Analysis of transcription factor abnormality in the progression of preleukemic state.

我们已经克隆了由t（11 ; 19）（q23 ; p13.1）产生的MIL/MEN（也称为ELL）嵌合基因，其通常在急性髓性白血病或骨髓增生异常综合征衍生的白血病中发现。MIL/MEN融合蛋白含有MLL的氨基末端一半，包括AT钩基序，其与具有富含赖氨酸序列的MEN蛋白融合（K.Mitani等人，BLOOD 85 ; 2017，1995）。因此，MII/MEN融合蛋白可以是嵌合转录因子。用MLL（645-974 a.a.）和MEN的麦芽糖结合融合物（100-179 a.a.），分别这些抗体在转染相应cDNA的Cos 1细胞中检测到MLL/MEN融合蛋白230 kD，MLL缺失型及其下游序列（tMLL）180 kD，MEN蛋白70 kD。使用免疫染色分析和表达MLL/MEN融合蛋白、tMLL和MEN蛋白的Cos 1细胞的亚细胞分级，我们发现， ...更多信息 Conaway等报道MEN是RNA聚合酶II延伸因子（SCIENCE 271 ; 1873，1996）。另一种延伸因子elongin的功能在体外被VHL肿瘤抑制蛋白抑制，这表明异常转录延伸与肿瘤发生可能存在关系。为了证明MEN蛋白的转化活性，将MEN cDNA逆转录病毒导入Rat 1细胞。过表达Men的细胞获得了非贴壁生长的能力。此外，在这些细胞中生长因子的需求减少。然而，表达缺少富含赖氨酸区域的MEN缺失突变体的细胞没有表现出这样的生物学能力。在MEN表达细胞中，c-Fos蛋白表达和AP-1活性升高，这可能是MEN转化的部分机制。c-fos mRNA的表达，其被称为部分地调节在转录延伸的阶段，出现在MEN表达细胞早于在用表皮生长因子刺激后用空载体或缺乏富含赖氨酸的区域的缺失突变体转染的细胞中。MEN的过表达或异常表达可能在白血病向干细胞疾病转化中起重要作用。少

We have cloned an MIL/MEN(also termed as ELL)chimeric gene generated by the t(11 ; 19) (q23 ; p13.1), which is usually found in acute myeloid leukemia or myelodysplastic syndrome-derived leukemia. The MIL/MEN fusion protein contains the amino-terminal half of MLL including AT hook motifs which is fused to the MEN protein with a lysine-rich sequence(K.Mitani, et al. BLOOD 85 ; 2017,1995). Thus, the Mll/MEN fusion protein could be a chimeric transcription factor. Polyclonal antibodies to MLL and MEN were raised in rabbits against GST fusion of MLL(645-974 a.a.)and maltose-binding fusion of MEN(100-179 a.a.), respectively. These antibodies detected the MLL/MEN fusion protein of 230 kD,truncated form of the MLL Without the zinc finger domain and its downstream sequence(tMLL)of 180kD,and MEN protein of 70kD in Cos1 cells transfected with the corresponding cDNAs. Using immunostaining assay and subcellular fractionation of Cos1 cells expressing the MLL/MEN fusion, tMLL,and MEN proteins, we ha … More ve demonstrated that all these proteins exist in the nucleus of the cells.Conaway et al.have reported that the MEN is an RNA polymerase II elongation factor (SCIENCE 271 ; 1873,1996). The function of another elongation factor, elongin, is known to be inhibited by VHL tumor suppressor protein in vitro, suggesting the possible relationship of aberrant transcriptional elongation to oncogenesis. To demonstrate the transforming activity of the MEN protein, the MEN cDNA was introduced retrovirally into Ratl cells. Men-overexpressing cells acquired capacity of anchorage independent growth. In addition, the growth factor requirement was decreased in these cells. However, cells expressing a deletion mutant of MEN lacking the lysine-rich region did not exhibit such biological abilities. The c-Fos protein expression and AP-1 activity were elevated in the MEN-expressing cells, which might be part of the responsible mechanism for the transformation. The c-fos mRNA,the expression of which is known to be regulated partly at the stage of transcriptional elongation, appeared earlier in the MEN-expressing cells than in cells transfected with an empty vector or the deletion mutant lacking the lysine-rich region after the stimulation with epidermal growth factor. Overexpression or aberrant expression of MEN may play an important role in the leukemic tranfformation of stem cell disease. Less