Differential Effects of HNF-1α Mutations Associated with Familial Young-Onset Diabetes on Target Gene Regulation
Differential Effects of HNF-1α Mutations Associated with Familial Young-Onset Diabetes on Target Gene Regulation
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DOI：
10.2119/molmed.2010.00097
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发表时间：
2011-03-01
影响因子：
5.7
通讯作者：
Navas, Maria-Angeles
中科院分区：
文献类型：
--
作者：
Galan, Maria;Garcia-Herrero, Carmen-Maria;Navas, Maria-Angeles
肝细胞核因子1-α（HNF-1 α）是一种在多种组织（包括肝脏和胰腺）中表达的同源域转录因子，可调节多种基因。编码HNF-1 α（HNF 1A）的基因中的杂合突变导致家族性糖尿病，也称为年轻人3型成熟型糖尿病（MODY 3）。MODY 3临床表型的变异性可能是由于环境和遗传因素以及突变的类型和位置。因此，HNF 1A突变的功能表征可能提供对解释MODY 3表型的可变性的分子缺陷的深入了解。我们已经在糖尿病患者中鉴定了六种HNF 1A突变的功能特征：两种新的突变，p.Glu235Gly和c-57- 64 delCACGCGGT;c-55 G>C;以及四种先前描述的突变，p.Val133Met，p.Thr196Ala，p.Arg271Trp和p.Pro379Arg。突变对转录活性的影响已经通过Cos 7和Min 6细胞中HNF-1 α靶启动子亚组的报告基因测定来测量。使用细菌表达的谷胱甘肽-S-转移酶（GST）-HNF-1 α融合蛋白和转染的Cos 7细胞的核提取物，通过电泳迁移率变动试验定量靶DNA结合亲和力。我们的功能研究表明，突变c-57- 64 delCACGCGGT;c-55 G>C降低了Min 6细胞中HNF 1A启动子的活性，并且错义突变具有不同的影响。突变p.Arg271Trp在所有测试条件下都损害HNF-1 α活性，而突变p.Val133Met、p.Glu235Gly和p.Pro379Arg根据靶启动子发挥不同的作用。相比之下，p.Thr196Ala取代似乎不会改变HNF-1 α的功能。我们的研究结果表明，HNF 1A突变可能对特定靶基因的调节有不同的影响，这可能有助于MODY 3临床表型的变异性。(C)2011 Feinstein医学研究所，www.feinsteininstitute.org在线地址：http://www.molmed.org doi：10.2119/molmed.2010.00097
Hepatocyte nuclear factor 1-alpha (HNF-1 alpha) is a homeodomain transcription factor expressed in a variety of tissues (including liver and pancreas) that regulates a wide range of genes. Heterozygous mutations in the gene encoding HNF-1 alpha (HNF1A) cause familial young-onset diabetes, also known as maturity-onset diabetes of the young, type 3 (MODY3). The variability of the MODY3 clinical phenotype can be due to environmental and genetic factors as well as to the type and position of mutations. Thus, functional characterization of HNF1A mutations might provide insight into the molecular defects explaining the variability of the MODY3 phenotype. We have functionally characterized six HNF1A mutations identified in diabetic patients: two novel ones, p.Glu235Gly and c-57-64delCACGCGGT;c-55G>C; and four previously described, p.Val133Met, p.Thr196Ala, p.Arg271Trp and p.Pro379Arg. The effects of mutations on transcriptional activity have been measured by reporter assays on a subset of HNF-1 alpha target promoters in Cos7 and Min6 cells. Target DNA binding affinities have been quantified by electrophoretic mobility shift assay using bacterially expressed glutathione-S-transferase (GST)-HNF-1 alpha fusion proteins and nuclear extracts of transfected Cos7 cells. Our functional studies revealed that mutation c-57-64delCACGCGGT;c-55G>C reduces HNF1A promoter activity in Min6 cells and that missense mutations have variable effects. Mutation p.Arg271Trp impairs HNF-1 alpha activity in all conditions tested, whereas mutations p.Val133Met, p.Glu235Gly and p.Pro379Arg exert differential effects depending on the target promoter. In contrast, substitution p.Thr196Ala does not appear to alter HNF-1 alpha function. Our results suggest that HNF1A mutations may have differential effects on the regulation of specific target genes, which could contribute to the variability of the MODY3 clinical phenotype. (C) 2011 The Feinstein Institute for Medical Research, www.feinsteininstitute.org Online address: http://www.molmed.org doi: 10.2119/molmed.2010.00097