Xue D, Finney M, Ruvkun G, Chalfie M
Department of Biological Sciences, Columbia University, New York, NY 10027.
EMBO J. 1992 Dec;11(13):4969-79. doi: 10.1002/j.1460-2075.1992.tb05604.x.
The mec-3 gene encodes a homeodomain protein with LIM repeats that is required for the specification of touch cell fate in Caenorhabditis elegans. Previous experiments suggested that mec-3 expression requires the product of the unc-86 gene, a POU-type homeoprotein, and mec-3 itself. We have analyzed the control of mec-3 expression by identifying potential cis regulatory elements in the mec-3 gene (by conservation in a related nematode and by DNase I footprinting using unc-86 and mec-3 proteins) and testing their importance by transforming C.elegans with mec-3lacZ fusions in which these sites have been mutagenized in vitro. Both unc-86 and mec-3 proteins bind specifically to the promoter of the mec-3 gene, suggesting that both proteins may be directly involved in the regulation of the mec-3 gene. In addition, the footprint pattern with mec-3 protein is altered in the presence of unc-86 protein. In vivo transformation experiments reveal that some of the binding regions of the two proteins are needed for general positive control and maintenance of mec-3 expression while others have no detectable, unique function. Interestingly, the unc-86 gene appears to be required not only to initiate mec-3 expression but also to maintain it.
mec-3基因编码一种带有LIM重复序列的同源结构域蛋白,该蛋白是秀丽隐杆线虫触觉细胞命运特化所必需的。先前的实验表明,mec-3的表达需要unc-86基因的产物(一种POU型同源蛋白)以及mec-3自身。我们通过鉴定mec-3基因中的潜在顺式调控元件(通过在相关线虫中的保守性以及使用unc-86和mec-3蛋白进行DNase I足迹分析)来分析mec-3表达的调控,并通过用mec-3lacZ融合体转化秀丽隐杆线虫来测试它们的重要性,其中这些位点已在体外进行了诱变。unc-86和mec-3蛋白都特异性地结合到mec-3基因的启动子上,这表明这两种蛋白可能都直接参与mec-3基因的调控。此外,在存在unc-86蛋白的情况下,mec-3蛋白的足迹模式会发生改变。体内转化实验表明,这两种蛋白的一些结合区域对于mec-3表达的一般阳性调控和维持是必需的,而其他区域则没有可检测到的独特功能。有趣的是,unc-86基因似乎不仅是启动mec-3表达所必需的,也是维持其表达所必需的。