Interactions among IQGAP1, Cdc42, and the cadherin/catenin protein complex regulate sertoli-germ cell adherens junction dynamics in the testis
Interactions among IQGAP1, Cdc42, and the cadherin/catenin protein complex regulate sertoli-germ cell adherens junction dynamics in the testis
复制标题
DOI：
10.1002/jcp.20098
复制
发表时间：
2005-01-01
影响因子：
5.6
通讯作者：
Cheng, CY
中科院分区：
文献类型：
--
作者：
Lui, WY;Mruk, DD;Cheng, CY
在精子发生过程中，发育中的生殖细胞穿过生精上皮的运动涉及支持细胞之间以及支持细胞和生殖细胞之间的广泛粘附连接（AJ）重建。在这份报告中，我们表明Cdc 42之间复杂的相互作用，（一种Rho家族蛋白，分子量为23 kDa，最初在人血小板和胎盘的膜中鉴定，是CDC 42 Sc的同系物，已知其调节酿酒酵母中的芽位点组装）及其效应物，（IQ）在bar基序下，含有（G）在bar Tactinase激活蛋白下（IQGAP 1，分子量189 kDa，也是一种肌动蛋白结合蛋白，与Cdc 42和Rac 1 GTP酶相互作用），调节支持-生殖细胞，但不调节支持-支持细胞，AJ动力学。使用睾丸裂解物进行免疫沉淀（IP），IQGAP 1显示与E-钙粘蛋白、N-钙粘蛋白和β-连环蛋白（但不与β 1-整联蛋白和nectin-2）以及肌动蛋白和波形蛋白（但不与pi-微管蛋白）相关。此外，IQGAP 1被发现本地化的支持和生殖细胞在生精上皮的细胞-细胞接触的网站的周边。使用荧光显微镜与双荧光探针，IQGAP 1被发现共定位，至少部分，与N-钙粘蛋白在生精上皮一致，其本地化在基底和顶端ES。使用Sertoli生殖细胞共培养物，证明AJ组装与Cdc 42和IQGAP 1的瞬时诱导相关，这在单独培养Sertoli细胞时没有发现。最后，Cdc 42，IQGAP 1，β-连环蛋白，和N-钙粘蛋白的相互作用的转变中检测到支持生殖细胞共培养使用C2+诱导的AJ破坏模型，这是用来检查AJ拆卸和重新组装。在Ca ~（2+）存在下，IQGAP 1优先与Cdc 42结合，而不是与β-连环蛋白结合。然而，当使用EGTA（一种Ca 2+螯合剂）从共培养物中耗尽Ca 2+时，IQGAP 1失去了对Cdc 42的亲和力，并与β-连环蛋白紧密结合，使支持细胞和生殖细胞之间的钙粘蛋白介导的AJs不稳定。然而，在单独培养的支持细胞中没有检测到这种蛋白质-蛋白质相互作用的转变。这些结果说明IQGAP 1、Cdc 42和β-catenin之间的相互作用对生精上皮中支持-生殖细胞的调控至关重要，但对支持-支持细胞、AJ动态的调控无关。(C)2004 Wiley-Liss，Inc.
The movement of developing germ cells across the seminiferous epithelium during spermatogenesis involves extensive adherens junction (AJ) restructuring between Sertoli cells, as well as between Sertoli and germ cells. In this report, we show that the intricate interactions between Cdc42 (a Rho family protein of Mr similar to23 kDa originally identified in membranes of human platelets and placenta, and is the homolog of CDC42Sc, which is known to regulate of bud-site assembly in Saccharomyces cerevisiae) and its effector, (IQ) under bar motif containing (G) under bar TPase activating protein (IQGAP1, Mr similar to189 kDa, it is also an actin-binding protein known to interact with Cdc42 and Rac1 GTPases), regulate Sertoli-germ cell, but not Sertoli-Sertoli cell, AJ dynamics. Using testis lysates for immunoprecipitation (IP), IQGAP1 was shown to associate with E-cadherin, N-cadherin, and beta-catenin (but not beta1-integrin and nectin-2), as well as with actin and vimentin (but not pi-tubulin). Moreover, IQGAP1 was found to localize to the periphery of both Sertoli and germ cells in the seminiferous epithelium, at sites of cell-cell contacts. Using fluorescent microscopy with dual fluorescent probes, IQGAP1 was found to co-localize, at least in part, with N-cadherin in the seminiferous epithelium consistent with their localization at the basal and apical ES. Using Sertoli-germ cell cocultures, it was demonstrated that AJ assembly associated with a transient induction of Cdc42 and IQGAP1, which was not found when Sertoli cells were cultured alone. Lastly, a shift in the interactions of Cdc42, IQGAP1, beta-catenin, and N-cadherin was detected in Sertoli-germ cell cocultures using an C2+-induced AJ disruption model, which was used to examine AJ disassembly and its reassembly. In the presence of Ca2+ IQGAP1 bound preferentially to Cdc42 rather than to P-catenin. However, when Ca2+ was depleted from cocultures using EGTA, a Ca2+ chelating agent, IQGAP1 lost its affinity for Cdc42 and became tightly associated with beta-catenin, destabilizing cadherin-mediated AJs between Sertoli and germ cells. Yet this shift of protein-protein interaction was not detected in Sertoli cells cultured alone. These results illustrate that the interactions among IQGAP1, Cdc42, and beta-catenin are crucial to the regulation of Sertoli-germ cell, but not Sertoli-Sertoli cell, AJ dynamics in the seminiferous epithelium. (C) 2004 Wiley-Liss, Inc.