PROMOTION OF CELL SURVIVAL BY ADENOVIRUS E1B 19K PROTEIN

受调控的细胞死亡是消除特定基因的重要过程 多细胞生物中的细胞在正常发育和 动态平衡。失控的细胞死亡可导致退行性疾病 例如阿尔茨海默氏症、帕金森氏症和艾滋病。对细胞的抑制 死亡也可能导致肿瘤形成和对化疗的抗药性。 治疗。人腺病毒E1B19kD蛋白是一种 一类与细胞抑制有关的蛋白质的重要成员 死亡。它促进了感染了腺病毒或 暴露在几种外部细胞死亡(凋亡)诱导刺激下。这个 19kD蛋白在功能上与Bc1-2原癌蛋白相似， Epstein-Barr病毒BHRF1蛋白。出现了19kD和Bcl2蛋白 共享抑制细胞死亡所必需的共同序列基序。 我们已经鉴定并克隆了四种与之相互作用的细胞蛋白 19 kD(19-kD相互作用蛋白，Nip1至4)和Bc1-2蛋白 通过这些同源序列基序。为了阐明共同之处 19kD和Bc1-2蛋白抑制细胞死亡的机制 建议从功能上描述不同的国家行动方案。这些因素的作用 不同细胞因子诱导的细胞死亡中的细胞蛋白 机制将使用多个细胞死亡/存活分析来检验。 这些蛋白的内源性表达和亚细胞定位 本课程将研究对各种细胞死亡刺激的反应。 其中一些细胞蛋白含有耐人寻味的序列同源性。 Nip1与某些特定的催化结构域有有限的同源性 哺乳动物的磷酸二酯酶。Nip2与(GTPase-)有同源性 刺激蛋白RhoGAP。Nip3定位于线粒体。Nip2 和Nip4含有可能的钙结合基序。这些蛋白质将是 特征是阐明它们在信号转导和钙离子中的作用 动员。由于腺病毒诱导的细胞死亡效应分子 感染情况尚不清楚，我们建议使用无细胞体外细胞死亡 识别和克隆这种效应器的系统(S)。自停用 19kD和Bc1-2蛋白的促存活活性 有利的是，我们建议探索使用突变体的可能性 作为显性负抑制因子的生存区域存在缺陷。 类似地，与生存结构域相对应的多肽也将 被检测为19kD和Bc1-2蛋白的竞争性抑制物。因此， 我们的建议可能解释19kD和Bc1-2 蛋白质抑制细胞死亡，也揭示了干扰策略 他们的活动。

Regulated cell death is an important process for elimination of specific cells in multicellular organisms during normal development and homeostasis. Deregulated cell death can lead to degenerative diseases such as Alzheimer's and Parkinson's disease and AIDS. Inhibition of cell death can also lead to oncogenesis and resistance to chemotherapeutic treatments. The E1B 19 kD protein of human adenovirus (Ad) is an important member of a class of proteins involved in suppression of cell death. It promotes the survival of cells infected with adenovirus or exposed to several external cell death (apoptosis)-inducing stimuli. The 19 kD protein is functionally similar to the Bc1-2 protooncoprotein and the Epstein-Barr virus BHRF1 protein. The 19 kD and Bcl-2 proteins appear to share common sequence motifs essential for inhibition of cell death. We have identified and cloned four cellular proteins that interact with the 19 kD (19-kD-interacting proteins, Nip1 to 4) and the Bc1-2 proteins through these homologous sequence motifs. To elucidate the common mechanism of cell death inhibition by the 19 kD and Bc1-2 proteins, we propose to functionally characterize the various Nips. The role of these cellular proteins in regulating cell death induced by different mechanisms will be examined using multiple cell death/survival assays. Endogenous expression and subcellular localization of these proteins in response to exposure to various cell death stimuli will be examined. Some of these cellular proteins contain intriguing sequence homologies. Nip1 shares limited homology with the catalytic domain of certain mammalian phosphodiesterases. Nip2 shares homology with the (GTPase- stimulating protein RhoGAP. Nip3 is localized in the mitochondria. Nip2 and Nip4 contain putative Ca2+ -binding motifs. These proteins will be characterized to elucidate their role in signal transduction and calcium mobilization. Since the cell death effectors induced by adenovirus infection is not known, we propose to use a cell free in vitro cell death system to identify and clone such effector(s). Since inactivation of survival-promoting activities of the 19 kD and Bc1-2 proteins would be advantageous, we propose to explore the possibility of using mutants defective in the survival domains as dominant negative inhibitors. Similarly, peptides corresponding to the survival domains will also be examined as competitive inhibitors of the 19 kD and Bc1-2 proteins. Thus, our proposal may illuminate the mechanism by which the 19 kD and Bc1-2 proteins suppress cell death and also reveal strategies to interfere with their activities.