Role of ER-membrane contacts in biogenesis of RNA-containing EVs

项目1总结： 细胞外小泡(EV)携带多种RNA，包括编码和非编码RNA。这些 RNA有可能影响细胞和组织的表型。事实上，对于miRNAs来说，现在有很多 EV携带的miRNA控制受体细胞中基因表达和功能的例子。RNA-Seq分析 与细胞内容相比，EVS中的一些RNA表现出特定的富集性。然而，非常 人们对RNA进入电动汽车的具体机制知之甚少。 基于几项研究，目前的研究范式是RNA结合蛋白(RBPs)负责 用于在电动汽车中选择性和特定地包括RNA。然而，这些限制性商业惯例如何连接到蜂窝 膜被结合到脱落的微囊泡(MVS)或晚内体衍生的外切体中 未知。一个值得注意的发现是，在EV中发现的许多限制性商业惯例通常与内质 细胞中的网状结构(ER)，这表明ER在将这些部分转移到其他细胞器中具有潜在的作用。 此外，rna诱导的沉默复合体组装在相关的mrna-核糖体复合体上。 与ER(粗略ER)的关系，建议了一条miRNA-RBP与ER相关联的路线，并随后与其他 膜。基于这些发现和我们的初步数据，我们假设内质网质膜 (PM)和ER-多囊泡内小体(MVE)膜接触部位(MCS)是RNA转运的关键 和限制性商业惯例进入脱落的微囊和外体。我们进一步假设信号和脂质转移 在这些接触处发生的事件进一步调节了RNA进入囊泡的运输。我们将测试这些 假设和确定MCS对RNA转移到受体细胞和结直肠癌生长的影响 和西妥昔单抗耐药。

Project 1 Summary: Extracellular vesicles (EVs) carry a variety of RNAs, including both coding and noncoding RNAs. These RNAs have the potential to influence cell and tissue phenotypes. Indeed for miRNAs there are now many examples of EV-carried miRNA controlling gene expression and function in recipient cells. RNA-Seq analyses have demonstrated specific enrichment of some RNAs in EVs, compared to the cellular content. However, very little is known about the specific mechanisms by which RNA is transported into EVs. The current paradigm, based on several studies, is that RNA-binding proteins (RBPs) are responsible for the selective and specific inclusion of RNAs in EVs. However, how those RBPs connect to cellular membranes to be incorporated into shed microvesicles (MVs) or late endosome-derived exosomes is unknown. A notable finding is that many RBPs identified in EVs are typically associated with the endoplasmic reticulum (ER) in cells, suggesting a potential role for the ER in transfer of those moieties to other organelles. Furthermore, the RNA-induced silencing complex is assembled on mRNA-ribosome complexes associated with the ER (rough ER), suggesting a route for miRNA-RBP association with the ER and subsequently other membranes. Based on these findings and our preliminary data, we hypothesize that ER-plasma membrane (PM) and ER-multivesicular endosome (MVE) membrane contact sites (MCS) are critical for transfer of RNAs and RBPs into shed microvesicles and exosomes. We further hypothesize that signaling and lipid transfer events taking place at these contacts further regulate RNA transport into vesicles. We will test these hypotheses and determine the impact of MCS on transfer of RNAs to recipient cells and CRC tumor growth and cetuximab resistance.