Transcriptional regulation of apoptosis

描述(由申请者提供)：对细胞凋亡的抵抗导致肿瘤的发生和不良的治疗结果。肿瘤抑制基因P53介导了两种主要的细胞反应，细胞周期停滞和细胞凋亡，但决定细胞命运的分子基础仍然不清楚。有人认为，p53非依赖性效应分子和p53依赖性基因表达之间的相互作用可能是细胞结局的关键决定因素。这些研究通过检验Sp/KLF家族成员和P53在调控基因表达上的相互作用来验证这一假说，重点关注p21CIP1作为凋亡反应的衰减器的作用，并阐明凋亡途径关键成员基础表达水平的变化是否有助于细胞命运的决定。位于启动子和第一内含子的多个p53反应元件参与了DNA损伤诱导的p21CIP1的上调，其中p21CIP1启动子的这两个位置以不同的方式受到调控。在第一个目标中，我们将阐明这两种类型P53位点差异的分子基础，并研究P53、Sp1和其他Sp/KLF家族成员之间的相互作用。其他细胞因子，如Sp/KLF家族成员，在p53依赖的基因调控中的关键作用将被视为影响细胞结果的一种手段。已发表的研究和初步数据表明，细胞周期蛋白依赖性激酶抑制剂p21CIP1具有干扰细胞凋亡和影响细胞命运的能力。在第二个目标中，将验证和表征p21CIP1在减弱细胞死亡反应中的作用，并阐明p21CIP1这一有趣效应的潜在分子基础。调节细胞凋亡途径关键成分的基础表达水平是决定细胞命运结果的另一种机制。Bax基因中与Bax基因转录调控有关的元件已被鉴定。NHE1是一个新的P53靶基因，已被证明通过脱酰胺作用调节Bclxl的抗细胞凋亡作用。在第三个目标中，将探索Bax、NHE1和Bclxl之间的分子相互作用，以确定这些机制是否可以解释特定肿瘤细胞的抗凋亡表型。根据特定的细胞条件，肿瘤抑制蛋白P53诱导生长停滞或介导凋亡反应。对许多化疗药物引起的DNA损伤的最佳治疗反应是细胞死亡，而不是抑制细胞周期进展。阐明调控P53诱导细胞凋亡的分子机制可能会导致更有效的治疗干预和克服在许多肿瘤中发现的化疗耐药表型。

DESCRIPTION (provided by applicant): Resistance to apoptosis contributes to tumorigenesis as well as poor therapeutic outcome. The tumor suppressor p53 mediates two predominant cellular responses, cell cycle arrest and apoptosis, but the molecular basis for this cell fate determination has remained elusive. It is proposed that an interplay between p53-independent effectors and p53-dependent gene expression may be the crucial determinant for cellular outcome. The proposed studies test this hypothesis by examining the interplay between Sp/KLF family members and p53 on regulating gene expression, focusing on the role of p21CIP1 as an attenuator of the apoptotic response, and elucidating whether alterations in basal levels of expression of key members of apoptotic pathways contribute to cell fate decisions. Multiple p53 response elements located in both the promoter and first intron are involved in the DNA damage-induced upregulation of p21CIP1 with the two sites in the p21CIP1 promoter being regulated in distinct manners. In the first aim, the molecular basis for the differences between the two types of p53 sites will be elucidated and the interplay between p53, Sp1, and other Sp/KLF family members will be examined. The key role of other cellular factors such as Sp/KLF family members in p53-dependent gene regulation will be addressed as a means for influencing cellular outcomes. The ability of the cyclin-dependent kinase inhibitor p21CIP1 to interfere with apoptosis and influence cell fate has been suggested by published studies as well as preliminary data. In the second aim, the role of p21CIP1 in attenuating the cell death response will be validated and characterized and the underlying molecular basis for this intriguing effect of p21CIP1 will be elucidated. Regulating the basal levels of expression of key components of apoptotic pathways is an alternative mechanism for determining cell fate outcomes. Elements in the bax gene that confer constitutive transcriptional regulation of Bax have been identified. NHE1 is a novel p53 target gene that has been shown to regulate the anti-apoptotic effects of Bcl-XL via deamidation. In the third aim, the molecular interplay between Bax, NHE1, and Bcl-XL will be explored to determine whether such mechanisms can explain the apoptotic resistant phenotype of particular tumor cells. Depending upon particular cellular conditions, the tumor suppressor protein p53 induces growth arrest or mediates an apoptotic response. The optimal therapeutic response to DNA damage caused by many chemotherapeutic agents is cell death rather than inhibition of cell cycle progression. Elucidating the molecular mechanisms that are responsible for regulating the ability of p53 to trigger apoptosis versus arrest may lead to more effective therapeutic intervention and a way to overcome the chemotherapeutic-resistant phenotype found in many tumors.