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The role of the mesenchyme homeobox genes in the regulation of vascular endothelial cell functionNorthcott, Josette M. D. 10 December 2011 (has links)
The mesenchyme homeobox genes, MEOX1 and MEOX2, encode homeodomain transcription factors. Studies of Meox1/Meox2 knockout mice established that these proteins are partially redundant during development, suggesting that they may regulate common target genes. In the adult vasculature, MEOX2 is expressed in vascular smooth muscle and endothelial cells. MEOX2 has been demonstrated to: i) inhibit proliferation, ii) activate apoptosis and iii) induce senescence. In contrast, the role of MEOX1 has not been studied in the vasculature. Currently, there are two known target genes of MEOX2: cyclin-dependent kinase inhibitor 1A (CDKN1A/p21CIP1/WAF1) and cyclin-dependent kinase inhibitor 2A (CDKN2A/p16INK4a), which regulate transient (quiescent) and permanent (senescent) cell cycle arrest. Senescence is postulated to contribute to the development of atherosclerotic vascular disease by promoting endothelial dysfunction.
We hypothesized that MEOX1 and MEOX2 would activate both p21CIP1/WAF1 and p16INK4a expression, as well as induce apoptosis, cell cycle arrest and senescence in endothelial cells. Furthermore, we postulated that the majority of newly identified MEOX target genes in endothelial cells would be regulated by both MEOX1 and MEOX2.
MEOX proteins were expressed in human endothelial cells via adenoviral transduction. Levels of target gene expression were measured by luciferase reporter gene assays, western blot and quantitative real-time PCR. Electrophoretic mobility shift assays were used to demonstrate MEOX binding to DNA. Cellular proliferation, senescence, and apoptosis were evaluated. For the identification of novel target genes, microarrays were used to compare levels of gene expression in endothelial cells transduced with MEOX constructs or control virus.
Both MEOX1 and MEOX2 activated p21CIP1/WAF1 and p16INK4a gene transcription, inhibited proliferation and induced apoptosis and senescence in endothelial cells. MEOX activation of p21CIP1/WAF1 transcription occurs via a DNA-binding independent mechanism that requires the SP1 transcription factor. In contrast, MEOX activation of p16INK4a transcription is dependent upon DNA-binding. Microarray analysis revealed that both MEOX1 and MEOX2 increased the expression of intercellular adhesion molecule 1 (ICAM-1) and decreased the expression of nitric oxide synthase 3 (NOS3/eNOS).
Taken together, we conclude that MEOX1 and MEOX2 have similar target genes in endothelial cells including p21CIP1/WAF1, p16INK4a and eNOS. As increased endothelial senescence and decreased nitric oxide production are hallmarks of endothelial dysfunction, this study proposes a role for the MEOX proteins in the progression of atherosclerotic vascular disease.
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The role of the mesenchyme homeobox genes in the regulation of vascular endothelial cell functionNorthcott, Josette M. D. 10 December 2011 (has links)
The mesenchyme homeobox genes, MEOX1 and MEOX2, encode homeodomain transcription factors. Studies of Meox1/Meox2 knockout mice established that these proteins are partially redundant during development, suggesting that they may regulate common target genes. In the adult vasculature, MEOX2 is expressed in vascular smooth muscle and endothelial cells. MEOX2 has been demonstrated to: i) inhibit proliferation, ii) activate apoptosis and iii) induce senescence. In contrast, the role of MEOX1 has not been studied in the vasculature. Currently, there are two known target genes of MEOX2: cyclin-dependent kinase inhibitor 1A (CDKN1A/p21CIP1/WAF1) and cyclin-dependent kinase inhibitor 2A (CDKN2A/p16INK4a), which regulate transient (quiescent) and permanent (senescent) cell cycle arrest. Senescence is postulated to contribute to the development of atherosclerotic vascular disease by promoting endothelial dysfunction.
We hypothesized that MEOX1 and MEOX2 would activate both p21CIP1/WAF1 and p16INK4a expression, as well as induce apoptosis, cell cycle arrest and senescence in endothelial cells. Furthermore, we postulated that the majority of newly identified MEOX target genes in endothelial cells would be regulated by both MEOX1 and MEOX2.
MEOX proteins were expressed in human endothelial cells via adenoviral transduction. Levels of target gene expression were measured by luciferase reporter gene assays, western blot and quantitative real-time PCR. Electrophoretic mobility shift assays were used to demonstrate MEOX binding to DNA. Cellular proliferation, senescence, and apoptosis were evaluated. For the identification of novel target genes, microarrays were used to compare levels of gene expression in endothelial cells transduced with MEOX constructs or control virus.
