Investigating the role of 4E-T, an eIF4E-binding protein resident in P-bodies, in gene expression control

研究 4E-T（一种驻留在 P 体中的 eIF4E 结合蛋白）在基因表达控制中的作用

• 批准号: BB/J00779X/1
• 负责人: Nancy Standart
• 金额: $ 54.13万
• 依托单位: University of Cambridge
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2012
• 资助国家: 英国
• 起止时间: 2012 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FJ00779X%2F1
• 关键词: Investigating; role; 4E; eIF4E; binding

DNA is copied into messenger RNA in the nucleus, and following transfer to the cytoplasm, mRNA is translated into protein. mRNAs are translated into protein at different rates, leading to vastly different protein levels, which in part explains why muscle and red blood cells, for example, are so distinct though sharing the same genes. Regulation of mRNA translation is principally controlled by eIF4E, which binds the unusual cap structure at the 5' end of mRNA. eIF4E is important because it interacts with eIF4G which recruits ribosomes, the translational machinery, to mRNAs. Alternative eIF4E-binding proteins exist, such as 4E-BPs and these prevent eIF4G binding to eIF4E and reduce translation. I propose to investigate a more recently discovered eIF4E-binding protein called 4E-T, which also competes with eIF4G for eIF4E-binding. Interestingly, 4E-T and a fraction of eIF4E is concentrated in punctate cytoplasmic "bodies" visible in the microscope, called P-bodies, which lack ribosomes, and are thought to be sites where mRNAs are stored and returned to translation, or where mRNAs are degraded. Despite considerable interest, the role of these P-bodies remains enigmatic, though they influence for example virus replication. Published and unpublished work from us and other investigators show that 4E-T regulates a sub-set of mRNAs, but it is not known what sort of proteins they encode, nor is it known whether 4E-T has to be in P-bodies to exert this control, what proteins it binds there, or what its modification by phosphorylation means for this control. These are the questions we want to answer, using human cells in culture. We believe this work is highly timely since aberrant levels of eIF4E (and its phosphorylation) has been implicated in cell growth, proliferaion and in cancer development, and therefore investigation of a factor that interacts with eIF4E is important. Moreover, the role of P-bodies is still relatively unexplored, and our work will make a significant contribution to our understanding of how mRNAs are regulated in these foci. Last, we're fortunate to collaborate with Professor Anne Willis from the MRC Toxicology Unit, Leicester, to undertake a part of the analysis of 4E-T mRNA targets, and to extend and benefit from a Translational Resource database there, containing data from our and similar experiments from other investigators. The proposed work is novel, focused and timely, and the outcomes will contribute to our understanding and possible exploitation of the regulation of translation initiation by eIF4E and its interacting factors.

DNA在细胞核中被复制成信使RNA，然后转移到细胞质中，mRNA被翻译成蛋白质。mRNA以不同的速率翻译成蛋白质，导致蛋白质水平的巨大差异，这在一定程度上解释了为什么肌肉和红细胞虽然共享相同的基因，但却如此不同。mRNA翻译的调节主要由eIF 4 E控制，其结合mRNA 5'端的不寻常帽结构。eIF 4 E是重要的，因为它与eIF 4G相互作用，eIF 4G募集核糖体，翻译机器，mRNA。存在替代的eIF 4 E结合蛋白，如4 E-BP，它们阻止eIF 4G与eIF 4 E结合并减少翻译。我建议研究一种最近发现的eIF 4 E结合蛋白，称为4 E-T，它也与eIF 4G竞争eIF 4 E结合。有趣的是，4 E-T和一部分eIF 4 E集中在显微镜下可见的点状细胞质“体”中，称为P体，其缺乏核糖体，被认为是mRNA储存和返回翻译的位点，或者是mRNA降解的位点。尽管有相当大的兴趣，这些P体的作用仍然是谜，虽然它们影响例如病毒复制。我们和其他研究人员已发表和未发表的工作表明，4 E-T调节一组mRNA，但不知道它们编码什么样的蛋白质，也不知道4 E-T是否必须在P体中才能发挥这种控制作用，它在那里结合了什么蛋白质，或者它通过磷酸化修饰对这种控制意味着什么。这些都是我们想用培养的人类细胞来回答的问题。我们相信这项工作是非常及时的，因为异常水平的eIF 4 E（及其磷酸化）已牵连在细胞生长，增殖和癌症的发展，因此调查的一个因素，与eIF 4 E相互作用是很重要的。此外，P体的作用仍然相对未被探索，我们的工作将为我们理解mRNA在这些病灶中的调控做出重大贡献。最后，我们很幸运地与莱斯特MRC毒理学单位的Anne Willis教授合作，进行4 E-T mRNA靶点的部分分析，并扩展和受益于那里的翻译资源数据库，其中包含来自我们和其他研究人员的类似实验的数据。这项工作是新颖的，有针对性的和及时的，其结果将有助于我们理解和可能利用的调控翻译起始的eIF 4 E及其相互作用的因素。

项目成果

The awesome power of ribosome profiling.

• DOI: 10.1261/rna.049908.115
• 发表时间: 2015-04
• 期刊: RNA (New York, N.Y.)
• 作者: Jackson R;Standart N
• 通讯作者: Standart N

The DDX6-4E-T interaction mediates translational repression and P-body assembly.

• DOI: 10.1093/nar/gkw565
• 发表时间: 2016-07-27
• 期刊: Nucleic acids research
• 影响因子: 14.9
• 作者: Kamenska A;Simpson C;Vindry C;Broomhead H;Bénard M;Ernoult-Lange M;Lee BP;Harries LW;Weil D;Standart N
• 通讯作者: Standart N

Investigating the consequences of eIF4E2 (4EHP) interaction with 4E-transporter on its cellular distribution in HeLa cells.

• DOI: 10.1371/journal.pone.0072761
• 发表时间: 2013
• 期刊: PloS one
• 影响因子: 3.7
• 作者: Kubacka D;Kamenska A;Broomhead H;Minshall N;Darzynkiewicz E;Standart N
• 通讯作者: Standart N

Human 4E-T represses translation of bound mRNAs and enhances microRNA-mediated silencing.

• DOI: 10.1093/nar/gkt1265
• 发表时间: 2014-03
• 期刊: Nucleic acids research
• 影响因子: 14.9
• 作者: Kamenska A;Lu WT;Kubacka D;Broomhead H;Minshall N;Bushell M;Standart N
• 通讯作者: Standart N

P-body assembly requires DDX6 repression complexes rather than decay or Ataxin2/2L complexes.

• DOI: 10.1091/mbc.e15-03-0136
• 发表时间: 2015-07-15
• 期刊: Molecular biology of the cell
• 影响因子: 3.3
• 作者: Ayache J;Bénard M;Ernoult-Lange M;Minshall N;Standart N;Kress M;Weil D
• 通讯作者: Weil D