ADENOVIRUS DEATH PROTEIN

人腺病毒2（Ad 2）编码E3 11，600 MW蛋白，命名为 腺病毒死亡蛋白（Adenovirus Death Protein，ADP）。 ADP是核（NM）糖蛋白 在感染后期大量合成。 ADP 与Ad 2 E1 B-19 K共定位，Ad 2 E1 B-19 K是Bcl-2的同源物，维持细胞 生存能力 ADP可能取代E1 B-19 K抑制细胞增殖的能力， 死亡;因此，细胞裂解并释放病毒。 结构功能 分析已经确定了ADP促进细胞死亡所需的结构域 并从ER到高尔基体和NM进行排序，其中ADP的功能是 促进细胞死亡。 我们建议确定ADP是否单独促进细胞死亡， E1 B-19 K抑制TNF或抗Fas细胞溶解的能力， 影响ICE家族的功能（导致死亡）， 通过凋亡杀死细胞。 ADP的“死亡”结构域将突变为 深入了解它们的运作方式。 蛋白质-蛋白质相互作用将通过免疫共沉淀进行检查 使用ADP的抗血清和表位标记的ADP，与GST- ADP融合蛋白，以及酵母双杂交系统。 细胞蛋白 将被克隆和表征。 我们将重点关注 ADP与E1 B-19 K结合，细胞蛋白与E1 B-19 K结合， Bcl-2家族成员 我们将确定ADP是否定位于内部或外部NM，然后 使用现有的突变体来鉴定瞬时转染中的结构域 ADP从trans-Golgi网络运输到 NM，以及ADP与E1 B-19 K的相互作用。 CD 8-ADP报告人将 用于鉴定ADP中足以靶向NM的序列。 ADP将通过共聚焦免疫荧光和平衡法监测。 膜级分的密度离心。 ADP是一种新的蛋白质，它代表了病毒学的一个新概念， 发病机制 ADP应该是理解细胞凋亡和 E1 B-19 K癌蛋白的功能，可能还有Bcl-2的成员， 家人 ADP也可能被用作杀死癌细胞的“自杀”基因。

Human adenovirus 2 (Ad2) encodes an E3 11,600 MW protein named the "Adenovirus Death Protein" (ADP). ADP is a nuclear (NM) glycoprotein synthesized in large amounts at late stages of infection. ADP colocalizes with Ad2 E1B-19K, a homolog of Bcl-2 which maintains cell viability. ADP may supersede the ability of E1B-19K to inhibit cell death; therefore, cells lyse and release virus. Structure-function analyses have identified domains required for ADP to promote cell death and be sorted from the ER to the Golgi and the NM where ADP functions to promote cell death. We propose to determine if ADP alone promotes cell death, it abrogates the ability of E1B-19K to inhibit cytolysis by TNF or anti-Fas, it affects the functions of the ICE family (which induces death), and it kills cells by apoptosis. The 'death' domains of ADP will be mutated to gain insight into how they function. Protein-protein interactions will be examined by coimmunoprecipitation using antisera to ADP and epitope tagged ADP, affinity binding to a GST- ADP fusion protein, and the yeast two-hybrid system. Cellular proteins that bind ADP will be cloned and characterized. We will focus on whether ADP binds to E1B-19K, cellular proteins that bind to E1B-19K, and to members of the Bcl-2 family. We will determine whether ADP localizes to the inner or outer NM, then use existing mutants to identify in transient transfections the domains in ADP necessary for transport of ADP from the trans-Golgi network to the NM, and for the interaction of ADP with E1B-19K. A CD8-ADP reporter will be used to identify sequences in ADP sufficient for targeting to the NM. ADP will be monitored by confocal immunofluorescence and by equilibrium density centrifugation of membrane fractions. ADP is a novel protein, and it represents a new concept in viral pathogenesis. ADP should be a powerful tool to understand apoptosis and the function of the E1B-19K oncoprotein, and perhaps members of the Bcl-2 family. ADP might also be used as a 'suicide' gene to kill cancer cells.