Deciphering the functions of the RNA binding protein T-STAR in mouse development

破译 RNA 结合蛋白 T-STAR 在小鼠发育中的功能

• 批准号: BB/K018957/1
• 负责人: David Elliott
• 金额: $ 40.68万
• 依托单位: Newcastle University
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2013
• 资助国家: 英国
• 起止时间: 2013 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FK018957%2F1
• 关键词: Deciphering; functions; RNA; binding; protein

RNA binding proteins play a key role in organising gene expression in the cell, and control the important process of alternative splicing through which a single gene can produce multiple different mRNAs and proteins. Most genes produce alternatively spliced mRNAs, and some individual genes also produce many splice isoforms. In the most common kind of alternative splicing in mammals entire exons are either spliced in or left out of the mRNA. The insertion of alternative exons can be very important for downstream protein function. Alternative splicing is controlled by nuclear RNA binding proteins. Many nuclear RNA binding proteins exist as families of closely related proteins, but whether family members have different or overlapping functions is usually a mystery. A good example of this is the T-STAR RNA binding protein, which is in the same family as Sam68. T-STAR and Sam68 are important for normal health. The T-STAR gene becomes amplified in some cases of medulloblastoma, and Sam68 becomes sequestered in the severe neurodegenerative disorder Fragile X Tremor Associated Ataxia. T-STAR and Sam68 function similarly in transfected cells to regulate splicing of the same target exons. We want to establish the roles of endogenous T-STAR protein and if it interacts with Sam68 to control splicing in vivo. Three aspects of this project are timely: (1) We have recently made a T-STAR null mouse to use in the current project. There is a window of opportunity here where we are the only group worldwide with a T-STAR null mouse (we have submitted our mouse to the European Mouse Mutant Archive for release next August). (2) In preliminary data using a powerful high throughput approach we have identified splicing defects in the knockout mice. The four already identified alternative exons which we have found to be controlled by T-STAR in the mouse brain include alternative exons in the Neurexin genes which encode proteins essential for wiring up the nervous system. The Neurexin genes have regional splicing patterns in the wild type brain, but this regional control totally breaks down in the absence of T-STAR protein. This indicates that T-STAR is the master regulator controlling this regional splicing pattern. (3) Through international collaboration with the Sette group in Rome we have access a Sam68 null allele to cross in to our T-STAR null mouse. Hence we will be able to analyse changes in phenotype and identify splicing defects in double knockout mice (missing Sam68 and T-STAR).Aims and objectives. In this current project we will comprehensively identify T-STAR target RNAs to discover if T-STAR predominantly regulates synaptic proteins or is responsible for establishing regional patterns of splicing in the brain (or both). We will establish if T-STAR and Sam68 function redundantly in the control of some target exons, and to what extent they overlap in expression in the brain. We will map where T-STAR binds RNA and work out the important features through which this binding regulates alternative splicing. We will examine the phenotypic consequences for the mouse when alternative splicing events are blocked by removing T-STAR. Finally we will address whether T-STAR and Sam68 work redundantly to control important developmental steps, particularly in the testis where both proteins are expressed at high levels as well as the brain. We expect that the results of this project will be significant in understanding how splicing factors interact to enable flexible use of information in the genome and the development of complex tissues like the brain and specialised cell types. Our project will discover new regulated targets of splicing control, interrogate mechanisms of regulation and phenotypic consequences when this is blocked. The main beneficiaries from this work will be scientists interested in gene expression, scientists and students who will be trained and members of the public that we will engage.

