Paz-Priel Ido, Cai Dong Hong, Wang Dehua, Kowalski Jeanne, Blackford Amanda, Liu Huaitian, Heckman Caroline A, Gombart Adrian F, Koeffler H Phillip, Boxer Linda M, Friedman Alan D
Division of Pediatric Oncology, Johns Hopkins University, CRB 253, 1650 Orleans Street, Baltimore, Maryland 21231, USA.
Mol Cancer Res. 2005 Oct;3(10):585-96. doi: 10.1158/1541-7786.MCR-05-0111.
The CEBPA gene is mutated in 10% of acute myeloid leukemia (AML) cases. We find that CEBPA and Bcl-2 RNA levels correlate highly in low-risk human AMLs, suggesting that inhibition of apoptosis via induction of bcl-2 by CCAAT/enhancer binding protein alpha (C/EBPalpha) or its mutant variants contributes to transformation. C/EBPalphap30, lacking a NH2-terminal transactivation domain, or C/EBPalphaLZ, carrying in-frame mutations in the leucine zipper that prevent DNA binding, induced bcl-2 in hematopoietic cell lines, and C/EBPalpha induced bcl-2 in normal murine myeloid progenitors and in the splenocytes of H2K-C/EBPalpha-Emu transgenic mice. C/EBPalpha protected Ba/F3 cells from apoptosis on interleukin-3 withdrawal but not if bcl-2 was knocked down. Remarkably, C/EBPalphaLZ oncoproteins activated the bcl-2 P2 promoter despite lack of DNA binding, and C/EBPalphap30 also activated the promoter. C/EBPalpha and the C/EBPalpha oncoproteins cooperated with nuclear factor-kappaB (NF-kappaB) p50, but not p65, to induce bcl-2 transcription. Endogenous C/EBPalpha preferentially coimmunoprecipitated with p50 versus p65 in myeloid cell extracts. Mutation of residues 297 to 302 in the C/EBPalpha basic region prevented induction of endogenous bcl-2 or the bcl-2 promoter and interaction with p50 but not p65. These findings suggest that C/EBPalpha or its mutant variants tether to a subset of NF-kappaB target genes, including Bcl-2, via p50 to facilitate gene activation and offer an explanation for preferential in-frame rather than out-of-frame mutation of the leucine zipper with sparing of the basic region in C/EBPalphaLZ oncoproteins. Targeting interaction between C/EBPalpha basic region and NF-kappaB p50 may contribute to the therapy of AML and other malignancies expressing C/EBPs.
在10%的急性髓系白血病（AML）病例中，CEBPA基因发生突变。我们发现，在低风险人类AML中，CEBPA和Bcl-2的RNA水平高度相关，这表明CCAAT/增强子结合蛋白α（C/EBPα）或其突变变体通过诱导bcl-2抑制凋亡有助于细胞转化。缺乏NH2末端反式激活结构域的C/EBPαp30或亮氨酸拉链中存在框内突变从而阻止DNA结合的C/EBPαLZ，在造血细胞系中诱导bcl-2表达，并且C/EBPα在正常小鼠髓系祖细胞和H2K-C/EBPα-Emu转基因小鼠的脾细胞中诱导bcl-2表达。在白细胞介素-3撤除时，C/EBPα保护Ba/F3细胞免于凋亡，但如果bcl-2被敲低则无此作用。值得注意的是，尽管缺乏DNA结合，C/EBPαLZ癌蛋白仍激活bcl-2 P2启动子，C/EBPαp30也激活该启动子。C/EBPα和C/EBPα癌蛋白与核因子-κB（NF-κB）p50而非p65协同作用以诱导bcl-2转录。在髓系细胞提取物中，内源性C/EBPα与p50的共免疫沉淀优先于与p65的共免疫沉淀。C/EBPα碱性区域中297至302位残基的突变阻止内源性bcl-2或bcl-2启动子的诱导以及与p50的相互作用，但不影响与p65的相互作用。这些发现表明，C/EBPα或其突变变体通过p50与包括Bcl-2在内的NF-κB靶基因子集相连，以促进基因激活，并为C/EBPαLZ癌蛋白中亮氨酸拉链优先发生框内而非框外突变且碱性区域未受影响提供了解释。靶向C/EBPα碱性区域与NF-κB p50之间的相互作用可能有助于AML和其他表达C/EBPs的恶性肿瘤的治疗。
