Regulation of Human Epidermal Tumorigenesis by the mRNA Degradation Pathway

项目总结/摘要 背景：调节表皮稳态的转录机制， 肿瘤已经很好地建立，但最近我们发现，后- 转录机制在维持表皮自我更新中起重要作用。 我们已经表明，由外泌体介导的3 '-5' mRNA降解途径， 复合物是维持表皮自我更新所必需的。特别是外泌体 亚基EXOSC 7、EXOSC 9和EXSCO 10是防止早产所必需的。 表皮干细胞的分化通过靶向和降解转录本， 寻找有效的促分化转录因子。 目的/假设：本提案旨在了解 使用RAS驱动的人类皮肤来控制从正常皮肤到肿瘤性皮肤的进展 表皮肿瘤模型。我们的初步数据表明，一个5亚基外泌体 亚复合物在肿瘤起始期间上调并靶向/降解转录物 编码抑制肿瘤生长和存活的因子。我们的目标是 表征每个肿瘤诱导的外泌体亚基在肿瘤进展中的作用， 正常到肿瘤性皮肤。此外，我们试图确定具体的成绩单， 每个外泌体亚单位在肿瘤起始期间结合以促进肿瘤发生。 具体目的：（1）确定外泌体亚单位在从 正常至肿瘤性皮肤和（2）鉴定和表征相关的转录物 与外泌体亚基结合。 研究设计：为了在更临床相关的环境中研究表皮稳态，我们 生成三维完整的人类皮肤，包含人类表皮细胞（其 已被永久敲除外泌体亚基）， 真皮基质和基底膜，在免疫受损小鼠上再生。 通过使用该模型，我们可以对外泌体亚基进行功能丧失实验 以表征它们在表皮生长、分化 并发展为肿瘤。我们将使用CLIP-Seq来确定相关的RNA 与外泌体亚基的关系。

Project Summary/Abstract Background: Transcriptional mechanisms that regulate epidermal homeostasis and neoplasia have been well established but recently we have discovered that post- transcriptional mechanisms play prominent roles in maintaining epidermal self-renewal. We have shown that the 3'-5' mRNA degradation pathway mediated by the exosome complex is necessary to maintain epidermal self-renewal. Specifically, the exosome subunits, EXOSC7, EXOSC9, and EXSCO10 are necessary to prevent premature differentiation of epidermal stem cells by targeting and degrading transcripts that code for potent pro-differentiation transcription factors. Objective/hypothesis: This proposal seeks to understand the molecular mechanisms governing the progression from normal to neoplastic skin using a RAS driven human epidermal tumor model. Our preliminary data suggests that a 5 subunit exosome subcomplex is upregulated during tumor initiation and targets/degrades transcripts coding for factors that would inhibit tumor growth and survival. Our objective is to characterize the role of each tumor induced exosome subunit in the progression of normal to neoplastic skin. Furthermore we seek to determine the specific transcripts that each exosome subunit binds during tumor initiation to promote tumorigenesis. Specific Aims: (1) To determine the role of exosome subunits on the progression from normal to neoplastic skin and (2) to identify and characterize the transcripts associated with exosome subunits. Study Design: To study epidermal homeostasis in a more clinically relevant setting, we generate 3-dimensionally intact human skin, containing human epidermal cells (that have been permanently knocked down for exosome subunits) in the context of human dermal stroma and basement membrane, regenerated on immune compromised mice. By using this model, we can perform loss of function experiments on exosome subunits in regenerated human skin to characterize their role in epidermal growth, differentiation, and progression to neoplasia. We will use CLIP-Seq to determine the RNAs associated with the exosome subunits during the progression from normal to neoplastic epidermis.