Apolipoprotein E (apoE) is a lipid transport protein which plays a key role in lipid metabolism. In addition to the well known polymorphic coding alleles epsilon2, epsilon3 and epsilon4, APOE promoter single nucleotide polymorphisms (SNPs) have also been reported to modify disease susceptibilities in humans. / In a case-control study involving 710 Chinese type 2 diabetes and 198 non-diabetic subjects, genotyping of three SNPs (-491A/T, -219G/T and +113G/C) within the APOE proximal promoter identified that -491A was associated with increased risk for type 2 diabetes in women (OR=2.44, 95%CI=1.15-5.19, p=0.017). However, the three tested SNPs were not associated with the risk of diabetic nephropathy (DN). Yeast one-hybrid screening of the human brain cDNA library using the polymorphic DNA sequences spanning the APOE promoter -491 site as the 'baits' identified one of the interacting transcription factors being the activating transcription factor 4 (ATF4). Electrophoretic-mobility-shift assay confirmed the physical interaction of the purified recombinant ATF4 protein and APOE promoter -491 A/T spanning region (-521 to -461). The binding of ATF4 to the -491T-containing sequence was stronger than that of the -491A-containing sequence. Chromatin immunoprecipitation (ChIP) assay further confirmed the interaction between ATF4 and APOE promoter -491-spanning region in vivo. The functional significance of APOE -491A/T polymorphism was supported by the dual-luciferase reporter assay showing that -491 A to T single nucleotide substitution significantly decreased the activity of the cloned APOE promoter (-1019 to +407) in human kidney (293), liver (WRL-68) and astrocyte (U-87) cell lines. Further analysis showed that ATF4 over-expression significantly down-regulated the activities of the cloned APOE promoter. The suppression of ATF4 on APOE promoter with -491A allelic form was significantly stronger than that with -491T allelic form in 293 cells (p<0.05). Interestingly, overexpression of recombinant ATF4 stimulated endogenous APOE transcription by about 10% in WRL-68 cells. / In conclusion, APOE promoter -491A/T polymorphism modifies the risk of type 2 diabetes in Hong Kong Chinese women. The -491A/T polymorphism controls APOE promoter activity and is interactive with transcription factor ATF4. / My thesis project aimed at testing two hypotheses: (1) APOE promoter SNPs associate with the risks of type 2 diabetes and diabetic nephropathy, (2) APOE promoter SNPs modify transcriptional control of the gene. / Geng, Hua. / "September 2007." / Source: Dissertation Abstracts International, Volume: 69-08, Section: B, page: 4559. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2007. / Includes bibliographical references (p. 140-151). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [200-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstracts in English and Chinese. / School code: 1307.
| Identifer | oai:union.ndltd.org:cuhk.edu.hk/oai:cuhk-dr:cuhk_344137 |
| Date | January 2007 |
| Contributors | Geng, Hua., Chinese University of Hong Kong Graduate School. Division of Biochemistry. |
| Source Sets | The Chinese University of Hong Kong |
| Language | English, Chinese |
| Detected Language | English |
| Type | Text, theses |
| Format | electronic resource, microform, microfiche, 1 online resource (xxi, 153 p. : ill.) |
| Rights | Use of this resource is governed by the terms and conditions of the Creative Commons “Attribution-NonCommercial-NoDerivatives 4.0 International” License (http://creativecommons.org/licenses/by-nc-nd/4.0/) |
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