MAP kinases regulate involucrin gene expression

Involucrin是角质形成细胞角化包膜的重要前体。在表皮中，总蛋白的表达 蛋白质和mRNA是角质形成细胞特有的和依赖于分化的(即，表达仅限于 超基底层)。我们的转基因小鼠和细胞培养研究表明，人总蛋白(HINV)基因 上游调控区域对于该表达和该区域中特定DNA元件bindAPI来说是足够的， C/EBP和SPL转录因子驱动表达。在过去的资助期间，我们确定了一个新的PKC， RAS、MEKK1、MEK3和p388-Erk1/2信号通路调节API、Sp1和P388的活性和水平 C/EBP转录因子增加了总蛋白基因的表达。尽管识别这个级联代表一个 我们对分化相关基因调控的理解向前迈进了一大步，我们对这一信号的理解 转导级联反应还远未完成。在确立了这一途径的重要性之后，我们现在建议 旨在扩大这些发现的实验。我们的研究表明，包括PKC5在内的新的PKC亚型是 正常角质形成细胞hINV基因表达的主要激活物及分化因子触发酪氨酸 通过磷酸化PKC8来调节基因的表达。然而，PKC8酪氨酸磷酸化在细胞周期中的重要性 角质形成细胞是复杂的、有争议的，而且还没有得到彻底的研究。我们的第一个主要目标是确定 PKC8酪氨酸磷酸化对PKC8活性、亚细胞定位和调控能力的影响 下行信令事件。一个重要的观察结果是正常人角质形成细胞中的信号转导 在p388上的MAPK级别收敛。我们的初步研究表明，MEK3和MEK3的差异相互作用 带有P38S的MEK6可能调节角质形成细胞分化和凋亡之间的平衡。这个 这项研究的第二个主要目标是表征这些激酶在调节平衡方面的作用 分化和细胞凋亡。我们将特别关注PKC8、MEK3和p388在调控中的作用 角质形成细胞分化。这项提议的总体目标是扩大我们对分子的认识 驱动角质形成细胞分化的机制。这项工作特别重要，因为这种有丝分裂原被激活 蛋白激酶(MAPK)信号转导通路参与调节角质形成细胞的存活、分化、 死亡和蜕变。人们希望，对这种级联的更好理解将导致新的有效的治疗方法。 治疗皮肤病。

Involucrin is an important precursor of the keratinocyte cornified envelope. In epidermis, expression of involucrin protein and mRNA is keratinocyte-specific and differentiation-dependent (i.e., expression is confined to the suprabasal layers). Our transgenic mouse and cell culture studies show that the human involucrin (hINV) gene upstream regulatory region is sufficient for this expression and that specific DNA elements in this region bindAPI, C/EBP and Spl transcription factors to drive expression. During the past funding period we identified a novelPKC, Ras, MEKK1, MEK3 and p388-ERKl/2 signaling cascade that regulates the activity and level of API, Spl and C/EBP transcription factors to increase involucrin gene expression. Although identifying this cascade represents a major step forward in our understanding of differentiation associated gene regulation, our understanding of this signal transduction cascade is far from complete. Having established the importance of this pathway, we now propose experiments designed to extend these findings. Our studies show that novel PKC isoforms, including PKC5, are the major activators of hINV gene expression in normal keratinocytes and that differentiation agents trigger tyrosine phosphorylation of PKC8 to regulate gene expression. However, the importance of PKC8 tyrosine phosphorylation in keratinocytes is complicated, controversial, and has not been thoroughly studied. Our first major goal is to identify the role of tyrosine phosphorylation of PKC8 on PKC8 activity, subcellular localization, and ability to regulate downstream signaling events. An important observation is that signal transduction in normal human keratinocytes converges at the MAPK level on p388. Our preliminary studies suggest that differential interaction of MEK3 and MEK6 with p38S may function to regulate the balance between keratinocyte differentiation and apoptosis. The second major goal of this study is to characterize the role of these kinases in regulating the balance between differentiation and apoptosis. In particular, we will focus on the role of PKC8, MEK3 and p388 in regulating keratinocyte differentiation. The overall goal of this proposal is to expand our knowledge regarding the molecular mechanisms that drive differentiation in keratinocytes. This work is particularly important, as this mitogen-activated protein kinase (MAPK) signaling cascade has been implicated in regulating keratinocyte survival, differentiation, death, and transformation. It is hoped that a better understand of this cascade will lead to new and effective therapies for epidermal disease.