PACAP: A NOVEL PROAPOPTOTIC CASPASE ADAPTER PROTEIN

描述（由申请人提供）：细胞凋亡或程序性细胞死亡是免疫系统功能发展和维持的关键过程。细胞凋亡依赖于半胱氨酸蛋白酶家族（半胱天冬酶），其激活受正、负两种调节机制的影响。最近，我们发现并部分描述了一种新的促凋亡caspase衔接蛋白（PACAP），它与两种引发caspase-2和-9相互作用。迄今为止，我们观察到：当PACAP在细胞系中过表达时，细胞凋亡被诱导，NF-kappaB通路被激活。在原代人B细胞中，内源性表达的PACAP的转录在B细胞激活后迅速而显著地下调，在细胞凋亡期间上调，这一发现支持了PACAP作为一种调节蛋白的作用。此外，PACAP与Bcl10形成复合物，Bcl10是一种与某些细胞类型和多种人类恶性肿瘤中NF-kappaB激活相关的蛋白质。因此，我们的假设是，PACAP可能将细胞外信号引导到casp-2或-9凋亡通路，并可能进一步调节凋亡通路中激活的其他细胞内通路。为了验证这一假设，我们提出了三个特定目标，利用分子和细胞生物学、免疫学和生化方法，在生理相关细胞模型的背景下进一步定义PACAP的特性，其中casp-2和caspase-9在细胞死亡途径中发挥重要作用。建议的具体目标是：1。提供PACAP相互作用的详细生化分析；2. 提供PACAP凋亡信号通路组成成分的综合定义；和3。进一步鉴定PACAP介导的细胞内信号传导的其他潜在调节因子，重点研究NF-kappaB通路。

DESCRIPTION (provided by applicant): Apoptosis or programmed cell death is a crucial process for the development and maintenance of a functional immune system. Apoptosis depends upon a family of cysteine proteases (caspases) whose activation is subject to both negative and positive regulatory mechanisms. Recently we discovered and have partially characterized a novel pro-apoptotic caspase adaptor protein (PACAP) that interacts with two initiator caspases, caspase-2 and -9. To date we have observed the following: when PACAP is overexpressed in cell lines, apoptosis is induced and the NF-kappaB pathway is activated. Support for a role for PACAP as a regulatory protein derives from the observations that the transcription of endogenously expressed PACAP in primary human B cells is rapidly and dramatically down regulated after B-cell activation and upregulated during apoptosis. Moreover PACAP forms complexes with Bcl10, a protein linked to NF-kappaB activation in some cell types and to a variety of human malignancies. Thus it is our hypothesis that PACAP may serve to direct extracellular signals to either casp-2 or -9 apoptotic pathways and may further regulate other intracellular pathways activated during the apoptotic pathway. To test this hypothesis we propose three Specific Aims that utilize molecular and cellular biological, immunological and biochemical approaches to further define the properties of PACAP in the context of physiologically relevant cellular models where casp-2 and caspase-9 play essential roles in cell death pathways. The proposed Specific Aims are: 1. To provide a detailed biochemical analysis of PACAP interactions; 2. To provide a comprehensive definition of the components that comprise the PACAP apoptotic signaling pathway; and 3. To provide further identification of other potential regulators of PACAP mediated intracellular signaling with an emphasis on the NF-kappaB pathway. The proposed studies are designed to provide fundamental information about this newly discovered protein that serves as a regulator of apoptotic pathways linked to both caspases-2 and-9. The long term goals of the proposed studies are to enhance our abilities to understand basic processes leading to human diseases linked to a dysfunctional immune system.