HB-EGF AND ITS RECEPTORS

HB-EGF被合成为一种膜锚定的旁分泌生长因子，它被分解为释放成熟的HB-EGF，HB-EGF是一种强大的有丝分裂原和平滑肌细胞(SMC)的趋化因子。Hb-EGF的活性由ErbB1和ErbB4两种受体介导。HB-EGF在体内的功能在很大程度上还没有得到很好的描述。在第一个目标中，我们将在体内和体外检测SMC来源的HB-EGF在介导SMC-EC在血管内相互作用中的作用。此外，将在体内分析跨膜型和成熟型HB-EGF在转基因小鼠中作用的潜在差异。第二个目标涉及新的HB-EGF受体的特征。一种是新近纯化和克隆的140 kDa蛋白，与HB-EGF特异性结合，作为可溶性受体是HB-EGF的特异性拮抗剂。此外，两种新的ErbB4亚型，一种是膜旁区域改变，不能脱落，另一种是缺乏PI3-激酶结合部位，不能激活PI3-激酶活性，将被进一步鉴定。这些关于HB-EGF功能和受体的研究具有重要意义，因为HB-EGF被认为有助于正常的生理反应，如伤口愈合和病理过程，如动脉粥样硬化和肺动脉高压。1.研究HB-EGF在血管中的作用，包括：a)分析HB-EGF启动子-LacZ报告基因在转基因小鼠血管中的时空表达；b)分析HB-EGF对体外培养的EC-SMC相互作用的影响；c)在转基因小鼠的血管中过度表达成熟的、跨膜的和不可切割的跨膜形式；d)通过删除跨膜和胞浆结构域，获得只表达成熟HB-EGF的转基因小鼠。2.鉴定新的HB-EGF受体，包括：a)新的140 kDa HB-EGF受体的结构和功能分析；b)在膜旁结构域和PI-3K结合区不同的新的选择性剪接ErbB4亚型的特征。

HB-EGF is synthesized as a membrane-anchored juxtacrine growth factor that is shred to released mature HB-EGF, a potent mitogen and chemotactic factor for smooth muscle cells (SMC). HB-EGF activity is mediated by two receptors, ErbB1 and ErbB4. For the most part HB-EGF function in vivo is not well characterized. In the first aim the role of SMC-derived HB-EGF in mediating SMC-EC interactions in blood vessels will be examined in vivo and in vitro. In addition, potential differences in the roles of transmembrane and mature HB-EGF will be analyzed in vivo in transgenic mice. The second aim involves characterization of novel HB-EGF receptors. One is a 140 kDa protein that has been recently purified and cloned, binds HB-EGF specifically and as a soluble receptor is a specific HB-EGF antagonist. In addition, two novel ErbB4 isoforms, one with an alteration in the juxtamembrane domain that can not be shed and one that lacks the PI3-kinase binding site and can not activate PI3-kinase activity will be further characterized. These proposed studies on HB-EGF function and receptors are significant since HB-EGF has been suggested to contribute to normal physiological responses such as wound healing and pathological processes such as atherosclerosis and pulmonary hypertension. The Specific Aims of the proposal are: 1. To Investigate the Role of HB-EGF in Blood Vessels including: a) analysis of temporal and spatial HB-EGF promoter-lacZ reporter gene expression in blood vessels of transgenic mice; b) analysis of the effects of HB-EGF on EC-SMC interactions in vitro; c) over- expression of mature, transmembrane and non-cleavable transmembrane forms in the blood vessels of transgenic mice; d) generation of transgenic mice expressing mature HB-EGF only, by deleting the transmembrane and cytoplasmic domains. 2. To Characterize Novel HB-EGF Receptors including: a) Structure and functional analysis of a novel specific 140 kDa HB-EGF receptor; b) characterization of novel alternatively spliced ErbB4 isoforms differing in the juxtamembrane domain and the PI-3K binding domains.