Targeted mRNA degradation in Drosophila spermatogenesis

果蝇精子发生中的靶向 mRNA 降解

• 批准号: BB/I007989/1
• 负责人: Sarah Newbury
• 金额: $ 55.21万
• 依托单位: University of Sussex
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2011
• 资助国家: 英国
• 起止时间: 2011 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FI007989%2F1
• 关键词: Targeted; mRNA; degradation; Drosophila; spermatogenesis

Stem cells have a vast potential in regenerative medicine for the replacement of defective tissue. They therefore offer a potential cure for injuries and also for degenerative diseases such as Alzheimers and Duchenne muscular dystrophy. In the testis, stem cells are required to maintain the supply of sperm, so that the male has potential to produce offspring for much of his life. In recent years, there have been rapid advances in the understanding of the genes which are required to be switched ON to produce self-renewing stem cells that are capable of developing into a wide range of tissues e.g. skin or nervous tissue. However, it is now known that it is also crucially important to make sure that certain genes are switched OFF to prevent stem cells from spontaneously differentiating into particular cell types or developing into cancer cells. We have recently discovered that an enzyme named Pacman, which is involved in the destruction of messenger RNA molecules, is necessary for stem cell function in testis cells of the fruit fly Drosophila. Messenger RNAs are the molecules which instruct the cell to make particular proteins. By comparing mutant flies with normal flies, we have found out that there are fewer testis stem cells in the pacman mutant leading to fewer sperm and offspring. Using various genetic and molecular techniques we have found out that Pacman is likely to destroy particular RNAs that would otherwise prevent stem cell division or cell death. This is interesting as it shows that these mRNAs must somehow be 'tagged' allowing Pacman and its partners to identify, 'hunt down' and destroy these particular RNAs. The specific aim of this project is to understand, in molecular terms, exactly how Pacman and its partners can identify and destroy its correct target RNAs and how this is controlled. The ability of Pacman to selectively destroy target RNAs is not intrinsic to itself as isolated Pacman protein in the test tube cannot distinguish between different RNAs. We know that different RNAs are selected in different tissues so how does Pacman and its partners do this? Our hypothesis is that a particular protein (or a tiny regulatory RNA) binds to a specific feature or the target RNA and 'tags' it for destruction. This tag is then recognised by Pacman and its partner proteins. We propose that Pacman, which is a large protein, can act as a scaffold to assemble other proteins upon it. When the correct partners are assembled, in the correct 3-dimensional shape, then a decapitation enzyme snips off the end of the RNA and the rest is rapidly chewed up. In this project, we aim to find out the details of this destruction pathway, including the ways that the target is tagged. This work will increase our general understanding of the ways that genes are specifically switched off in response to their cellular context. Since Pacman is known to be important in other important cellular events such as wound healing and migration of cell sheets, this work may also shed light on the molecular mechanisms of these processes. Although this project will be carried out using the fruit fly Drosophila, it also has relevance for stem cell function in humans as the cellular processes are surprisingly similar in both organisms. In addition, the Pacman enzyme is extremely similar between flies and humans therefore the insights we gain during this project may help us to improve treatment for fertility and also help us to understand the ways that stem cells remain virtually immortal for the lifetime of the organism. This project will therefore provide valuable insights into the ways in which specific RNAs are targeted in a particular cell type which can be used in the development of new therapeutics.

