Corticotropin-releasing hormone stimulates proopiomelanocortin transcription by cFos-dependent and -independent pathways: characterization of an AP1 site in exon 1.
Corticotropin-releasing hormone stimulates proopiomelanocortin transcription by cFos-dependent and -independent pathways: characterization of an AP1 site in exon 1.
复制标题
DOI：
10.1210/mend.9.6.8592520
复制
发表时间：
1995-06
影响因子：
--
通讯作者：
A. Boutillier;D. Monnier;D. Lorang;J. Lundblad;J. Roberts;J. Loeffler
中科院分区：
文献类型：
--
作者：
A. Boutillier;D. Monnier;D. Lorang;J. Lundblad;J. Roberts;J. Loeffler
POMC基因编码激素前体蛋白，主要在垂体中以组织特异性方式表达。POMC基因受多种激素和神经肽以及第二信使cAMP和Ca++的转录调节。使用促肾上腺皮质激素衍生的AtT 20细胞系，我们以前已经表明，cFos的过度表达刺激POMC转录。本工作的目的是分析cFos是否直接与基础和促肾上腺皮质激素释放激素（CRH）刺激的细胞中的POMC基因相互作用。使用逐渐缺失的POMC启动子序列或与氯霉素乙酰转移酶报告基因偶联的异源启动子构建体，我们证明了POMC基因第一个外显子内存在主要的cFos响应序列。这个序列，TGACTAA，似乎从典型的AP 1结合位点功能上无法区分。当与最小启动子融合时，该序列赋予cFos和CRH的诱导作用。凝胶位移分析与CRH刺激AtT 20核提取物或在体外合成的蛋白质显示，该序列有效地结合Fos和Jun。表达c-fos反义mRNA减少CRH刺激POMC转录，从而表明，至少部分地，cFos介导CRH对POMC转录的影响。然而，从POMC构建体中删除该主要外显子AP 1位点大大降低了c-fos过表达的作用，但没有抑制CRH对POMC的刺激，表明CRH通过一种或多种cFos非依赖性机制刺激POMC转录。
The POMC gene, encoding a hormonal precursor protein, is primarily expressed in the pituitary in a tissue-specific manner. The POMC gene is transcriptionally regulated by a variety of hormones and neuropeptides and the second messengers cAMP and Ca++. Using the corticotrope-derived AtT20 cell line, we have previously shown that overexpression of cFos stimulates POMC transcription. The aim of this work was to analyze whether cFos directly interacts with the POMC gene in basal and corticotropin-releasing hormone (CRH) stimulated cells. Using progressively deleted POMC promoter sequences or heterologous promoter constructs coupled to the chloramphenicol acetyl transferase reporter gene, we demonstrate the existence of a major cFos- responsive sequence within the first exon of the POMC gene. This sequence, TGACTAA, appears functionally indistinguishable from the canonical AP1 binding site. When fused to a minimal promoter, this sequence confers inducibility by cFos and CRH. Gel shift analyses with CRH-stimulated AtT20 nuclear extracts or in vitro synthesized proteins revealed that this sequence efficiently binds Fos and Jun. Expression of c-fos anti-sense mRNA reduced CRH-stimulated POMC transcription, thus indicating that, at least in part, cFos mediates the effect of CRH on POMC transcription. However, deletion of this major exonic AP1 site from the POMC constructs greatly reduced the effect of c-fos overexpression but did not suppress POMC stimulation by CRH, indicating that CRH stimulates POMC transcription by one or more cFos-independent mechanism(s).