Hepatic Nuclear Factor 3 and Nuclear Factor 1 Regulate 5-Aminolevulinate Synthase Gene Expression and Are Involved in Insulin Repression*
Hepatic Nuclear Factor 3 and Nuclear Factor 1 Regulate 5-Aminolevulinate Synthase Gene Expression and Are Involved in Insulin Repression*
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肝核因子 3 和核因子 1 调节 5-氨基乙酰丙酸合酶基因表达并参与胰岛素抑制*
DOI：
10.1074/jbc.m401792200
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发表时间：
2004
影响因子：
4.8
通讯作者：
E. Cánepa
中科院分区：
文献类型：
--
作者：
M. E. Scassa;Alejandra S. Guberman;J. M. Ceruti;E. Cánepa
尽管胰岛素对基因表达的负调控已经得到了广泛的研究，但负责胰岛素效应的转录因子仍不清楚。本工作的目的是探讨5-氨基酮戊酸合成酶(ALAS)基因胰岛素抑制的分子机制。对5‘-调节区的缺失分析使我们能够确定位于-459到-354个碱基之间的胰岛素反应区。该片段包含一个高度同源的胰岛素反应(IRE)序列。通过瞬时转染实验，我们确定了肝核因子3(HNF3)和核因子1(NF1)是ALAS基因适当表达所必需的。胰岛素可抑制HNF3β或HNF3β加NF1介导的ALAS转录活性的刺激。凝胶迁移率改变分析和Southwest blotting表明HNF3与ALAS启动子结合。对该区域的突变分析表明，IRE的中断会取消胰岛素的作用，而HNF3元件的突变会维持激素的反应性。这种HNF3结合和胰岛素作用之间的分离表明HNF3β不是胰岛素诱导的转录抑制的唯一生理中介。此外，Southwest blotting分析表明，除HNF3β外，至少有两个多肽可以与ALAS启动子结合，并且这种结合依赖于IRE的完整性。我们提出了一个模型，在该模型中，胰岛素通过干扰HNF3AKT对转录因子β的结合或激活而发挥负效应，这可能是由于Akt对该转录因子的特异性磷酸化所致。在这一点上，从使用激酶抑制剂的转染实验中获得的结果支持这一假说。由于这一事件，NF1将失去对发起人的可及性。HNF3的翻译后修饰将允许识别核心IRE的蛋白质复合体结合。这些结果为胰岛素介导的含IRE启动子的抑制提供了一个潜在的机制。
Although the negative regulation of gene expression by insulin has been widely studied, the transcription factors responsible for the insulin effect are still unknown. The purpose of this work was to explore the molecular mechanisms involved in the insulin repression of the 5-aminolevulinate synthase (ALAS) gene. Deletion analysis of the 5′-regulatory region allowed us to identify an insulin-responsive region located at –459 to –354 bp. This fragment contains a highly homologous insulin-responsive (IRE) sequence. By transient transfection assays, we determined that hepatic nuclear factor 3 (HNF3) and nuclear factor 1 (NF1) are necessary for an appropriate expression of the ALAS gene. Insulin overrides the HNF3β or HNF3β plus NF1-mediated stimulation of ALAS transcriptional activity. Electrophoretic mobility shift assay and Southwestern blotting indicate that HNF3 binds to the ALAS promoter. Mutational analysis of this region revealed that IRE disruption abrogates insulin action, whereas mutation of the HNF3 element maintains hormone responsiveness. This dissociation between HNF3 binding and insulin action suggests that HNF3β is not the sole physiologic mediator of insulin-induced transcriptional repression. Furthermore, Southwestern blotting assay shows that at least two polypeptides other than HNF3β can bind to ALAS promoter and that this binding is dependent on the integrity of the IRE. We propose a model in which insulin exerts its negative effect through the disturbance of HNF3β binding or transactivation potential, probably due to specific phosphorylation of this transcription factor by Akt. In this regard, results obtained from transfection experiments using kinase inhibitors support this hypothesis. Due to this event, NF1 would lose accessibility to the promoter. The posttranslational modification of HNF3 would allow the binding of a protein complex that recognizes the core IRE. These results provide a potential mechanism for the insulin-mediated repression of IRE-containing promoters.
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影响因子：
10.5
作者：
Shim, EY;Woodcock, C;Zaret, KS
通讯作者：
Zaret, KS
影响因子：
15.9
作者：
Li, WW;Dammerman, MM;Leff, T
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Leff, T
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10.5
作者：
Sund, NJ;Vatamaniuk, MZ;Kaestner, KH
通讯作者：
Kaestner, KH
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3.7
作者：
Dong, Y;Asch, HL;Asch, BB
通讯作者：
Asch, BB
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作者：
Gartel, AL;Goufman, E;Tyner, AL
通讯作者：
Tyner, AL