the lungs of CF patients and the growth of P. aeruginosa biofilms on indwelling medical devices, such as endotracheal tubes and catheters, is a significant source of nosocomial infection. By growing as a biofilm, P. aeruginosa resists clearance by the immune system and increases its resistance to antimicrobial therapy. In this thesis, I describe the characterization of a sigma factor and anti-sigma factor implicated in P. aeruginosa virulence and cell envelope stress that control the expression of a novel regulator of swarming motility and biofilm formation. In addition, I describe work done to investigate the role of a post-translational regulator of flagellar motility that has been described in other bacteria, but has not been studied extensively P. aeruginosa. In bacteria, RNA polymerase (RNAP) requires sigma factors for promoter-specific transcription initiation. sigma factors guide RNAP to promoters by recognizing conserved DNA sequences within the promoter called the -10 and -35 elements. Most bacteria encode a primary sigma factor and several alternative sigma factors, each of which recognizes different promoter -10 and -35 sequences. By modulating the activity of alternative sigma factors, bacteria can rapidly alter their transcriptional program in response to changes in growth, morphological development, and environmental conditions. The extracytoplasmic function (ECF) sigma factors are the largest and most diverse group of bacterial sigma factors. The gene encoding an ECF sigma factor is often cotranscribed with its own negative regulator, called an anti-sigma factor, which directly binds to and inhibits its partner sigma factor until the appropriate extracytoplasmic signal stimulates sigma factor release and expression of the sigma factor’s regulon. In this thesis, I describe the characterization of the P. aeruginosa ECF sigma factor PA2896 and its cognate anti-sigma factor PA2895. Using immunoprecipitation, we show that the ECF sigma factor PA2896 and RNAP co-purify in vivo. Utilizing DNA microarrays, we identify the genes that constitute the PA2895 and PA2896 regulon and infer the putative -10 and -35 consensus sequences recognized by PA2896. Genetic analysis revealed a subset of genes within the PA2896 regulon that share the putative promoter consensus sequence are positively regulated by the ECF sigma factor PA2896 and negatively regulated by the anti-sigma factor PA2895. Using a bacterial two-hybrid assay, we show that PA2895 directly interacts with PA2896. We present evidence that increased expression of the PA2896 regulon in ∆PA2895 mutants cells leads to the inhibition of swarming motility and enhanced biofilm formation. We further show that one gene in the PA2896 regulon, PA1494, is necessary and sufficient for the inhibition of swarming motility and promotion of biofilm formation. Thus we report the discovery of a system that may respond to a stress signal by activating PA2896-dependent expression of PA1494 to inhibit swarming motility and promote the formation of a protective biofilm. In many bacteria, swarming motility and biofilm formation are controlled by the second messenger c-di-GMP. In P. aeruginosa, elevated intracellular c-di-GMP generally results in the inhibition of flagellar-dependent swarming motility and enhanced biofilm formation. In some species of bacteria, c-di-GMP-mediated repression of flagellar motility is achieved by repressing the expression of the flagellar genes. However, flagellar gene expression in P. aeruginosa does not appear to be influenced by elevated c-di-GMP, suggesting c-di-GMP controls flagellar motility post transcriptionally in this bacterium. Mechanisms by which c-di-GMP controls flagellar function post translationally have been described in both Gram-negative and Gram-positive bacteria, however the mechanism in P. aeruginosa remains unclear. Gram-negative bacteria appear to utilize a “flagellar brake” to control flagellar function in response to c-di-GMP. P. aeruginosa encodes a homolog of this brake and in this thesis I present evidence that this homolog is involved in controlling flagellar function in P. aeruginosa. Together, the characterization of PA2895 and PA2896, the identification of PA1494 as a novel regulator of swarming motility and biofilm formation, and evidence of a functional flagellar brake in P. aeruginosa advance our understanding of how this bacterium controls the transition from motile cell to sessile biofilm.
Medical Sciences

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42

Poleo, Camejo German Antonio. "Fibroblast growth factor 8 and cell proliferation in zebrafish fins." Thesis, University of Ottawa (Canada), 1998. http://hdl.handle.net/10393/4546.

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The fins of some teleost fishes, like many urodele amphibian limbs, have the ability to regenerate after injury. This regeneration process occurs in a sequential manner beginning with the closure of the wound by epithelial cells and followed by the establishment of a region of undifferentiated proliferating mesenchymal cells called the blastema. To examine the pattern of cell proliferation taking place in the different tissues during the regeneration process of zebrafish (Danio rerio) caudal fins, I followed the pattern of incorporation of bromodeoxyuridine (BrdU) in the DNA of proliferating cells at various times after amputation. Growing evidence suggests that key molecules involved in signaling pathways leading to limb bud development are also acting during fin bud development. Some of these signals are members of the fibroblast growth factor (Fgf) gene family which locally regulate growth and patterning in vertebrate embryos and during limb regeneration. I sequenced a cDNA clone encoding for the zebrafish FGF8 protein and analyzed its pattern of expression during zebrafish embryonic development and during fin regeneration. (Abstract shortened by UMI.)

