Post-Transcriptional Regulators of Epidermal Homeostasis and Neoplasia

项目概要/摘要 背景：调节表皮稳态的转录机制 已经得到证实，但最近我们发现转录后 机制在维持表皮自我更新中起重要作用。我们有 表明RNA解旋酶DDX 6是维持表皮自我更新所必需的 通过mRNA降解和翻译途径。通过与特定的 翻译途径的成员DDX 6结合并介导 自我更新和增殖转录本以维持自我更新。DDX 6也 与mRNA降解蛋白结合， 转录本，以促进其降解，防止过早分化。 目的/假设：本提案旨在了解 表皮稳态和肿瘤起始通过转录后机制。 我们以前鉴定了与DDX 6相关的蛋白质，我们的目标是 表征每种蛋白质在调节表皮生长、分化 和进展为瘤形成以及作用机制。此外我们 试图确定DDX 6及其相关复合物的特异性转录本， 在体内平衡和肿瘤起始期间结合。 具体目的：（1）研究DDX 6相关蛋白在表皮细胞凋亡中的作用， 体内平衡和肿瘤起始以及（2）鉴定和表征转录物 与DDX 6复合物有关。 研究设计：为了在更临床相关的环境中研究表皮稳态， 我们生成了三维完整的人类皮肤， (that已经被DDX 6或其相关的 蛋白质）在人真皮基质和基底膜的情况下， 在免疫受损的小鼠身上再生通过这个模型，我们可以 DDX 6及其相关蛋白在再生肝细胞中的功能丧失实验 人类皮肤，以表征其在表皮生长、分化和 进展为瘤形成。我们还将使用CLIP-Seq来确定RNA 与DDX 6复合物相关，从正常进展为肿瘤 表皮

Project Summary/Abstract Background: Transcriptional mechanisms that regulate epidermal homeostasis 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 RNA helicase DDX6 is necessary to maintain epidermal self-renewal through the mRNA degradation and translation pathway. By associating with specific members of the translation pathway DDX6 binds to and mediates the translation of self- renewal and proliferation transcripts to maintain self-renewal. DDX6 also associates with mRNA degradation proteins to bind differentiation-inducing transcripts to promote their degradation to prevent premature differentiation. Objective/hypothesis: This proposal seeks to understand the regulation of epidermal homeostasis and tumor initiation through post-transcriptional mechanisms. We previously identified proteins associated with DDX6 and our objective is to characterize the role of each protein in regulating epidermal growth, differentiation, and progression to neoplasia as well as the mechanisms of action. Furthermore we seek to determine the specific transcripts that DDX6 and its associated complexes bind during homeostasis and tumor initiation. Specific Aims: (1) To determine the role of DDX6 associated proteins on epidermal homeostasis and tumor initiation and (2) to identify and characterize the transcripts associated with DDX6 complexes. 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 either DDX6 or its associated proteins) 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 DDX6 and its associated proteins in regenerated human skin to characterize their role in epidermal growth, differentiation, and progression to neoplasia. We will also use CLIP- Seq to determine the RNAs associated with DDX6 complexes during the progression from normal to neoplastic epidermis.