CHIMERIC E2A-HLF TRANSCRIPTION FACTOR IN ACUTE LEUKEMIA

在正常发育过程中，多达90%的淋巴细胞 前体经历非生产性抗原受体基因重排 通过一种有序的，遗传的 由三个不同阶段组成的受控程序：细胞命运 决定，执行死亡信号和诱导半胱天冬酶- 介导的蛋白水解。 E2A-HLF嵌合转录因子，发现 白血病前B淋巴细胞染色体易位 t（17; 19）（q22; p13）被认为激活SLUG的表达， ces-1存活基因的哺乳动物直系同源物。elegans，其 增加的转录抑制因子活性阻断了 下游细胞死亡效应物。 因此，淋巴细胞前体， 通常将被指定用于破坏（例如，由于缺陷抗原 受体基因重排，缺乏外源生存信号 或DNA损伤与p53激活）继续生存，一些 细胞获得额外的突变，促进白血病表型。 该提案的目标1旨在确定细胞命运基因 在E2A-HLF结合位点下游驱动白血病发生， 在正常淋巴细胞生成过程中介导细胞存活决定。 的 实验将利用诱导型基因表达系统， 代表性差异分析（RDA）和其他新兴的 技术来评估mRNA转录的存在和不存在的 嵌合蛋白的DNA结合。 转基因小鼠将在 检查以确定哺乳动物CES-1的正常发育作用， 类似基因（例如，Snail/Slug家族）和Nfil3/E4bp4，一种细胞因子- 调节的哺乳动物C.线虫ces-2基因。 分离形式 经典的基因反式激活，由保守的bZIP结构域介导， 具有禁用的bZIP DNA结合结构域的E2A-HLF突变体可以保护细胞 通过AD 1和AD 2反式激活因子的活性从凋亡中 在E2A分子的氨基末端内的结构域。 因此，目标2 导致Groucho样蛋白表达的试验， 凋亡所需的第二个下游遗传程序。 电流 对哺乳动物细胞凋亡途径的深入了解 与ces/ced相比， 途径（C. elegans）。 该研究计划将扩大 通过揭示许多仍然难以理解的中间步骤， 在细胞凋亡的起始事件和细胞凋亡的激活之间， 半胱天冬酶介导的蛋白水解级联反应。 了解E2A-HLF如何 颠覆了白血病淋巴母细胞的细胞死亡程序， 提出了白血病细胞耐药性的新基础， 揭示了改善暴露后临床结局的有吸引力的靶点 到细胞毒性剂。

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.