THE BIOCHEMISTRY AND FUNCTION OF HRK

程序性细胞死亡(PCD)通常通过细胞凋亡发生，这是一种生理性的 胚胎发育过程中常见的细胞死亡形式，组织周转 以及细胞群体的选择。最近有几个基因被发现 确定激活、执行或抑制细胞死亡途径。 BCL-2家族的凋亡调控基因调控细胞死亡 由一系列广泛的信号引起。该家族的两名成员，BCL-2 和bcl-x在许多组织中表达，它们的重要性是 突显这一发现的是缺乏bcl2或bclx基因的突变小鼠 由于异常的细胞凋亡而表现出致命性表型。此外， 失控的Bcl2和Bclx蛋白的表达起着重要的作用 在癌症发展和肿瘤细胞对治疗的耐药性中的作用- 诱导细胞凋亡。然而，Bcl2家族的生化机制 成员对细胞凋亡的调控知之甚少。我们已经确定了一个 调控Bcl2和BclX的新基因harakiri/L 以及哺乳动物细胞的凋亡性死亡。哈拉基里产品(HRK) 与Bcl2或BclX/L无显著同源性，缺乏 保守的BH1和BH2结构域，与BCL-2家族成员共享。 HRK含有一个8个氨基酸的BH3基序，它存在于 BCL-2家族。值得注意的是，HRK在物理上与反 细胞凋亡蛋白Bcl2和BclX/L与促死亡蛋白Bcl2无关 BAX和BAK等成员。HRK基因的表达诱导急性心肌梗死的发生 哺乳动物细胞的细胞死亡。重要的是促进死亡的活动 HRK的表达受Bcl2和BclX/L的抑制，提示HRK是一种常见的 抗细胞凋亡蛋白Bcl2和Bclxl的靶点。在这 建议我们提出实验(I)，以进一步表征 HRK与Bcl2/Bclk-X/L的分子相互作用及鉴定 HRK的其他靶点；(Ii)检查表达、调控、 亚细胞定位；(Iii)研制重组腺病毒 研究HRK的细胞靶点及其激活细胞死亡的能力 正常细胞和癌细胞，以及(Iv)产生缺陷的突变小鼠 以确定其在体内的生理作用。概述的研究 应该为研究细胞凋亡的途径提供新的视角 这可能导致替代治疗策略的治疗 癌症。

Programmed cell death (PCD) often occurs via apoptosis, a physiological form of cellular demise common during embryogenesis, turnover of tissues and selection of cell populations. Several genes have been recently identified that activate, execute or inhibit the cell death pathway. The bcl-2 family of apoptosis-regulatory genes modulates cell death induced by a wide array of signals. Two members of the family, bcl-2 and bcl-x are expressed in many tissues and their importance is underscored by the finding that mutant mice deficient in bcl-2 or bcl-x exhibit lethal phenotypes due to abnormal apoptosis. Moreover, deregulated expression of Bcl-2 and Bcl-x proteins play an important role in cancer development and resistance of tumor cells to therapy- induced apoptosis. Yet, the biochemical mechanism by which Bcl-2 family members regulate apoptosis is poorly understood. We have identified a novel gene, harakiri, that functions as a regulator of Bcl-2 and Bcl-X/L and apoptotic cell death in mammalian cells. The harakiri product (HRK) does not exhibit significant homology to Bcl-2 or Bcl-X/L and lacks conserved BH1 and BH2 domains which are shared by BCL-2 family members. HRK contains an 8 amino acid BH3 motif which is found in proteins of the BCL-2 family. Significantly, HRK physically interacts with anti- apoptosis proteins Bcl-2 and Bcl-X/L but not with death-promoting Bcl-2 members such as Bax and Bak. Expression of HRK induces rapid onset of cell death in mammalian cells. Importantly the death-promoting activity of HRK is repressed by Bcl-2 and Bcl-X/L suggesting that HRK is a common target for the anti-apoptosis proteins Bcl-2 and Bcl-XL. In this proposal we propose experiments (i) to further characterize the molecular interaction between HRK and Bcl-2/Bclk-X/L and identify additional targets of HRK; (ii) to examine the expression, regulation, subcellular localization; (iii) develop recombinant adenoviruses to study cellular targets of HRK and its ability to activate cell death in normal and cancer cells, and (iv) to generate mutant mice deficient in HRK to determine its physiological role in vivo. The studies outlined in this proposal should provide novel sight into the apoptosis pathway which could lead to alternative therapeutic strategies for treatment of cancer.