Molecular characterization of 4A, a novel member of the MAGUK family

• Main Authors: Lin-Chun Chang, 張翎君
• Other Authors: Wey-Jinq Lin
• Format: Others
• Language: zh-TW
• Published: 2004
• Online Access: http://ndltd.ncl.edu.tw/handle/96243816799088605189

碩士 === 國立陽明大學 === 生物藥學研究所 === 92 === Abstract MAGUKs (membrane-associated guanylate kinase homologues) family members function in a variety of cellular processes including tumor suppression and receptor clustering. The previous results from ours lab reported that the rat 4A gene, cloned by a yeast two-hybrid screening using the antiproliferative gene TIS21 as a bait, encodes a novel member of MAGUK family. The deduced protein sequence of 4A shows 71% identity to that of mouse p55/MPP1. MPP1 does not interact with TIS21 in the yeast two-hybrid analysis. In contrast to 4A, MPP1 does not enhance the accumulation of TIS21 protein in vivo. The 4A and MPP1 messages are expressed at different levels in various human cell lines. Antibodies obtained using a peptide specific to hMPP1 recognizes MPP1 but not 4A in western blotting analysis using GST-hMPP1 as antigens. TIS21 is an immediated-early gene with antiproliferative acitivity. In the presence of the protein synthesis inhibitor cycloheximide, the calculated half-life of TIS21 protein is 30 minutes in NIH3T3 cells. 4A has been shown to enhance accumulation of TIS21 proteins. In this studies, we show that 4A stabilizes TIS21 and prolongs its half-life to 6 hours. TIS21 proteins are degraded by ubiquitin-proteasome system in NIH3T3 cells. The TIS21S147A protein in which CDK1 phosphorylation site is mutated, shows a similar protein degradation pattern upon 4A overexpression. This indicates that the CDK1 phosphorylation site may not be involved in TIS21 degradation processes.