Kawata Akira, Iida Junko, Ikeda Mitsunobu, Sato Yuji, Mori Hiroki, Kansaku Ai, Sumita Kazutaka, Fujiwara Naoyuki, Rokukawa Chiaki, Hamano Mamiko, Hirabayashi Susumu, Hata Yutaka
Department of Medical Biochemistry, Graduate School of Medicine, Tokyo Medical and Dental University, Tokyo 113-8519.
J Biochem. 2006 May;139(5):931-9. doi: 10.1093/jb/mvj105.
Membrane-associated guanylate kinase inverted (MAGI)-1 plays a role as a scaffold at cell junctions in non-neuronal cells, while S-SCAM, its neuronal isoform, is involved in the organization of synapses. A search for MAGI-1-interacting proteins by yeast two-hybrid screening of a kidney cDNA library yielded dendrin. As dendrin was originally reported as a brain-specific postsynaptic protein, we tested the interaction between dendrin and S-SCAM and revealed that dendrin binds to the WW domains of S-SCAM. Dendrin is known to be dendritically translated but its function is largely unknown. To gain insights into the physiological meaning of the interaction, we performed a second yeast two-hybrid screening using dendrin as a bait. We identified CIN85, an endocytic scaffold protein, as a putative dendrin-interactor. Immunocytochemistry and subcellular fractionation analysis supported the synaptic localization of CIN85. The first SH3 domain and the C-terminal region of CIN85 bind to the proline-rich region and the N-terminal region of dendrin, respectively. In vitro experiments suggest that dendrin forms a ternary complex with CIN85 and S-SCAM and that this complex formation facilitates the recruitment of dendrin and S-SCAM to vesicle-like structures where CIN85 is accumulated.
膜相关鸟苷酸激酶反向异构体（MAGI）-1在非神经元细胞的细胞连接处作为支架发挥作用，而其神经元异构体S-SCAM则参与突触的组织。通过对肾脏cDNA文库进行酵母双杂交筛选寻找与MAGI-1相互作用的蛋白质，得到了树突蛋白。由于树突蛋白最初被报道为一种脑特异性突触后蛋白，我们测试了树突蛋白与S-SCAM之间的相互作用，发现树突蛋白与S-SCAM的WW结构域结合。已知树突蛋白是在树突中翻译的，但其功能在很大程度上尚不清楚。为了深入了解这种相互作用的生理意义，我们以树突蛋白为诱饵进行了第二轮酵母双杂交筛选。我们鉴定出一种内吞支架蛋白CIN85为假定的树突蛋白相互作用蛋白。免疫细胞化学和亚细胞分级分析支持CIN85在突触中的定位。CIN85的第一个SH3结构域和C末端区域分别与树突蛋白的富含脯氨酸区域和N末端区域结合。体外实验表明，树突蛋白与CIN85和S-SCAM形成三元复合物，这种复合物的形成促进了树突蛋白和S-SCAM向CIN85积累的囊泡样结构的募集。
