Regulation of epidermal growth and differentiation through mRNA export

项目摘要/摘要 背景：调节表皮动态平衡的转录机制有 已经很好地建立了，但最近我们发现了信使核糖核酸输出机制 在维持表皮自我更新方面发挥突出作用。我们已经证明了RBM15 ZC3H18只在干细胞和祖细胞中与Nxf1导出子结合 在分化的细胞中与Nxf1相关。此关联允许RBM15或ZC3H18 控制与表皮生长有关的关键转录本的mRNA输出 差异化。 目的/假设：这项提议试图了解表皮的调节 干细胞和祖细胞通过转录后的自我更新和分化 机械装置。我们已经确定了出口所必需的RNA结合蛋白 自我更新的mRNAs以促进表皮自我更新。同样，我们已经确定了 将诱导分化的mRNAs输出到 促进表皮分化。此外，这些蛋白质的突变可能导致 皮肤的克隆性扩张是由于改变了对表皮生长的调节和 差异化。 具体目标：(1)确定RBM15和ZC3H18在表皮生长中的作用 和差异化。(2)确定RBM15和ZC3H18的分子机制 野生型和突变型蛋白影响表皮的动态平衡。 研究设计：为了在更具临床相关性的环境中研究表皮动态平衡，我们 生成三维完整的人类皮肤，包含人类表皮细胞(即 已因RBM15或ZC3H18)在人类的上下文中被永久拆除 真皮基质和基底膜，在免疫受损的小鼠身上再生。 利用该模型，我们可以对RBM15或ZC3H18进行功能损失实验 以确定它们在表皮生长和生长中的作用 差异化。我们还将使用RNA免疫沉淀，然后是下一代 测序以确定与这些蛋白质相关的RNA。

Project Summary/Abstract Background: Transcriptional mechanisms that regulate epidermal homeostasis have been well established but recently we have discovered that mRNA export mechanisms play prominent roles in maintaining epidermal self-renewal. We have shown that RBM15 associates with the NXF1 exporter only in stem and progenitor cells while ZC3H18 associates with NXF1 in differentiated cells. This association allows RBM15 or ZC3H18 to control the mRNA export of key transcripts involved in epidermal growth and differentiation. Objective/hypothesis: This proposal seeks to understand the regulation of epidermal stem and progenitor cell self-renewal and differentiation through post-transcriptional mechanisms. We have identified RNA binding proteins that are necessary for the export of self-renewal mRNAs to promote epidermal self-renewal. Similarly we have identified RNA binding proteins that are necessary to export differentiation inducing mRNAs to promote epidermal differentiation. Furthermore mutations in these proteins can lead to clonal expansion of the skin due to altered regulation of epidermal growth and differentiation. Specific Aims: (1) To determine the role of RBM15 and ZC3H18 on epidermal growth and differentiation. (2) To determine the molecular mechanisms of RBM15 and ZC3H18 wildtype and mutant proteins impact on epidermal homeostasis. 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 RBM15 or ZC3H18) 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 RBM15 or ZC3H18 in regenerated human skin to characterize their role in epidermal growth and differentiation. We will also use RNA immunoprecipitations followed by next generation sequencing to determine the RNAs associated with these proteins.