Characterization of the 5-Region Regulatory Sequences of Human Angiotensin-Converting Enzyme 2 Gene
| Autor: | Yang, Tzu-Hui, 楊子慧 |
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| Rok vydání: | 2010 |
| Druh dokumentu: | 學位論文 ; thesis |
| Popis: | 98 The renin-angiotensin system (RAS) is a key regulatory axis on blood pressure and fluid homeostasis and critically involves in cardiovascular pathophysiology. Angiotensin- converting enzyme 2 (ACE2) has been proposed as a potential cardioprotective target in regulating cardiovascular functions owing to its key role in the formation of vasoprotective peptides angiotensin-(1-7) [Ang-(1-7)] from angiotensin II (Ang II), an effector peptide of RAS with vasoconstrictive, pro-inflammatory and pro-fibrotic properties. However, the regulatory mechanism of human ace2 expression remains to be explored. In this study, we investigated the human ace2 promoter to identify regulatory elements of the gene. The human ace2 gene promoter from position -2,069 to +20 was cloned and series 5’ deletion mutants were constructed in a luciferase reporter vector. The reporter expressions were analyzed by transient transfection of the constructants in human cardiofibroblast (HCF) cells. The results indicate one activating region at -516/-481 and one repressing region at -627/-516 in HCF cells. Furthermore, deletion of domain (-516/-481) within the ace2 gene construct (-2,069/+20) resulted in a significant decrease in luciferase activity. Mobility shift electrophoresis with double stranded oligonucleotides (-516/-481) resulted in a DNA-protein complex. The effects of angiotensin peptides, Ang II and Ang-(1-7), and pro-inflammatory factors, TGF-beta?? and TNF-alpha, on the transcriptional activity of the ace2 promoter variants in HCF cells were investigated. The experimental results show that the relative luciferase activities of the (-516/+20) construct increase significantly in response to Ang II and Ang-(1-7) indicating that the angiotensin peptides can upregulate transcription of ace2 gene. However, the transcriptional activity of the ace2 promoter in HCF cells did not seem to be affected by TGF-beta?? and TNF-alpha treatments?| In conclusion, our results suggest that domain (-516/-481) of ace2 gene may be a binding domain for as yet unidentified regulatory factors that can activate ace2 expression and the activation is associated with angiotensin peptides signaling. |
| Databáze: | Networked Digital Library of Theses & Dissertations |
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