Phillips-Mason Polly J, Mourton Tracy, Major Denice L, Brady-Kalnay Susann M
Department of Molecular Biology and Microbiology, School of Medicine, Case Western Reserve University, Cleveland, Ohio 44106-4960, USA.
J Cell Biochem. 2008 Nov 1;105(4):1059-72. doi: 10.1002/jcb.21907.
The receptor protein tyrosine phosphatase PTPmu belongs to a family of adhesion molecules that contain cell-cell adhesion motifs in their extracellular segments and catalytic domains within their intracellular segments. The ability of PTPmu both to mediate adhesion and exhibit enzymatic activity makes PTPmu an excellent candidate to transduce signals in response to cell-cell adhesion. In an effort to identify downstream signaling partners of PTPmu, we performed a modified yeast two-hybrid screen using the first tyrosine phosphatase domain of PTPmu as bait. We isolated an interacting clone encoding BRCA2 and CDKN1A interacting protein (BCCIP) from a HeLa cell library. BCCIP is a p21 and BRCA2 interacting protein that has been shown to play roles in both cell cycle arrest and DNA repair. In this manuscript, we confirm the interaction between BCCIP and PTPmu identified in yeast using in vitro biochemical studies and characterize BCCIP as a PTPmu binding protein. We demonstrate that BCCIP is phosphorylated by the Src tyrosine kinase and dephosphorylated by the PTPmu tyrosine phosphatase in vitro. Furthermore, we show that BCCIP is required for both the permissive and repulsive functions of PTPmu in neurite outgrowth assays, suggesting BCCIP and PTPmu are in a common signal transduction pathway.
受体蛋白酪氨酸磷酸酶PTPmu属于一类粘附分子家族,其细胞外段含有细胞间粘附基序,细胞内段含有催化结构域。PTPmu既能介导粘附又能展现酶活性,这使得它成为响应细胞间粘附来转导信号的优秀候选者。为了鉴定PTPmu的下游信号传导伙伴,我们使用PTPmu的第一个酪氨酸磷酸酶结构域作为诱饵进行了改良的酵母双杂交筛选。我们从HeLa细胞文库中分离出一个编码BRCA2和CDKN1A相互作用蛋白(BCCIP)的相互作用克隆。BCCIP是一种p21和BRCA2相互作用蛋白,已被证明在细胞周期停滞和DNA修复中都发挥作用。在本论文中,我们使用体外生化研究证实了酵母中鉴定出的BCCIP与PTPmu之间的相互作用,并将BCCIP表征为一种PTPmu结合蛋白。我们证明BCCIP在体外被Src酪氨酸激酶磷酸化,并被PTPmu酪氨酸磷酸酶去磷酸化。此外,我们表明在神经突生长试验中,BCCIP是PTPmu的允许和排斥功能所必需的,这表明BCCIP和PTPmu处于共同的信号转导途径中。