Both MEOX1 and MEOX2 activated p21CIP1/WAF1 and p16INK4a gene transcription, inhibited proliferation and induced apoptosis and senescence in endothelial cells. MEOX activation of p21CIP1/WAF1 transcription occurs via a DNA-binding independent mechanism that requires the SP1 transcription factor. In contrast, MEOX activation of p16INK4a transcription is dependent upon DNA-binding. Microarray analysis revealed that both MEOX1 and MEOX2 increased the expression of intercellular adhesion molecule 1 (ICAM-1) and decreased the expression of nitric oxide synthase 3 (NOS3/eNOS).
Taken together, we conclude that MEOX1 and MEOX2 have similar target genes in endothelial cells including p21CIP1/WAF1, p16INK4a and eNOS. As increased endothelial senescence and decreased nitric oxide production are hallmarks of endothelial dysfunction, this study proposes a role for the MEOX proteins in the progression of atherosclerotic vascular disease.
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Molecular cloning and characterization of FHL2, a novel lim domain protein preferentially expressed in human heart. / CUHK electronic theses & dissertations collectionJanuary 1998 (has links)
by Kwok-keung Chan. / Thesis (Ph.D.)--Chinese University of Hong Kong, 1998. / Includes bibliographical references (p. 177-195). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Mode of access: World Wide Web. / Abstracts in English and Chinese.
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Mechanisms of Nkx6.1 governing [B] cell development and function /Taylor, David G January 2006 (has links)
Thesis (Ph. D.)--University of Virginia, 2006. / Bracketed B in title is the Greek letter beta. Includes bibliographical references (leaves150-169). Also available online through Digital Dissertations.
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Molecular analyses of four-and-a-half LIM-only (FHL) protein family. / CUHK electronic theses & dissertations collectionJanuary 2000 (has links)
Hoi-Yeung Li. / "May 2000." / Thesis (Ph.D.)--Chinese University of Hong Kong, 2000. / Includes bibliographical references (p. 151-168). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Mode of access: World Wide Web. / Abstracts in English and Chinese.
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Functional studies of MEIS1, a HOX co-factorGoh, Siew-Lee. January 2007 (has links)
HOX proteins are evolutionarily conserved homeodomain-containing transcription factors involved in hematopoiesis and patterning during embryogenesis. Their tasks as master regulators of embryonic development are achieved in large part through their ability to interact with co-factors of the PBX and MEIS/PREP families, which constitute the broader three amino-acid loop extension (TALE) class of homeodomain proteins. HOX, MEIS, and PBX have been implicated in leukemic hematopoiesis due to their association with hematological malignancies. The oncogenic function of MEIS1 in accelerating the onset of acute myeloid leukemia induced by HOX was mapped to its C-terminal transactivation domain, which is responsive to PKA signaling. This thesis extends our understanding regarding the mechanism by which MEIS1A executes its C-terminal transactivation function in vivo. We describe the involvement of CREB and its co-activators CBP and TORC in conferring the PKA-responsiveness of the ME1S1A C terminus. CREB mutants that fail to bind CBP or TORC also fail to promote PKA induction mediated by the C terminus of ME1S1A. TORC was further shown to be capable of bypassing the need for PKA to activate transcription by MEIS1, an ability endowed by its physical interaction with MEIS1. Chromatin immunoprecipitation (ChIP) demonstrated a concerted recruitment of endogenous MEIS1, TORC2, and CREB proteins on ME1S1 target genes. In addition, this thesis also characterizes the promoter of the murine Meis1 gene. Meis1 possesses multiple transcription start sites upstream of its translation initiation site. We identified a ME1S·PBX consensus recognition site within the Meis1 promoter and showed that PBX1 binds to this sequence in vitro. Our ChIP assay results further suggest an autoregulatory mode for the Meis1 gene as revealed by a co-occupancy of endogenous CREB, TORC2, PBX1, and MEIS1 itself on the Meis1 promoter. Collectively, this thesis proposes a mechanistic action conferred by CREB, CBP and TORC in the PKA-inducible transactivation of ME1S1A, and provides new information on the Meis1 promoter.
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Biochemical analysis of the role of the truncated HOXA1 transcript in cell differentiation /Fernandez, Cristina Carmen. January 2008 (has links)
Thesis (Ph. D.)--Cornell University, January, 2008. / Vita. Includes bibliographical references.
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Functional studies of MEIS1, a HOX co-factorGoh, Siew-Lee. January 2007 (has links)
No description available.
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Molecular cloning and characterization of a cardiac and skeletal muscle LIM domain protein family (FHL). / CUHK electronic theses & dissertations collectionJanuary 1999 (has links)
Simon, Ming-yuen Lee. / Thesis (Ph.D.)--Chinese University of Hong Kong, 1999. / Includes bibliographical references (p. 239-257). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Mode of access: World Wide Web. / Abstracts in English and Chinese.
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Homeobox genes play an important role in smooth muscle cell developmentPerlegas, Demetra Georgia. January 2008 (has links)
Thesis (Ph. D.)--University of Virginia, 2008. / Title from title page. Includes bibliographical references. Also available online through Digital Dissertations.
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