REGULATION OF BCL XL BY CASPASES

程序性细胞死亡的失调是许多人类疾病的核心。细胞死亡不足会导致癌症和一些自身免疫性疾病，而过度的细胞死亡会导致许多神经系统疾病和艾滋病。细胞凋亡抑制剂Bcl-2的过度表达源于滤泡性淋巴瘤患者的染色体易位特征。Bcl-2家族成员Bcl-XL在艾滋病患者的卡波西氏肉瘤病变中经常被发现过表达。在肿瘤发展的晚期，一个或多个Bcl-2家族成员的上调是一种常见的现象。然而，Bcl-2家族成员调控程序性细胞死亡的分子机制才刚刚开始被理解。同样，由于未知的机制，Bcl-XL和Bcl-2阻止半胱天冬酶的激活，半胱氨酸蛋白酶家族是凋亡细胞死亡的关键促进因素。最近，Bcl-2家族成员被发现作为caspase底物。caspase在n端附近的环域中切割Bcl-XL和Bcl-2，导致BH4同源结构域的丢失，这是抑制细胞死亡所必需的结构域。因此，半胱天冬酶对Bcl-XL和Bcl-2的切割将这些蛋白从细胞死亡的有效抑制剂转化为细胞死亡的有效诱导剂。然而，除了Bcl-XL切割位点P1位置普遍保守的天冬氨酸外，这些位点与任何其他已知的caspase切割位点都不相似。实验旨在确定细胞激酶/磷酸酶对Bcl-XL切割位点的翻译后修饰是否会调节半胱天蛋白酶对这些位点的识别。我们将在神经元和肿瘤细胞系中研究磷酸化和相关激酶在细胞凋亡过程中调控BcI-XL功能的作用。被切割的Bcl-XL的促死亡活性所需的蛋白质结构域将由广泛的突变决定。将进行生化和功能分析，以探索这种促凋亡活性背后的分子机制。这些研究有望显著推进我们对程序性细胞死亡的分子过程的理解，这些过程影响着广泛的人类疾病。

Dysregulation of programmed cell death is central to a number of human diseases. Insufficient cell death leads to cancer and some autoimmune diseases while excessive cell death contributes to a number of neurological disorders and AIDS. Overexpression of Bcl-2, an inhibitor of apoptosis, results from the chromosomal translocations characteristic of follicular lymphoma patients. Overexpression of a Bcl-2 family member, Bcl-XL, is frequently found in the Kaposi's sarcoma lesions of AIDS patients. Upregulation of one or more Bcl-2 family members in the late stage of tumor development is a common occurrence. However, the molecular mechanisms by which Bcl-2 family members regulate programmed cell death are only beginning to be understood. Again by unknown mechanisms Bcl-XL and Bcl-2 prevent the activation of caspases, a family of cysteine proteases that are key facilitators of apoptotic cell death. Recently, Bcl-2 family members were found to serve as caspase substrates. Caspases cleave Bcl-XL and Bcl-2 in the loop domain near the N-terminus which results in loss of the BH4 homology domain, a domain that is required for inhibition of cell death. As a result, cleavage of Bcl-XL and Bcl-2 by caspases converts these proteins from potent inhibitors of cell death to potent inducers of cell death. However, except for the universally conserved aspartate at the P1 position of the Bcl-XL cleavage sites, these sites do not resemble any other known caspase cleavage sites. Experiments are proposed to determine if posttranslational modification of the Bcl-XL cleavage sites by cellular kinases/phosphatases modulates the recognition of these site by caspases. The role of phosphorylation and the responsible kinases that regulate BcI-XL function during apoptosis will be studied in neurons and tumor cell lines. The protein domains required for the pro-death activity of cleaved Bcl-XL will be determined by extensive mutagenesis. Biochemical and functional analyses will be performed to explore the molecular mechanism behind this pro-apoptotic activity. These studies are expected to significantly advance our understanding of the molecular processes of programmed cell death which impact on a wide range of human disorders.