Posttranscriptional Regulation Of Genes Controling Cell

控制细胞基因的转录后调控

• 批准号: 6663572
• 负责人: MYRIAM none GOROSPE
• 金额: --
• 依托单位: NATIONAL INSTITUTE ON AGING
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/6663572
• 关键词: RNA binding protein; biological signal transduction; cell growth regulation; cell proliferation; cyclins; developmental genetics; gene induction /repression; human tissue; messenger RNA; oncoprotein p21; posttranscriptional RNA processing; serial analysis of gene expression; tissue /cell culture; tumor suppressor proteins

In response to signals of either endogenous or exogenous origin, mammalian cells activate a series of events leading to changes in gene expression. In turn, these alterations in gene expression directly participate in the implementation of a global cellular response. While the transcriptional events regulating such changes in gene expression have been thoroughly studied, it is becoming increasingly apparent that posttranscriptional events, which are less well understood, are also major regulatory mechanisms that profoundly modify gene expression. Posttranscriptional events of gene regulation include mRNA processing, transport, stability and translation, as well as protein processing, phosphorylation and degradation. With respect to mRNA stability, we are investigating the mechanisms regulating the expression of various cell cycle regulatory and proliferation-associated genes. We have shown that the expression of cyclins A and B1 throughout the cell division cycle was regulated through the cyclic association of their respective mRNAs with the RNA-binding protein HuR, which results in transcript stabilization during the S phase. We have also reported that high levels of HuR expression in young human fibroblasts contributes to the heightened presence of cyclins A and B1, as HuR stabilizes their mRNAs. Conversely, in senescent human fibroblasts, lower HuR levels directly cause a reduction in the stability, and hence the expression of cyclin A and cyclin B1 mRNAs. Other studies from our laboratory have shown that decreased expression of cyclin D1 following treatment with the stress agent prostaglandin A2 was accomplished through cyclin D1 mRNA destabilization and likely involved the RNA-binding protein AUF1. We previously demonstrated that induction of the cyclin-dependent kinase inhibitor p21 by ultraviolet light and other stresses occurs through stabilization of its mRNA by HuR. Over the past year, we reported that the localization of HuR within the cell is modulated by the AMP-activated kinase (AMPK). This regulatory event is critical for HuR function, as its mRNA stabilizing influence occurs in the cytoplasm. More recently, we have undertaken a high-throughput approach to assess the relative roles of transcription and mRNA turnover in governing gene expression during the cellular stress response. Using cDNA array technology, we have developed a novel approach which involves comparison of large-scale hybridization patterns generated with steady-state mRNA versus newly transcribed (nuclear run-on) RNA. This methodology has allowed us to demonstrate the importance of mRNA turnover in regulating gene expression following several conditions of stress. Our long-term efforts are thus focused on searching for RNA-binding proteins, target mRNA regions, and signaling pathways involved in regulating the stability of mRNAs encoding growth control and cell cycle regulatory genes. The product of the tumor suppressor gene von Hippel- Lindau (pVHL) is believed to modulate gene expression at the levels of transcription elongation, mRNA stability and protein degradation. Our investigation of pVHL's global influence on gene expression using SAGE was recently reported. Now, we have focused our efforts towards understanding VHL?s influence on protein translation. We anticipate that these studies will help elucidate pVHL's function and its tumor suppressor properties.

为了响应内源性或外源性起源的信号，哺乳动物细胞激活一系列事件，导致基因表达变化。反过来，基因表达的这些改变直接参与了全局细胞反应的实施。虽然已经对调节基因表达中这种变化的转录事件进行了彻底的研究，但越来越明显的是，文章后事件的理解较低，也是深刻地改变基因表达的主要调节机制。基因调节的转录后事件包括mRNA处理，运输，稳定性和翻译，以及蛋白质加工，磷酸化和降解。 关于mRNA稳定性，我们正在研究调节各种细胞周期调节和增殖相关基因表达的机制。我们已经表明，整个细胞分裂周期中细胞周期蛋白A和B1的表达通过其各自的mRNA与RNA结合蛋白HUR的循环结合进行调节，从而导致在S期间稳定转录本。我们还报告说，年轻人类成纤维细胞中的HUR表达高水平有助于细胞周期蛋白A和B1的存在，因为HUR稳定了其mRNA。相反，在衰老的人成纤维细胞中，较低的HUR水平直接导致稳定性降低，从而表达细胞周期蛋白A和Cyclin b1 mRNA。我们实验室的其他研究表明，用胁迫剂前列腺素A2治疗后，细胞周期蛋白D1的表达降低是通过细胞周期蛋白D1 mRNA不稳定完成的，并且可能涉及RNA结合蛋白AUF1。我们先前证明，紫外线和其他应力通过HUR稳定其mRNA来诱导细胞周期蛋白依赖性激酶抑制剂p21。在过去的一年中，我们报告说，HUR在细胞内的定位是由AMP激活激酶（AMPK）调节的。该调节事件对于HUR功能至关重要，因为其mRNA稳定影响发生在细胞质中。最近，我们采取了一种高通量方法来评估在细胞应激反应过程中转录和mRNA转换基因表达中的相对作用。使用cDNA阵列技术，我们开发了一种新颖的方法，涉及比较用稳态mRNA与新转录（核跑步）RNA产生的大规模杂交模式。这种方法使我们能够证明在多种压力条件下，在调节基因表达中mRNA转换的重要性。因此，我们的长期努力集中在寻找RNA结合蛋白，靶mRNA区域以及涉及调节编码生长控制和细胞周期调节基因的mRNA稳定性的信号通路。 据信肿瘤抑制基因von Hippellindau（PVHL）的乘积在转录伸长，mRNA稳定性和蛋白质降解水平下调节基因表达。最近，我们使用SAGE对PVHL对基因表达的全球影响进行了研究。现在，我们集中精力理解VHL对蛋白质翻译的影响。我们预计这些研究将有助于阐明PVHL的功能及其肿瘤抑制特性。