Mechanism of the spatio-temporal regulation of Ras and Rap1.
Epidermal growth factor (EGF) activates Ras and Rap1 at distinct intracellular regions. Here, we explored the mechanism underlying this phenomenon. We originally noticed that in cells expressing Epac, a cAMP-dependent Rap1 GEF (guanine nucleotide exchange factor), cAMP activated Rap1 at the perinuclear region, as did EGF. However, in cells expressing e-GRF, a recombinant cAMP-responsive Ras GEF, cAMP activated Ras at the peripheral plasma membrane. Based on the uniform cytoplasmic expression of Epac and e-GRF, GEF did not appear to account for the non-uniform increase in the activities of Ras and Rap1. In contrast, when we used probes with reduced sensitivity to GTPase-activating proteins (GAPs), both Ras and Rap1 appeared to be activated uniformly in the EGF-stimulated cells. Furthermore, we calculated the local rate constants of GEFs and GAPs from the video images of Ras activation and found that GAP activity was higher at the central plasma membrane than the periphery. Thus we propose that GAP primarily dictates the spatial regulation of Ras family G proteins, whereas GEF primarily determines the timing of Ras activation.
Ohba
Yusuke
Y
Department of Tumor Virology, Research Institute for Microbial Diseases, Osaka University, Suita, Osaka 565-0871, Japan.
Kurokawa
Kazuo
K
Matsuda
Michiyuki
M
D000818:Animals; D019556:COS Cells; D000242:Cyclic AMP / Q000378:metabolism; D004705:Endocytosis / Q000502:physiology; D004815:Epidermal Growth Factor / Q000378:metabolism; D066246:ErbB Receptors / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000378:metabolism; D020690:GTPase-Activating Proteins / Q000378:metabolism; D005786:Gene Expression Regulation / Q000502:physiology*; D020662:Guanine Nucleotide Exchange Factors / Q000378:metabolism; D020827:rap1 GTP-Binding Proteins / Q000096:biosynthesis / Q000235:genetics*; D018631:ras Proteins / Q000096:biosynthesis / Q000235:genetics*
D020690:GTPase-Activating Proteins; D020662:Guanine Nucleotide Exchange Factors; D004815:Epidermal Growth Factor; D000242:Cyclic AMP; D066246:ErbB Receptors; D019204:GTP-Binding Proteins; D020827:rap1 GTP-Binding Proteins; D018631:ras Proteins