Zeleznik-Le N J, Harden A M, Rowley J D
Department of Medicine, University of Chicago, IL 60637.
Proc Natl Acad Sci U S A. 1994 Oct 25;91(22):10610-4. doi: 10.1073/pnas.91.22.10610.
Translocations involving chromosome band 11q23, found in acute lymphoid and myeloid leukemias, disrupt the MLL gene. This gene encodes a putative transcription factor with homology to the zinc fingers and other domains of the Drosophila trithorax gene product and to the "AT-hook" motif of high mobility group proteins. To map potential transcriptional activation or repression domains of the MLL protein, yeast GAL4 DNA-binding domain and MLL hybrid protein-expressing plasmids were cotransfected with chloramphenicol acetyltransferase reporter plasmids in a transient transfection system. We found that MLL contains a strong activation domain and a repression domain. The former, located telomeric (3') to the breakpoint region, activated transcription 18-fold to > 200-fold, depending on the promoter and cell line used for transfection. A repression domain that repressed transcription 4-fold was located centromeric (5') to the breakpoint region of MLL. The MLL AT-hook domain protein was expressed in bacteria and was utilized in a gel mobility shift assay to assess DNA-binding activity. The MLL AT-hook domain could bind cruciform DNA, recognizing structure rather than sequence of the target DNA. In translocations involving MLL, loss of an activation domain with retention of a repression domain and a DNA-binding domain on the der(11) chromosome could alter the expression of downstream target genes, suggesting a potential mechanism of action for MLL in leukemia.
在急性淋巴细胞白血病和急性髓细胞白血病中发现的涉及染色体带11q23的易位会破坏MLL基因。该基因编码一种推定的转录因子,与果蝇三体胸节基因产物的锌指及其他结构域以及高迁移率族蛋白的“AT钩”基序具有同源性。为了绘制MLL蛋白潜在的转录激活或抑制结构域,在瞬时转染系统中,将酵母GAL4 DNA结合结构域和表达MLL杂交蛋白的质粒与氯霉素乙酰转移酶报告质粒共转染。我们发现MLL含有一个强激活结构域和一个抑制结构域。前者位于断点区域的端粒侧(3'),根据用于转染的启动子和细胞系,可激活转录18倍至>200倍。一个抑制转录4倍的抑制结构域位于MLL断点区域的着丝粒侧(5')。MLL AT钩结构域蛋白在细菌中表达,并用于凝胶迁移率变动分析以评估DNA结合活性。MLL AT钩结构域可以结合十字形DNA,识别的是靶DNA的结构而非序列。在涉及MLL的易位中,der(11)染色体上激活结构域的缺失以及抑制结构域和DNA结合结构域的保留可能会改变下游靶基因的表达,这提示了MLL在白血病中的一种潜在作用机制。