Cytokine-Mediated Regulation of Cholesterogenesis

描述(申请人提供)：角鲨烯合成酶(SQS)催化 致力于胆固醇的类异戊二烯代谢途径的第一反应 生物合成及其活性调节中间体向甾醇的流动。 SQS受类固醇、脂多糖和促炎因子的调节。 细胞因子肿瘤坏死因子-α和白介素1-β (IL-1-β)。甾醇介导的SQS转录调控良好 了解，但对其调控机制知之甚少。 炎症反应。我们建议检验这一假设，即内毒素和 细胞因子调控SQS mRNA转录和稳定性及酶蛋白 翻译后修饰、稳定性和亚细胞定位。在……里面 具体目标1我们将确定内毒素和细胞因子对肝脏的影响 不同饲料对叙利亚仓鼠SQS转录和mRNA稳定性的影响 高胆固醇饮食。我们还将确定SQS mRNA水平和 细胞因子应答过程中的大小和特异性 这些过程中的细胞因子。在具体目标2中，我们建议澄清 脂多糖和细胞因子调节SQS基因表达的分子机制。 在培养的细胞中，这将通过检查启动子序列来实现 负责细胞因子转录抑制的元件，通过 鉴定参与这种抑制的转录因子，并通过 参与转录的细胞信号通路的阐明 监管。此外，SQS mRNA所需的序列通过以下方式失稳 细胞因子将被本地化。在具体目标3中，我们建议阐明内毒素-和 细胞因子诱导的翻译后机制 SQS酶活性。我们将研究SQS蛋白质失稳，机制 对于它的降解，通过磷酸化的翻译后修饰，以及 内毒素和细胞因子对其亚细胞定位的影响。 这三个具体目标的实现将直接有助于我们 阐明SQS对类异戊二烯调节的重要性的长期目标 代谢流。这将增加我们对类固醇代谢的了解。 对感染和炎症的急性时相反应(APR)及SQS的作用 这一过程中的监管。最后，它将提高我们的总体认识 在正常和病理情况下，肝脏胆固醇的产生。

DESCRIPTION (provided by applicant): Squalene synthase (SQS) catalyzes the first reaction of the isoprenoid metabolic pathway committed to cholesterol biosynthesis and its activity regulates the flux of intermediates to sterols. SQS is regulated by sterols, lipopolysaccharide (LPS), and the pro-inflammatory cytokines tumor necrosis factor-alpha (TNF-alpha) and interlukin-1-beta (IL-1-beta). Sterol-mediated transcriptional regulation of SQS is well understood, but little is known about the mechanism of its regulation during the inflammatory response. We propose to test the hypothesis that LPS and cytokines regulate SQS mRNA transcription and stability and enzyme protein posttranslational modification, stability and subcellular localization. In Specific Aim 1 we will determine the effects of LPS and cytokines on hepatic SQS transcription and mRNA stability in Syrian hamsters fed with different cholesterogenic diets. We will also determine changes in SQS mRNA level and size during response to cytokines and the contribution of the specific cytokines to these processes. In Specific Aim 2 we propose to elucidate the molecular mechanisms responsible for SQS mRNA regulation by LPS and cytokines. This will be achieved, in cultured cells, by examining promoter sequence elements responsible for the transcriptional repression by cytokines, by identification of transcription factors involved in this suppression, and by the elucidation of cellular signaling pathways involved in the transcriptional regulation. In addition, sequences required for SQS mRNA destabilization by cytokines will be localized. In Specific Aim 3 we propose to elucidate the LPS-and cytokine-induced, post-translational mechanisms underlying the decrease in SQS enzymic activity. We will examine SQS protein destabilization, mechanisms for its degradation, post-translational modification by phosphorylation, and change in its subcellular localization in response to LPS and cytokines. Achievement of the three Specific Aims will contribute directly to our long-term goal to elucidate the importance of SQS regulation on isoprenoid metabolic flux. It will increase our understanding of sterol metabolism in acute phase response (APR) to infection and inflammation and the role of SQS regulation in this process. Finally, it will enhance our overall understanding of hepatic cholesterol production in normal and pathological situations.