Trask R V, Koster J C, Ritchie M E, Billadello J J
Department of Medicine, Washington University School of Medicine, St Louis, MO 63110.
Nucleic Acids Res. 1992 May 11;20(9):2313-20. doi: 10.1093/nar/20.9.2313.
Cis-elements (-933 to -641) upstream of the human M creatine kinase gene cap site contain an enhancer that confers developmental and tissue-specific expression to the chloramphenicol acetyltransferase gene in C2C12 myogenic cells transfected in culture. Division of the enhancer at -770 into a 5' fragment that includes the MyoD binding sites (-933 to -770) and a 3' fragment that includes the MEF-2 binding site (-770 to -641) resulted in two subfragments that showed minimal activity but in combination interacted in a position- and orientation-independent fashion to enhance activity of the SV40 promoter in transient transfection experiments. A 5' enhancer construct (-877 to -832) including only one (the low affinity) MyoD binding site was active when present in multiple copies. In contrast, a 3' enhancer construct (-749 to -732) including the MEF-2 binding site was inactive even when present in multiple copies. However, if the 5' construct was extended to include the high-affinity MyoD binding site (-877 to -803) the 5' and 3' constructs interacted in a position- and orientation-independent fashion to activate the SV40 promoter. Thus, the human M creatine kinase enhancer comprises multiple functional interacting domains.
人类M型肌酸激酶基因帽位点上游的顺式元件（-933至-641）包含一个增强子，该增强子可赋予氯霉素乙酰转移酶基因在培养中转染的C2C12成肌细胞中进行发育和组织特异性表达。将-770处的增强子分为一个包含MyoD结合位点的5'片段（-933至-770）和一个包含MEF-2结合位点的3'片段（-770至-641），产生了两个亚片段，它们显示出最小的活性，但在瞬时转染实验中，它们以位置和方向独立的方式相互作用，以增强SV40启动子的活性。当以多拷贝形式存在时，仅包含一个（低亲和力）MyoD结合位点的5'增强子构建体（-877至-832）具有活性。相比之下，包含MEF-2结合位点的3'增强子构建体（-749至-732）即使以多拷贝形式存在也无活性。然而，如果将5'构建体扩展以包含高亲和力MyoD结合位点（-877至-803），则5'和3'构建体以位置和方向独立的方式相互作用以激活SV40启动子。因此，人类M型肌酸激酶增强子包含多个功能性相互作用结构域。
