Synergistic activation of the inhibin alpha-promoter by steroidogenic factor-1 and cyclic adenosine 3',5'-monophosphate.
Synergistic activation of the inhibin alpha-promoter by steroidogenic factor-1 and cyclic adenosine 3',5'-monophosphate.
复制标题
DOI：
10.1210/mend.14.1.0410
复制
发表时间：
2000
影响因子：
--
通讯作者：
Masafumi Ito;Youngkyu Park;J. Weck;K. Mayo;J. Jameson
中科院分区：
文献类型：
--
作者：
Masafumi Ito;Youngkyu Park;J. Weck;K. Mayo;J. Jameson
α-亚基基因在卵巢、睾丸、肾上腺和垂体中表达。由于这种表达模式对应于孤儿核受体，类固醇生成因子-1（SF-1），我们假设，可能是由SF-1调节的α启动子。外源SF-1的表达，在SF-1缺陷的细胞系，引起适度的刺激的α启动子。然而，cAMP途径的激活，这是已知的调节α表达，大大增强SF-1的作用。SF-1与cAMP依赖性蛋白激酶A的催化亚基的共表达引起大于250倍的刺激，而仅通过SF-1或蛋白激酶A途径观察到4或7倍的刺激。在表达内源性SF-1的GRMO 2颗粒细胞中也观察到SF-1和cAMP途径的协同刺激。缺失和定点突变定位了一个新的SF-1调节元件（TCA GGGCCA; -137至-129），与变体cAMP反应元件（CRE; -120至-114）相邻。SF-1和cAMP刺激的协同特性在该复合的α-腺苷酸片段（-146和-112）内是固有的，因为它可转移到异源启动子。CRE或SF-1调节元件中的突变完全消除了这些途径的协同激活。SF-1和CRE结合蛋白（CREB）的结合，以α调节元件是相对较弱的凝胶迁移率变动分析，一致的偏离共识结合位点。然而，SF-1被发现与CREB相互作用，使用一种测定法，其中表位标记的SF-1在细胞中表达，并用于拉下体外翻译的CREB。CREB结合蛋白（CBP）的表达，与SF-1和CREB相互作用的共激活因子，进一步增强了这些途径的转录。SF-1和cAMP通路的刺激与组蛋白H4乙酰化增加相关，表明染色质重塑伴随着它们的作用。我们提出了一个模型，其中SF-1，CREB，和相关的辅激活因子，如CBP的直接相互作用诱导强烈的合作transactivation的途径，单独有相对较弱的影响转录。
The inhibin alpha-subunit gene is expressed in the ovary, testis, adrenal, and pituitary. Because this pattern of expression corresponds to that of the orphan nuclear receptor, steroidogenic factor-1 (SF-1), we hypothesized that the inhibin alpha promoter might be regulated by SF-1. Expression of exogenous SF-1, in an SF-1 deficient cell line, caused modest stimulation of the inhibin alpha promoter. However, activation of the cAMP pathway, which is known to regulate inhibin alpha expression, greatly enhanced the actions of SF-1. Coexpression of SF-1 with the catalytic subunit of cAMP-dependent protein kinase A caused greater than 250-fold stimulation, whereas only 4- or 7-fold stimulation was seen by the SF-1 or protein kinase A pathway alone. Synergistic stimulation by SF-1 and the cAMP pathway was also seen in GRMO2 granulosa cells, which express endogenous SF-1. Deletion and site-directed mutagenesis localized a novel SF-1 regulatory element (TCA GGGCCA; -137 to -129) adjacent to a variant cAMP-response element (CRE; -120 to -114). The synergistic property of SF-1 and cAMP stimulation was inherent within this composite inhibin alpha fragment (-146 and -112), as it was transferable to heterologous promoters. Mutations in either the CRE or the SF-1 regulatory element completely eliminated synergistic activation by these pathways. The binding of SF-1 and CRE binding protein (CREB) to the inhibin alpha regulatory elements was relatively weak in gel mobility shift assays, consistent with their deviation from consensus binding sites. However, SF-1 was found to interact with CREB using an assay in which epitope-tagged SF-1 was expressed in cells and used to pull down in vitro translated CREB. Expression of CREB binding protein (CBP), a coactivator that interacts with SF-1 and CREB, further enhanced transcription by these pathways. Stimulation by the SF-1 and cAMP pathways was associated with increased histone H4 acetylation, suggesting that chromatin remodeling accompanies their actions. We propose a model in which direct interactions of SF-1, CREB, and associated coactivators like CBP induce strongly cooperative transactivation by pathways that individually have relatively weak effects on transcription.