AMPHIREGULIN PROCESSING IN HUMAN KERATINOCYTES

双调蛋白（Amphiregulin，AR）是表皮生长因子受体的一员 （EGFR）配体家族。在培养的人角质形成细胞中，双调蛋白 是主要的自分泌生长因子。此外，促有丝分裂 AR对角质形成细胞的作用可以通过与 其他蛋白质如CD 9，并被肝素抑制。虽然 AR在正常成人表皮中的作用还不完全清楚， AR蛋白和mRNA水平的分析提供了一些见解， 它的功能。AR在正常成人角质形成细胞中弱表达 但在几种过度增殖性疾病中上调 例如牛皮癣和肿瘤。两者之间可能存在因果关系 最近的研究表明，AR水平升高与银屑病有关， 发现在Kl 4下过表达人AR cDNA的转基因小鼠 角蛋白启动子显示银屑病表型。这一结果表明， 与TGF α过度表达的作用形成直接对比，表明 这两种配体的不同作用。此外，AR，而不是TGF α， 在几种创伤模型中显著诱导（10-30倍）。 双调蛋白是一种糖基化的膜锚定蛋白， 前体（proAR）。在极化的上皮细胞中，我们检查了 proAR的生物合成和加工，以及证明的复合物 proAR胞外域的顺序加工以产生多个细胞 和可溶性AR形式。一种主要的43 kD可溶性AR形式是新的， 含有N-末端前区并具有C-末端延伸。的 多种AR种类的生物学意义尚不清楚。的目标 这项拨款提案的目的是确定细胞和可溶性形式 正常人角质形成细胞中产生的AR，并开始解决其 这种细胞的生物学功能。这些研究将是第一个 为了全面研究proAR的生物合成和加工， 正常原代上皮细胞类型。所涉机制 调节角质形成细胞中的proAR胞外域切割也将提供 理解金属蛋白酶在这一过程中的作用的基础 过程中，更一般地在膜蛋白脱落的过程中。 最后，这些研究应该加强我们对 在不同环境条件下表达的各种AR形式 (eg紫外线暴露）和不同的皮肤病状态。

Amphiregulin (AR) is a member of the epidermal growth factor receptor (EGFR) family of ligands. In cultured human keratinocytes, amphiregulin is the major autocrine growth factor. Additionally, the mitogenic effects of AR on keratinocytes can be enhanced by interaction with other proteins such as CD9 and are inhibited by heparin. Although the role of AR in normal adult epidermis is not completely understood, analysis of AR protein and mRNA levels has provided some insights into its function. AR is weakly expressed in keratinocytes in normal adult epidermis but is up-regulated in several hyperproliferative disorders such as psoriasis and in tumors. A possible causal link between elevated AR levels and psoriasis has been suggested by the recent finding that transgenic mice overexpressing human AR cDNA under the Kl4 keratin promoter display a psoriatic phenotype. This result stands in direct contrast to the effects of TGFalpha overexpression, suggesting distinct roles for these two ligands. Moreover, AR, but not TGFalpha, is dramatically induced (10-30 fold) in several wounding models. Amphiregulin is synthesized as a glycosylated membrane-anchored precursor (proAR). In polarized epithelial cells, we have examined the biosynthesis and processing of proAR and demonstrated complex sequential processing of proAR ectodomain to produce multiple cellular and soluble AR forms. A predominant 43 kD soluble AR form is novel, contains the N-terminal proregion and has a C-terminal extension. The biological significance of multiple AR species is not known. The goals of this grant proposal are to identify the cellular and soluble forms of AR produced in normal human keratinocytes and begin to address their biological function in this cell type. These studies will be the first to comprehensively examine proAR biosynthesis and processing in a normal primary epithelial cell type. The mechanism(s) involved in regulating proAR ectodomain cleavage in keratinocytes will also provide the basis for understanding the role of metalloproteases in this process and more generally in the process of membrane protein shedding. Finally, these studies should enhance our understanding of the role of the various AR forms expressed under different environmental conditions (eg, UV exposure) and in different skin disease states.