StAR expression: Integration of Transcription with regulation via the mRNA 3'UTR

描述（由申请人提供）：肾上腺皮质中的皮质醇合成在应激反应、饮食控制和心血管调节中起着重要作用。下丘脑-垂体-肾上腺轴的这种控制的异常在代谢综合征中起重要作用，代谢综合征通常与2型糖尿病相关。ACTH刺激皮质醇合成始于胆固醇通过线粒体细胞色素P450 11 A1转化为胆固醇烯醇酮。这一步骤的激活依赖于类固醇生成急性调节因子（星星）的新合成及其被蛋白激酶A（PKA）磷酸化。星星缺乏会导致高脂血症性肾上腺功能不全。这项研究的重点是发现星星的表达是由PKA和蛋白激酶C（PKC）以非常不同的方式刺激的。我们将展示这些过程是如何明确区分的重要贡献，分别从组蛋白去乙酰化酶和ERK激酶。我们将描述这些过程的不同贡献的核因子，包括分化调节剂，SF-1。这包括在转录期间上下循环的时间依赖性修饰（磷酸化/去磷酸化和乙酰化/去乙酰化）。将有证据表明PKA还可以增强PKC过程的后期阶段，从而在星星转录中产生强大的协同作用。星星的转录后调节提供了PKA和PKC调节之间的另一个区别点。一种新的调节剂，锌指蛋白，TIS 11b，靶向在3.5 kb星星mRNA的延伸的3 '非翻译区的特定序列。TIS 11b被PKA快速激活，但被PKC抑制。我们将在星星3.5 kb mRNA的末端确定特定的TIS 11b识别位点，并表明这种相互作用可以增强星星蛋白的翻译，同时增加mRNA的降解。我们将测试TIS 11b是否由于共享转录因子（包括SF-1）而与星星共转录。我们将确定这些机制如何在促肾上腺皮质激素刺激过程中相互作用，包括在原代肾上腺细胞系和体内肾上腺。在星星表达期间指定SF-1和TIS 11b相互作用的启动子和mRNA序列从小鼠到人类基本上是保守的。我们将测试这些小鼠机制是否保留在人肾上腺H295细胞中。我们的研究表明，TIS 11b与星星的相互作用以及星星mRNA的优势在原代牛肾上腺细胞中是保守的，并且对ACTH有反应。这项工作将汇集几个实验室，以提供PKA，SF-1和TIS 11b（或与每个相关的因子）之间的相互作用的第一个视图，这可能发生在多种类固醇生成组织中。肾上腺皮质中的皮质醇合成在应激反应、饮食控制和心血管调节中起着中心作用。促肾上腺皮质激素，这是由压力升高，刺激皮质醇的合成，通过增强胆固醇转化为去甲烯醇酮。这一步取决于类固醇生成急性调节剂（星星）的新合成。这项研究提出了一个新的过程，ACTH不仅刺激星星的转录，还刺激一种名为TIS 11b的蛋白质，该蛋白质分别调节星星蛋白质翻译和mRNA降解。我们对控制星星和TIS 11b转录的调控过程进行了详细分析，以了解它们之间的潜在协调。TIS 11b可以加速对压力的反应时间，同时也有助于在ACTH刺激被去除时去除星星。TIS 11b调节存在于产生类固醇的其他组织（睾丸）中。这种mRNA调节器的缺乏可能会导致对ACTH和压力的反应更加迟缓，以及皮质醇输出异常。

DESCRIPTION (provided by applicant): Cortisol synthesis in the adrenal cortex plays a central role in stress responses, in dietary control, and in cardiovascular regulation. Aberrations in this control of the hypothalamic pituitary adrenal axis play an important part in the Metabolic Syndrome that is often associated with Type 2 diabetes. ACTH stimulation of cortisol synthesis starts with the conversion of cholesterol to pregnenolone by mitochondrial cytochrome P450 11A1. Activation of this step depends on new synthesis of the steroidogenesis acute regulator (StAR) and its phosphorylation by protein kinase A (PKA). StAR deficiency causes hyperlipidemic adrenal insufficiency. This research focuses on the finding that StAR expression is stimulated in very different ways by PKA and by protein kinase C (PKC). We will show how these processes are clearly distinguished by essential contributions from, respectively, histone de-acetylatases and Erk kinase. We will characterize these processes for different contributions from nuclear factors, including the differentiation regulator, SF-1. This includes time-dependent modifications that cycle up and down during transcription (phosphorylation/de-phosphorylation and acetylation/de- acetylation). Evidence will be developed that PKA can also enhance a late stage in the PKC process, thus producing strong synergy in StAR transcription. Post transcriptional regulation of StAR provides another point of distinction between PKA and PKC regulation. A novel regulator, the Zn finger protein, TIS11b, targets specific sequences in the extended 3'untranslated region of the 3.5 kb StAR mRNA. TIS11b is rapidly stimulated by PKA, but suppressed by PKC. We will identify specific TIS11b recognition sites at the end of the StAR 3.5 kb mRNA and show that this interaction can enhance StAR protein translation, while increasing mRNA degradation. We will test whether TIS11b is co-transcribed with StAR due to shared transcription factors, including SF-1. We will determine how these mechanisms interplay during ACTH stimulation, including in primary adrenal cell lines and in adrenals in vivo. The promoter and mRNA sequences that specify SF-1 and TIS11b interactions during StAR expression are substantially conserved from mouse to humans. We will test whether these mouse mechanisms are retained in human adrenal H295 cells. Our research shows that TIS11b interaction with StAR and predominance of an extended StAR mRNA are conserved in primary bovine adrenal cells and respond to ACTH. This work will bring together several laboratories to provide a first look at the interplay between PKA, SF-1, and TIS11b (or factors related to each), which is likely to occur in multiple steroidogenic tissues. Cortisol synthesis in the adrenal cortex plays a central role in stress responses, in dietary control, and in cardiovascular regulation. ACTH, which is elevated by stress, stimulates cortisol synthesis through enhanced conversion of cholesterol to pregnenolone. This step depends on new synthesis of the steroidogenesis acute regulator (StAR). The proposed research addresses a novel process, whereby ACTH stimulates not only transcription of StAR, but also a protein called TIS11b, which separately regulates StAR protein translation and mRNA degradation. We make a detailed analysis of regulatory processes that control StAR and TIS11b transcription in order to understand their potential coordination. TIS11b may accelerate the response time to stress, while also aiding in the removal of StAR when the ACTH stimulus is removed. TIS11b regulation is present in other tissues that make steroids (testis). Deficiency in this mRNA regulator may lead to more sluggish responses to ACTH and stress as well as abnormal cortisol output.