Regulation of pre-mRNA splicing fidelity by the Nineteen Complex (NTC)

十九复合物 (NTC) 对前 mRNA 剪接保真度的调节

• 批准号: BB/I019510/1
• 负责人: Raymond O'Keefe
• 金额: $ 40.04万
• 依托单位: University of Manchester
• 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%2FI019510%2F1
• 关键词: Regulation; pre; mRNA; splicing; fidelity

Genes within cells are copied into a pre-messenger RNA (pre-mRNA) which is used as a template for protein production. All the information contained within genes is not required for making proteins. The unwanted information, therefore, must be removed from the pre-mRNA before it is used for protein production. The unwanted information is removed, or 'spliced', from pre-mRNA by a process similar to the editing of unwanted frames from a film. One end of the region to be removed is first cut then the other end is cut while the two remaining pieces are 'spliced' together. This 'splicing' of the pre-mRNA is very important because it must occur accurately in order for functional proteins to be produced. The regulation of splicing is essential for all aspects of human biology. Splicing is required for proper embryo development and differentiation of all tissues and organs. Regulation of pre-mRNA splicing is also vital for organisms to respond to their environment and adapt to stresses and nutrient deprivation. Defects in pre-mRNA splicing are associated with a wide range of diseases including diabetes, cancer and age related diseases. Alternative splicing has also allowed humans to expand their cellular complexity without having to increase the size of their genome. The work that will be undertaken here will address how the process of splicing occurs as there are still some key unanswered questions on how pre-mRNA splicing is regulated. Splicing is carried out by a large RNA/protein complex called the spliceosome. The spliceosome must arrange itself into specific conformations to identify and 'splice' out the unwanted regions. An increasing amount of evidence points to a complex of proteins called the NineTeen Complex, or NTC, that associates with the spliceosome to act as an essential regulator of spliceosome function. In work leading up to this proposal we have identified an NTC protein called Cwc2 that provides the direct link between proteins of the NTC and the active site of the spliceosome. Discovering this link between the NTC and the spliceosome has implications for how scientists now understand the mechanisms of pre-mRNA splicing regulation. As we are the first researchers to discover this link we now have an advantage in discovering exactly how this link contributes to the regulation of pre-mRNA splicing. Work proposed during the tenure of this research grant will address how Cwc2 is involved in regulating spliceosome conformations required for the two steps of splicing through a set of detailed molecular experiments. As the basic mechanisms of pre-mRNA splicing are identical in yeast and human cells, we avoid using animals in our research by using yeast cells to perform our experiments .

细胞内的基因被复制到前信使 RNA (pre-mRNA) 中，用作蛋白质生产的模板。基因中包含的所有信息并不是制造蛋白质所必需的。因此，在用于蛋白质生产之前，必须从前 mRNA 中去除不需要的信息。通过类似于剪辑电影中不需要的帧的过程，从前 mRNA 中去除或“拼接”不需要的信息。首先切割要去除的区域的一端，然后切割另一端，同时将剩余的两块“拼接”在一起。 mRNA 前体的这种“剪接”非常重要，因为它必须准确发生才能产生功能性蛋白质。剪接的调控对于人类生物学的各个方面都是至关重要的。剪接对于胚胎的正常发育以及所有组织和器官的分化是必需的。 mRNA 前体剪接的调节对于生物体对其环境做出反应并适应压力和营养缺乏也至关重要。 mRNA 前体剪接缺陷与多种疾病相关，包括糖尿病、癌症和年龄相关疾病。选择性剪接还使人类能够扩大细胞的复杂性，而无需增加基因组的大小。这里将要开展的工作将解决剪接过程是如何发生的，因为关于前 mRNA 剪接如何调节仍存在一些未解答的关键问题。剪接是由称为剪接体的大型 RNA/蛋白质复合物进行的。剪接体必须将自身排列成特定的构象，以识别并“剪接”掉不需要的区域。越来越多的证据表明，有一种称为 NineTeen Complex（NTC）的蛋白质复合物，它与剪接体结合，充当剪接体功能的重要调节剂。在提出这一建议的工作中，我们鉴定了一种称为 Cwc2 的 NTC 蛋白，它提供了 NTC 蛋白与剪接体活性位点之间的直接连接。发现 NTC 和剪接体之间的这种联系对于科学家现在如何理解前 mRNA 剪接调节机制具有重要意义。由于我们是第一批发现这种联系的研究人员，因此我们现在有优势准确地发现这种联系如何有助于前体 mRNA 剪接的调节。在这项研究资助期间提出的工作将通过一系列详细的分子实验来解决 Cwc2 如何参与调节剪接两个步骤所需的剪接体构象。由于前体 mRNA 剪接的基本机制在酵母和人类细胞中是相同的，因此我们通过使用酵母细胞进行实验来避免在我们的研究中使用动物。

项目成果

The NineTeen Complex (NTC) and NTC-associated proteins as targets for spliceosomal ATPase action during pre-mRNA splicing.

• DOI: 10.1080/15476286.2015.1008926
• 发表时间: 2015
• 期刊: RNA biology
• 影响因子: 4.1
• 作者: de Almeida RA;O'Keefe RT
• 通讯作者: O'Keefe RT

Compound heterozygosity of low-frequency promoter deletions and rare loss-of-function mutations in TXNL4A causes Burn-McKeown syndrome.

• DOI: 10.1016/j.ajhg.2014.10.014
• 发表时间: 2014-12
• 期刊: American journal of human genetics
• 影响因子: 9.8
• 作者: D. Wieczorek;W. Newman;T. Wieland;Tea Berulava;Maria Kaffe;D. Falkenstein;C. Beetz;E. Graf;T. Schwarzmayr;S. Douzgou;J. Clayton-Smith;Sarah B. Daly;S. Williams;S. Bhaskar;J. Urquhart;Beverley H Anderson;J. O’Sullivan;O. Boute;Jasmin Gundlach;J. Czeschik;A. V. van Essen;F. Hazan;Sarah S. Park;A. Hing;A. Kuechler;D. Lohmann;K. Ludwig;E. Mangold;L. Steenpass;M. Zeschnigk;J. Lemke;C. Lourenço;U. Hehr;E. Prott;M. Waldenberger;A. Böhmer;B. Horsthemke;R. O’Keefe;T. Meitinger;J. Burn;H. Lüdecke;T. Strom

Remodeling of U2-U6 snRNA helix I during pre-mRNA splicing by Prp16 and the NineTeen Complex protein Cwc2.

• DOI: 10.1093/nar/gku431
• 发表时间: 2014-07
• 期刊: Nucleic acids research
• 影响因子: 14.9
• 作者: Hogg R;de Almeida RA;Ruckshanthi JP;O'Keefe RT
• 通讯作者: O'Keefe RT