The roles of midbody associated mRNAs in regulating cell proliferation and differentiation

中体相关mRNA在调节细胞增殖和分化中的作用

• 批准号: 10624620
• 负责人: Rytis Prekeris
• 金额: $ 7.32万
• 依托单位: UNIVERSITY OF COLORADO DENVER
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-20 至 2025-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10624620
• 关键词: Binding Proteins; CENP-E protein; CRISPR screen; Cell Differentiation process; Cell Proliferation; Cells; Complex; Cytokinesis; Excision; Extracellular Space; Goals; Maps; Mediating; Messenger RNA; Microtubules; Mitosis; Mitotic; Molecular; Mothers; Organelles; Play; Proteins; Proteomics; Protocols documentation; RNA-Binding Proteins; Resolution; Role; Signal Pathway; Signal Transduction; Structure; System; Testing; Thinness; Translations; Untranslated Regions; base; cancer cell; daughter cell; novel; recruit; stem cells; transcriptome sequencing

Project Summary This Project Summary/Abstract was originally submitted with R01 GM143774 and is included here unchanged to satisfy submission system requirements. During mitosis, the mother cell divides by the formation of a cleavage furrow, leaving two daughter cells connected by a thin intercellular bridge. The resolution of this bridge, abscission, leads to the separation of the two daughter cells. During ingression of the cleavage furrow, the central spindle microtubules are compacted to form a structure known as the midbody (MB). It is now well established that MB regulates cytokinesis by recruiting abscission-mediating proteins, such as ESCRT complex, as well as several regulators of abscission checkpoint. Until recently, the MB was thought to be discarded after division by releasing it into extracellular space. However, recently it was shown that MBs accumulate in stem and cancer cells after mitosis has been completed (called MBsomes) and it has been proposed that MBsomes function as novel signalling platforms that regulate cell differentiation and proliferation. Recently we developed a protocol for purification of post- mitotic MBs and completed their proteomic and RNAseq analyses that led to identification of several mRNAs and mRNA-binding proteins that accumulate at the MB. Importantly, MB-enriched mRNAs encode several ESCRT complex subunits, as well as proteins that stimulate cell proliferation. Furthermore, we show that these MB-enriched mRNAs can be transferred to the neighboring cells via post-mitotic MB internalization. Based on all of these findings, we hypothesize that targeting of selected mRNAs to the MB during cytokinesis play a key role in regulating cell abcission and post-mitotic MBsome signaling. Here we propose three specific aims to test this hypothesis. First, we will map and characterize the domain(s) within 3'-UTR that are needed for mRNA targeting during cytokinesis. We will then use candidate approach, as well as proteomic and CRISPR screens, to identify RNA-binding proteins that interact with these 3”-UTR domains and regulate mRNA targeting and localized translation at the MB. Second, we will test the possibility that MB accumulation/translation of ESCRT mRNAs mediates ESCRP complex targeting to the MB. Third, we will test whether MBsome-dependent transfer of specific mRNAs, such as mRNA encoding proliferation regulator CENP-E contribute to MBsome-induced cell proliferation.

项目概要 本项目摘要/摘要最初与 R01 GM143774 一起提交，并原封不动地包含在此处 以满足提交系统的要求。 在有丝分裂过程中，母细胞通过形成卵裂沟进行分裂，留下两个子细胞 由薄的细胞间桥连接。这座桥的解决，即分离，导致了分离 两个子细胞。在卵裂沟进入期间，中央纺锤体微管被压实 形成称为中体 (MB) 的结构。现已明确，MB 通过以下方式调节胞质分裂： 招募脱落介导蛋白，例如 ESCRT 复合物，以及几种脱落调节因子 检查站。直到最近，MB 还被认为在分裂后通过释放到细胞外而被丢弃。 空间。然而，最近的研究表明，有丝分裂后，MB 在干细胞和癌细胞中积累。 已完成（称为 MBsomes），并且有人提议 MBsomes 作为新型信号平台 调节细胞分化和增殖。最近我们开发了一种纯化后的方案 有丝分裂 MB 并完成了蛋白质组学和 RNAseq 分析，从而鉴定了几种 mRNA 以及在 MB 处积累的 mRNA 结合蛋白。重要的是，富含 MB 的 mRNA 编码多种 ESCRT 复合亚基以及刺激细胞增殖的蛋白质。此外，我们表明这些 富含 MB 的 mRNA 可以通过有丝分裂后 MB 内化转移到邻近细胞。基于 所有这些发现，我们假设在胞质分裂过程中，选定的 mRNA 靶向 MB 在调节细胞脱落和有丝分裂后 MBsome 信号传导中发挥关键作用。这里我们提出三个 具体目的是检验这一假设。首先，我们将映射并表征 3'-UTR 内的域 胞质分裂期间 mRNA 靶向所需。然后我们将使用候选方法以及蛋白质组学和 CRISPR 筛选，以识别与这些 3”-UTR 结构域相互作用并调节的 RNA 结合蛋白 MB 处的 mRNA 靶向和本地化翻译。其次，我们将测试MB的可能性 ESCRT mRNA 的积累/翻译介导 ESCRP 复合物靶向 MB。第三，我们要测试 特定 mRNA 是否依赖 MBsome 转移，例如编码增殖调节因子的 mRNA CENP-E 有助于 MBsome 诱导的细胞增殖。