Bivona Trever G, Wiener Heidi H, Ahearn Ian M, Silletti Joseph, Chiu Vi K, Philips Mark R
Deparment of Cell Biology, New York University School of Medicine, 550 First Ave., New York, NY 10016, USA.
J Cell Biol. 2004 Feb 2;164(3):461-70. doi: 10.1083/jcb.200311093.
Rap1 and Ras are closely related GTPases that share some effectors but have distinct functions. We studied the subcellular localization of Rap1 and its sites of activation in living cells. Both GFP-tagged Rap1 and endogenous Rap1 were localized to the plasma membrane (PM) and endosomes. The PM association of GFP-Rap1 was dependent on GTP binding, and GFP-Rap1 was rapidly up-regulated on this compartment in response to mitogens, a process blocked by inhibitors of endosome recycling. A novel fluorescent probe for GTP-bound Rap1 revealed that this GTPase was transiently activated only on the PM of both fibroblasts and T cells. Activation on the PM was blocked by inhibitors of endosome recycling. Moreover, inhibition of endosome recycling blocked the ability of Rap1 to promote integrin-mediated adhesion of T cells. Thus, unlike Ras, the membrane localizations of Rap1 are dynamically regulated, and the PM is the principle platform from which Rap1 signaling emanates. These observations may explain some of the biological differences between these GTPases.
Rap1和Ras是密切相关的GTP酶，它们共享一些效应器，但具有不同的功能。我们研究了Rap1在活细胞中的亚细胞定位及其激活位点。绿色荧光蛋白（GFP）标记的Rap1和内源性Rap1都定位于质膜（PM）和内体。GFP-Rap1与质膜的结合依赖于GTP结合，并且GFP-Rap1在该区室中响应有丝分裂原而迅速上调，这一过程被内体再循环抑制剂所阻断。一种用于检测结合GTP的Rap1的新型荧光探针显示，这种GTP酶仅在成纤维细胞和T细胞的质膜上被短暂激活。质膜上的激活被内体再循环抑制剂所阻断。此外，内体再循环的抑制阻断了Rap1促进T细胞整合素介导的粘附的能力。因此，与Ras不同，Rap1的膜定位是动态调节的，质膜是Rap1信号传导发出的主要平台。这些观察结果可能解释了这些GTP酶之间的一些生物学差异。
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