RNA结合蛋白在组织细胞中的基因表达方面起着关键作用，并控制着重要的选择性剪接过程，通过该过程，单个基因可以产生多个不同的mRNAs和蛋白质。大多数基因产生交替剪接的mRNA，一些单独的基因也产生许多剪接异构体。在哺乳动物中最常见的选择性剪接中，整个外显子要么被剪接进信使核糖核酸，要么被排除在信使核糖核酸之外。替代外显子的插入对于下游蛋白质的功能可能非常重要。选择性剪接由核RNA结合蛋白控制。许多核RNA结合蛋白以密切相关的蛋白质家族的形式存在，但家族成员是否具有不同或重叠的功能通常是一个谜。T-star RNA结合蛋白就是一个很好的例子，它与Sam68属于同一家族。T-STAR和Sam68对正常健康很重要。在某些髓母细胞瘤中，T-STAR基因被扩增，而在严重的神经退行性疾病脆性X震颤相关性共济失调中，Sam68变得孤立。T-STAR和Sam68在转基因细胞中的功能相似，以调节相同目标外显子的剪接。我们想要确定内源性T-star蛋白的作用，以及它是否与Sam68相互作用来控制体内的剪接。这个项目有三个方面是及时的：(1)我们最近制造了一个T-STAR空鼠标，用于当前项目。这里有一个机会之窗，我们是世界上唯一拥有T-STAR空鼠标的团体(我们已经将我们的鼠标提交给欧洲老鼠突变档案馆，准备明年8月发布)。(2)在使用强大的高通量方法的初步数据中，我们已经在基因敲除小鼠中发现了剪接缺陷。我们在小鼠大脑中发现了四个由T-STAR控制的替代外显子，其中包括neuresin基因中的替代外显子，它编码连接神经系统所必需的蛋白质。在野生型大脑中，neuresin基因具有区域剪接模式，但这种区域控制在缺乏T-star蛋白的情况下完全崩溃。这表明T-STAR是控制这种区域剪接模式的主调控因子。(3)通过与罗马赛特小组的国际合作，我们获得了一个Sam68零等位基因，以杂交到我们的T-STAR零小鼠。因此，我们将能够分析双基因敲除小鼠(缺失Sam68和T-STAR)的表型变化并识别剪接缺陷。在这个目前的项目中，我们将全面识别T-STAR靶RNA，以发现T-STAR是否主要调节突触蛋白或负责在大脑中建立区域剪接模式(或两者兼而有之)。我们将确定T-STAR和Sam68是否在某些目标外显子的控制中冗余地发挥作用，以及它们在大脑中的表达重叠程度。我们将绘制T-STAR与RNA结合的位置，并找出这种结合调节选择性剪接的重要特征。我们将研究当选择性剪接事件通过移除T-STAR而被阻止时，对小鼠的表型后果。最后，我们将讨论T-STAR和Sam68是否冗余地控制重要的发育步骤，特别是在睾丸和大脑中，这两种蛋白都高水平表达。我们预计，该项目的结果将对理解剪接因子如何相互作用以灵活使用基因组中的信息以及脑和特殊细胞类型等复杂组织的发育具有重要意义。我们的项目将发现剪接控制的新调控靶点，询问调控机制和当这被阻止时的表型后果。这项工作的主要受益者将是对基因表达感兴趣的科学家、将接受培训的科学家和学生以及我们将参与的公众成员。

项目成果

The tissue-specific RNA binding protein T-STAR controls regional splicing patterns of neurexin pre-mRNAs in the brain.

• DOI: 10.1371/journal.pgen.1003474
• 发表时间: 2013-04
• 期刊: PLoS genetics
• 影响因子: 4.5
• 作者: Ehrmann I;Dalgliesh C;Liu Y;Danilenko M;Crosier M;Overman L;Arthur HM;Lindsay S;Clowry GJ;Venables JP;Fort P;Elliott DJ
• 通讯作者: Elliott DJ

The cancer-associated cell migration protein TSPAN1 is under control of androgens and its upregulation increases prostate cancer cell migration.

• DOI: 10.1038/s41598-017-05489-5
• 发表时间: 2017-07-12
• 期刊: Scientific reports
• 影响因子: 4.6
• 作者: Munkley J;McClurg UL;Livermore KE;Ehrmann I;Knight B;Mccullagh P;Mcgrath J;Crundwell M;Harries LW;Leung HY;Mills IG;Robson CN;Rajan P;Elliott DJ
• 通讯作者: Elliott DJ

Structural basis of RNA recognition and dimerization by the STAR proteins T-STAR and Sam68.

• DOI: 10.1038/ncomms10355
• 发表时间: 2016-01-13
• 期刊: Nature communications
• 影响因子: 16.6
• 作者: Feracci M;Foot JN;Grellscheid SN;Danilenko M;Stehle R;Gonchar O;Kang HS;Dalgliesh C;Meyer NH;Liu Y;Lahat A;Sattler M;Eperon IC;Elliott DJ;Dominguez C
• 通讯作者: Dominguez C

Differential expression of paralog RNA binding proteins establishes a dynamic splicing program required for normal cerebral cortex development

• DOI: 10.1093/nar/gkae071
• 发表时间: 2024-02-07
• 期刊: NUCLEIC ACIDS RESEARCH
• 影响因子: 14.9
• 作者: Cesari，Eleonora;Farini，Donatella;Sette，Claudio
• 通讯作者: Sette，Claudio

Analysis of human ES cell differentiation establishes that the dominant isoforms of the lncRNAs RMST and FIRRE are circular.

• DOI: 10.1186/s12864-018-4660-7
• 发表时间: 2018-04-20
• 期刊: BMC genomics
• 影响因子: 4.4
• 作者: Izuogu OG;Alhasan AA;Mellough C;Collin J;Gallon R;Hyslop J;Mastrorosa FK;Ehrmann I;Lako M;Elliott DJ;Santibanez-Koref M;Jackson MS
• 通讯作者: Jackson MS