Characterization of mRNA-protein complexes as modulators of cellular hypoxia response.

mRNA-蛋白质复合物作为细胞缺氧反应调节剂的表征。

• 批准号: 280542466
• 负责人: Dr. Isabel Naarmann-de Vries
• 金额: --
• 依托单位: Klinik für Operative Intensivmedizin und Intermediate Care
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2015
• 资助国家: 德国
• 起止时间: 2014-12-31 至 2018-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/280542466?language=en
• 关键词: Characterization; mRNA; protein; complexes; modulators

Hypoxia is described as cellular lack of oxygen and strongly affects energy consuming processes as the activity of the Na-K-ATPase and mRNA translation. Thus, under hypoxic conditions global, 5´cap-dependent mRNA translation is inhibited. However, a subset of mRNAs escapes this inhibition by different mechanisms. Internal ribosome entry sites (IRESs) mediate the 5´cap-independent initiation of translation. Under hypoxia translation is favored at ER-bound ribosomes. Cis-elements in the untranslated regions (UTRs) of certain mRNAs and trans-acting proteins mediate their localization to the ER and thereby efficient translation under hypoxic conditions. Employing the MCF-7 cell system I will study the post-transcriptional control of gene expression during the hypoxic response utilizing protein occupancy profiling. RNA motifs differentially occupied with proteins will be identified by this method. Interacting RNA binding proteins (RBPs) will either be identified by RNA affinity chromatography in combination with mass spectrometry analysis or based on their known RNA binding motifs. The function of dynamic RNA-protein interactions in post-transcriptional control of gene expression during the hypoxic response will be analyzed in RNA interference, mRNA stability, ribosome profiling and in vitro translation experiments. These analyses will give insight into mechanisms, by which tumor cells escape hypoxic conditions limiting tumor growth.

缺氧被描述为细胞缺氧，并强烈影响na - k - atp酶活性和mRNA翻译等能量消耗过程。因此，在全球缺氧条件下，依赖5´cap的mRNA翻译受到抑制。然而，mrna的一个子集通过不同的机制逃脱了这种抑制。内部核糖体进入位点（IRESs）介导与5´cap无关的翻译起始。在缺氧条件下，er结合的核糖体更倾向于翻译。某些mrna和反式蛋白的非翻译区（UTRs）中的顺式元件介导其定位到内质网，从而在缺氧条件下有效翻译。利用MCF-7细胞系统，我将利用蛋白占用谱研究缺氧反应过程中基因表达的转录后调控。用这种方法可以鉴定与蛋白质不同的RNA基序。相互作用RNA结合蛋白（rbp）可以通过RNA亲和层析结合质谱分析或基于其已知的RNA结合基序来鉴定。我们将通过RNA干扰、mRNA稳定性、核糖体谱分析和体外翻译实验分析RNA-蛋白动态相互作用在缺氧反应过程中转录后调控基因表达中的作用。这些分析将深入了解肿瘤细胞逃避限制肿瘤生长的缺氧条件的机制。