Shi Q, Li R K, Mickle D A, Jackowski G
Toronto Hospital-General Division, Ontario, Canada.
J Mol Cell Cardiol. 1992 Nov;24(11):1221-9. doi: 10.1016/0022-2828(92)93089-3.
To explore the mechanisms regulating expression of ventricular myosin light chain 1, the human gene including 5'-flanking DNA was cloned and characterized by Southern blot and restriction mapping. A 2 kb 5'-flanking DNA was sequenced and linked to a chloramphenicol acetyltransferase reporter gene. The constructs then were transfected into cultured human and rat cardiomyocytes as well as rat aortic endothelial cells. Deletion analysis of constructs revealed that the basal promoter sequences, which were located within 62 base pairs of the cap site, could direct high levels of chloramphenicol acetyltransferase gene expression in the cardiomyocytes and endothelial cells. The region between -62 to -312 base pairs strongly repressed the chloramphenicol acetyltransferase gene expression in the cardiomyocytes and endothelial cells. Positive elements were found between -312 and -2000 base pairs of the cap site. These results are indicative, among other possibilities, that the human ventricular myosin light chain 1 gene is turned on in cardiomyocytes by the presence of trans-acting factors that are bound to upstream positive elements and is turned off in non-muscle cells by the presence of repressor-binding proteins. But this mechanism remains to be established.
为了探究调节心室肌球蛋白轻链1表达的机制，克隆了包含5'侧翼DNA的人类基因，并通过Southern印迹和限制性图谱分析对其进行了表征。对一段2 kb的5'侧翼DNA进行了测序，并将其与氯霉素乙酰转移酶报告基因相连。然后将构建体转染到培养的人类和大鼠心肌细胞以及大鼠主动脉内皮细胞中。对构建体的缺失分析表明，位于帽位点62个碱基对内的基础启动子序列可在心肌细胞和内皮细胞中指导高水平的氯霉素乙酰转移酶基因表达。-62至-312个碱基对之间的区域强烈抑制心肌细胞和内皮细胞中氯霉素乙酰转移酶基因的表达。在帽位点的-312至-2000个碱基对之间发现了正向元件。这些结果表明，除其他可能性外，人类心室肌球蛋白轻链1基因在心肌细胞中通过与上游正向元件结合的反式作用因子的存在而开启，在非肌肉细胞中通过阻遏物结合蛋白的存在而关闭。但这一机制仍有待确定。