干细胞在再生医学中具有巨大的潜力，可以替代有缺陷的组织。因此，它们提供了一种潜在的治疗损伤和退行性疾病，如阿尔茨海默病和杜氏肌营养不良症。在睾丸中，需要干细胞来维持精子的供应，这样男性就有潜力在一生的大部分时间里产生后代。近年来，在理解基因方面取得了快速进展，这些基因需要被打开以产生能够发育成广泛组织（例如皮肤或神经组织）的自我更新干细胞。然而，现在已知，确保某些基因被关闭以防止干细胞自发分化为特定细胞类型或发展成癌细胞也至关重要。我们最近发现，一种名为Pacman的酶，参与破坏信使RNA分子，是果蝇睾丸细胞干细胞功能所必需的。信使RNA是指导细胞制造特定蛋白质的分子。通过比较突变果蝇和正常果蝇，我们发现pacman突变体中睾丸干细胞较少，导致精子和后代较少。使用各种遗传和分子技术，我们发现Pacman可能会破坏特定的RNA，否则会阻止干细胞分裂或细胞死亡。这很有趣，因为它表明这些mRNA必须以某种方式被“标记”，允许Pacman及其合作伙伴识别，“追捕”和破坏这些特定的RNA。该项目的具体目标是从分子角度了解Pacman及其合作伙伴如何识别和破坏其正确的靶RNA以及如何控制。Pacman选择性破坏靶RNA的能力不是其本身固有的，因为试管中分离的Pacman蛋白不能区分不同的RNA。我们知道不同的RNA在不同的组织中被选择，那么Pacman及其合作伙伴是如何做到这一点的呢？我们的假设是，一种特定的蛋白质（或一种微小的调节RNA）与一种特定的特征或靶RNA结合，并“标记”它以进行破坏。这个标签然后被Pacman及其伴侣蛋白识别。我们认为，作为一种大蛋白质，Pacman可以充当支架，在其上组装其他蛋白质。当正确的伴侣组装成正确的三维形状时，然后去头酶剪断RNA的末端，其余部分被迅速咀嚼。在这个项目中，我们的目标是找出这种破坏途径的详细信息，包括标记目标的方式。这项工作将增加我们对基因在细胞环境中特异性关闭的方式的总体理解。由于已知Pacman在其他重要的细胞事件中很重要，如伤口愈合和细胞片迁移，这项工作也可能揭示这些过程的分子机制。虽然该项目将使用果蝇果蝇进行，但它也与人类干细胞功能相关，因为两种生物的细胞过程惊人地相似。此外，Pacman酶在苍蝇和人类之间非常相似，因此我们在这个项目中获得的见解可能有助于我们改善生育治疗，也有助于我们了解干细胞在生物体的一生中几乎保持不朽的方式。因此，该项目将为特定RNA在特定细胞类型中的靶向方式提供有价值的见解，这些细胞类型可用于开发新的治疗方法。

项目成果

Identification of circulating microRNAs as diagnostic biomarkers for use in multiple myeloma.

• DOI: 10.1038/bjc.2012.525
• 发表时间: 2012-12-04
• 期刊: BRITISH JOURNAL OF CANCER
• 影响因子: 8.8
• 作者: Jones, C. I.;Zabolotskaya, M. V.;King, A. J.;Stewart, H. J. S.;Horne, G. A.;Chevassut, T. J.;Newbury, S. F.
• 通讯作者: Newbury, S. F.

Severity of Systemic Inflammatory Response Syndrome Affects the Blood Levels of Circulating Inflammatory-Relevant MicroRNAs.

• DOI: 10.3389/fimmu.2017.01977
• 发表时间: 2017
• 期刊: Frontiers in immunology
• 影响因子: 7.3
• 作者: Caserta S;Mengozzi M;Kern F;Newbury SF;Ghezzi P;Llewelyn MJ
• 通讯作者: Llewelyn MJ

Circulating MicroRNA Biomarkers in Melanoma: Tools and Challenges in Personalised Medicine.

• DOI: 10.3390/biom8020021
• 发表时间: 2018-04-26
• 期刊: Biomolecules
• 影响因子: 5.5
• 作者: Mumford SL;Towler BP;Pashler AL;Gilleard O;Martin Y;Newbury SF
• 通讯作者: Newbury SF

Circulating Plasma microRNAs can differentiate Human Sepsis and Systemic Inflammatory Response Syndrome (SIRS).

• DOI: 10.1038/srep28006
• 发表时间: 2016-06-20
• 期刊: Scientific reports
• 影响因子: 4.6
• 作者: Caserta S;Kern F;Cohen J;Drage S;Newbury SF;Llewelyn MJ
• 通讯作者: Llewelyn MJ

DIS3 isoforms vary in their endoribonuclease activity and are differentially expressed within haematological cancers.

• DOI: 10.1042/bcj20170962
• 发表时间: 2018-06-29
• 期刊: The Biochemical journal
• 作者: Robinson SR;Viegas SC;Matos RG;Domingues S;Bedir M;Stewart HJS;Chevassut TJ;Oliver AW;Arraiano CM;Newbury SF
• 通讯作者: Newbury SF