Inukai T, Inaba T, Ikushima S, Look A T
Department of Experimental Oncology, St. Jude Children's Research Hospital, Memphis, Tennessee 38105, USA.
Mol Cell Biol. 1998 Oct;18(10):6035-43. doi: 10.1128/MCB.18.10.6035.
The chimeric oncoprotein E2A-HLF, generated by the t(17;19) chromosomal translocation in pro-B-cell acute lymphoblastic leukemia, incorporates the transactivation domains of E2A and the basic leucine zipper (bZIP) DNA-binding and protein dimerization domain of HLF (hepatic leukemic factor). The ability of E2A-HLF to prolong the survival of interleukin-3 (IL-3)-dependent murine pro-B cells after IL-3 withdrawal suggests that it disrupts signaling pathways normally responsible for cell suicide, allowing the cells to accumulate as transformed lymphoblasts. To determine the structural motifs that contribute to this antiapoptotic effect, we constructed a panel of E2A-HLF mutants and programmed their expression in IL-3-dependent murine pro-B cells (FL5.12 line), using a zinc-inducible vector. Neither the E12 nor the E47 product of the E2A gene nor the wild-type HLF protein was able to protect the cells from apoptosis induced by IL-3 deprivation. Surprisingly, different combinations of disabling mutations within the HLF bZIP domain had little effect on the antiapoptotic property of the chimeric protein, so long as the amino-terminal portion of E2A remained intact. In the context of a bZIP domain defective in DNA binding, mutants retaining either of the two transactivation domains of E2A were able to extend cell survival after growth factor deprivation. Thus, the block of apoptosis imposed by E2A-HLF in pro-B lymphocytes depends critically on the transactivating regions of E2A. Since neither DNA binding nor protein dimerization through the bZIP domain of HLF is required for this effect, we propose mechanisms whereby protein-protein interactions with the amino-terminal region of E2A allow the chimera to act as a transcriptional cofactor to alter the expression of genes regulating the apoptotic machinery in pro-B cells.
嵌合癌蛋白E2A-HLF由前B细胞急性淋巴细胞白血病中的t(17;19)染色体易位产生，它融合了E2A的反式激活结构域以及HLF（肝白血病因子）的碱性亮氨酸拉链（bZIP）DNA结合和蛋白质二聚化结构域。E2A-HLF能够在白细胞介素-3（IL-3）撤除后延长依赖IL-3的小鼠前B细胞的存活期，这表明它破坏了通常负责细胞自杀的信号通路，使细胞能够以转化的淋巴母细胞形式积累。为了确定促成这种抗凋亡作用的结构基序，我们构建了一组E2A-HLF突变体，并使用锌诱导载体在依赖IL-3的小鼠前B细胞（FL5.12系）中编程表达这些突变体。E2A基因的E12或E47产物以及野生型HLF蛋白均无法保护细胞免受IL-3剥夺诱导的凋亡。令人惊讶的是，只要E2A的氨基末端部分保持完整，HLF bZIP结构域内不同的失活突变组合对嵌合蛋白的抗凋亡特性影响很小。在DNA结合缺陷的bZIP结构域背景下，保留E2A两个反式激活结构域之一的突变体能够在生长因子剥夺后延长细胞存活期。因此，E2A-HLF在前B淋巴细胞中施加的凋亡阻断关键取决于E2A的反式激活区域。由于这种作用既不需要通过HLF的bZIP结构域进行DNA结合也不需要蛋白质二聚化，我们提出了一些机制，即与E2A氨基末端区域的蛋白质-蛋白质相互作用使嵌合体能够作为转录辅因子来改变调节前B细胞凋亡机制的基因的表达。