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43

Pause, Arnim. "Mutational analysis of the mammalian translation initiation factor eIF-4A." Thesis, McGill University, 1994. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=41744.

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eIF-4A is a eukaryotic translation initiation factor and DEAD box RNA helicase that is thought to be responsible for the melting of secondary structure in the 5$ sp prime$ untranslated region of messenger RNAs to facilitate ribosome binding. A mutational analysis of eIF-4A revealed that the ATPase A motif (AXXXXGKT) is involved in ATP binding, the ATPase B motif (DEAD) is implicated in ATP hydrolysis, the SAT region is essential for RNA unwinding, and the HRIGRXXR region is required for ATP hydrolysis-dependent RNA binding. Furthermore, defective eIF-4A mutants exhibit a strong dominant negative effect on in vitro translation of several mRNAs, including those translated by a cap-independent internal initiation mechanism. It is demonstrated that eIF-4A functions primarily as a subunit of eIF-4F, and singular eIF-4A is required to recycle through the eIF-4F during translation. Accordingly, eIF-4F appears to be required for cap-dependent and internal initiation of translation.

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44

Lavallée, François. "Chromatin structure of the atrial natriuretic factor gene." Thesis, McGill University, 1990. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=60460.

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Atrial natriuretic factor (ANF) is a 28 amino acid hormone which is secreted by the heart and which appears to a play a role in blood pressure regulation and blood volume homeostasis. The role of ANF in the control of blood pressure prompted several investigations in relation with the regulation of its gene expression. The work presented in this thesis describes the putative localization of some regulatory elements of the rat ANF gene. These elements were investigated by chromation hypersensitivity and methylation studies of the promoter region spanning the first 3.5 kb upstream of the transcription initiation site.
Preliminary data suggested that there might be DNAse hypersensitive site(s) at two positions ($-$2.5 kb and $-$0.5 kb) in the promoter. These positions are consistent with other experiments performed in our laboratory.
Digestion with several methylation-sensitive restriction endonucleases suggested some tissue-specific differences in the methylation status of HpaII, SaII, SnaBI and HhaI restriction sites in the ANF gene when comparing expressing and non-expressing tissues.
Taken together, the hypersensitivity and methylation studies have identified sequences in the 5$ sp prime$ region of the rat ANF gene as being putative target for regulatory proteins.

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45

Mohammed, Hesham Hamada Taha. "Molecular analysis of adenylyl cyclase : bacillus anthracis edema factor exotoxin." kostenfrei, 2009. http://www.opus-bayern.de/uni-regensburg/volltexte/2010/1411/.

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46

Reinhardt, Christoph. "Analyses on molecular mechanisms of activation of intravascular Tissue Factor." Diss., lmu, 2007. http://nbn-resolving.de/urn:nbn:de:bvb:19-77216.

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47

Lowry, Jason Allen. "Molecular and evolutionary analysis of the GATA transcription factor family." NCSU, 2002. http://www.lib.ncsu.edu/theses/available/etd-11272002-123604/.

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The objective of this research has been to characterize the evolution of the GATA family of transcription factors through phylogenetic, molecular, and biochemical analyses. From a phylogenetic perspective, we address three major questions. First, does this protein family represent a monophyletic or polyphyletic group? Second, what methods of gene or modular duplication are utilized within different organisms to propagate and maintain this group of proteins? Third, what are the structural or functional constraints on evolution of the conserved DNA-binding domain? These questions are addressed through a combination of computational and molecular methods. Phylogenetic analyses provide evidence of monophyletic origin for the GATA family followed by gene duplication and modular evolution, accompanied by considerable divergence outside the conserved zinc finger DNA-binding domain. Genomic comparisons permit the tracing of GATA factor evolution and provide insight into mechanisms utilized by respective organisms. Finally, mutational and biochemical analyses enable the separation of phylogenetic and structural/functional constraints on the conserved zinc finger DNA-binding domain. The result of this research is a predictive motif for classifying potentially homologous proteins to be discovered in future studies.

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48

Bibb, Lindsay Claire. "Molecular studies on the cone-rod homeobox (CRX) transcription factor." Thesis, Imperial College London, 2003. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.404407.

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49

Davis, Jason A. "Molecular studies of epidermal growth factor like domains in CRB1." Thesis, University of Oxford, 2005. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.413077.

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50

Stratford, Anna. "Molecular mechanisms of the PTTG Binding Factor (PBF) in tumourigenesis." Thesis, University of Birmingham, 2005. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.433637.

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