极性蛋白Crumbs胞内域功能保守性研究1Functional-遗传.PDFVIP

极性蛋白Crumbs 胞内域功能保守性研究1 史其萍，曹浩伟，许蕊，张丹丹，黄娟 南京医科大学基础医学院，南京 211166 摘要：跨膜蛋白Crumbs （Crb ）是细胞顶部的决定因子，对上皮细胞顶-底极性的建立和维持起着关键的作 用。其胞内域虽然仅有37 个氨基酸，但对Crb 的功能必不可少。在果蝇 （Drosophila melanogaster ）中， 如果胞内域发生突变，将造成胚胎发育异常、上皮细胞顶底极性丧失等严重后果。Crb 胞内域从果蝇到小 鼠 （Mus musculus ）和人类 （Homo sapiens ）具有很高的同源性，但线虫 （Caenorhabditis elegans ）的两个 Crb 蛋白的胞内域与果蝇和哺乳动物却较为不同。为验证线虫Crb 蛋白胞内域是否功能保守，本文利用基 因组工程法 （Genomic engineering ），将果蝇基因组中Crb 基因编码胞内域的部分替换为一致性和相似性较 远的线虫Crb2 基因的相应区段。与其他Crb 胞内域突变果蝇不同，替换突变体胚胎发育正常，Crb 及其他 极性蛋白的表达和定位正常，胚胎上皮细胞顶底极性能够正确的建立和维持。这些结果证实虽然线虫和果 蝇Crb 蛋白胞内域之间存在大量序列变异，但重要的氨基酸位点和功能模块则完全保守。 关键词：Crumbs ；胞内域；功能保守性；细胞极性 Functional conservation study of polarity protein Crumbs intracellular domain Qiping Shi, Haowei Cao, Rui Xu, Dandan Zhang, Juan Huang Nanjing Medical University, School of Basic Medical Sciences, Nanjing 211166, China Abstract: The transmembrane protein Crumbs (Crb) plays key roles in the establishing and maintaining cell apical-basal polarity in epithelial cells by determining the apical plasma membrane identity. Although its intracellular domain contains only 37 amino acids, it is absolutely essential for its function. In Drosophila, mutations in this intracellular domain result in severe defects in epithelial polarity and abnormal embryonic development. The intracellular domain of Crb shows high homology across species from Drosophila to Mus musculus and Homo sapiens . However, the intracellular domains of the two Crb proteins in C. elegans are rather divergent from those of Drosophila and mammals, raising the question on whether the function of the intracellular domain of the Crb protein is conserved in C. elegans. Using genomic engineering approaches, we replaced the intracellular domain of the Drosophila Crb with that of C. elegans Crb2 (CeCrb2), which has extremely low 收稿日期：201