CHIMERIC E2A-HLF TRANSCRIPTION FACTOR IN ACUTE LEUKEMIA

在正常发育过程中，多达90%的淋巴细胞 前体细胞经历非生产性抗原受体基因重排 是通过一种有序的、基因上的 由三个不同阶段组成的控制程序：细胞命运 决定，死亡信号的执行和caspase的诱导- 介导的蛋白质分解。发现E2a-HLF嵌合转录因子 伴有染色体易位的白血病前B淋巴细胞 T(17；19)(q22；p13)，被认为激活了slug的表达，a 线虫Ces-1生存基因的哺乳动物同源基因，其 转录抑制因子活性增强可阻断蛋白的表达 下游细胞死亡效应器。因此，淋巴细胞前体 通常会被指定为销毁(例如，由于有缺陷的抗原 受体基因重排，缺乏外源生存信号 或与P53激活引起的DNA损伤)继续存活，一些 获得促进白血病表型的额外突变的细胞。 这项提议的目标1试图确定细胞命运基因 在E2A-HLF结合位点下游，驱动白血病发生和 在正常的淋巴生成过程中调节细胞的存活决定。这个 实验将利用可诱导的基因表达系统， 代表性差异分析(RDA)和其他新兴技术 评估在存在和不存在的情况下转录的mRNA的技术 由嵌合蛋白结合的DNA。转基因小鼠将成为 研究确定哺乳动物CES-1-1的正常发育作用 像基因(如蜗牛/鼻涕虫家族)和Nfil3/E4bp4，一种细胞因子- 线虫Ces-2基因的哺乳动物同源基因。分开的形式 经典的基因反式激活，由保守的bZIP结构域介导， 缺失bZIP DNA结合域的E2A-HLF突变体对细胞的保护作用 通过AD1和AD2反式激活因子的活性而引起的细胞凋亡 E2A分子氨基末端的结构域。因此，目标2 导致表达Groucho样蛋白的测试 细胞凋亡所需的第二个下游遗传程序。当前 对哺乳动物细胞凋亡途径的洞察 与CES/CED相比，淋巴系造血术受益匪浅。 线虫(线虫)的途径。拟议的研究将延长 通过揭示许多仍然难以捉摸的中间步骤 在细胞凋亡的启动事件和细胞的激活 Caspase介导的蛋白水解级联反应。对E2A-HLF的理解 颠覆白血病淋巴母细胞的细胞死亡程序最终可能 为白血病细胞的耐药性提供了新的基础，并可能 揭示暴露后改善临床结果的有吸引力的目标 到细胞毒剂。

During normal development, as many as 90 percent of lymphocyte precursors undergo nonproductive antigen receptor gene rearrangements are selected for apoptotic death through an orderly, genetically controlled program consisting of three distinct stages: cell fate decisions, execution of death signals and the induction of caspase- mediated proteolysis. The E2A-HLF chimeric transcription factor, found in leukemic pro-B lymphocytes with the chromosomal translocation t(17;19)(q22;p13), is postulated to activate expression of SLUG, a mammalian ortholog of the ces-1 survival gene of C. elegans, whose increased transcriptional repressor activity blocks the expression of downstream cell death effectors. Thus, lymphocyte precursors that normally would be slated for destruction (e.g, due to defective antigen receptor gene rearrangements, the absence of exogenous survival signals or to DNA damage with p53 activation) continue to survive, with some cells acquiring additional mutations that promote a leukemic phenotype. Aim 1 of this proposal seeks to identify the cell fate genes downstream from the E2A-HLF binding site that drive leukemogenesis and mediate cell survival decisions during normal lymphopoiesis. The experiments will exploit inducible gene expression systems, representational difference analysis (RDA) and other emerging technologies to assess mRNAs transcribed in the presence and absence of DNA binding by the chimeric protein. Genetically altered mice will be examined to define the normal developmental roles of mammalian ces-1- like genes (e.g., the Snail/Slug family) and Nfil3/E4bp4, a cytokine- regulated mammalian ortholog of the C. elegans ces-2 gene. Apart form classical gene transactivation, mediated by a conserved bZIP domain, E2A-HLF mutants with disabled bZIP DNA-binding domains can protect cells from apoptosis through the activity of the AD1 and AD2 transactivator domains within the amino-terminus of the E2A molecule. Thus, Aim 2 tests leading to expression of a Groucho-like protein that blocks a second downstream genetic program required for apoptosis. Current insights into the apoptotic pathways operating in mammalian lymphopoiesis have profited enormously from comparisons with the ces/ced pathway in nematodes (C. elegans). The proposed research will extend this knowledge by revealing many of the still-elusive intermediate steps between the initiating events of apoptosis and activation of the caspase-mediated proteolytic cascade. An understanding of how E2A-HLF subverts the cell death program in leukemic lymphoblasts may ultimately suggest a new basis for drug resistance in leukemic cells and could reveal attractive targets for improving clinical outcomes after exposure to cytotoxic agents.