Sec14 homology domain targets p50RhoGAP to endosomes and provides a link between Rab and Rho GTPases.
Sec14 protein was first identified in Saccharomyces cerevisiae, where it serves as a phosphatidylinositol transfer protein that is essential for the transport of secretory proteins from the Golgi complex. A protein domain homologous to Sec14 was identified in several mammalian proteins that regulates Rho GTPases, including exchange factors and GTPase activating proteins. P50RhoGAP, the first identified GTPase activating protein for Rho GTPases, is composed of a Sec14-like domain and a Rho-GTPase activating protein (GAP) domain. The biological function of its Sec14-like domain is still unknown. Here we show that p50RhoGAP is present on endosomal membranes, where it colocalizes with internalized transferrin receptor. We demonstrate that the Sec14-like domain of P50RhoGAP is responsible for the endosomal targeting of the protein. We also show that overexpression of p50RhoGAP or its Sec14-like domain inhibits transferrin uptake. Furthermore, both P50RhoGAP and its Sec14-like domain show colocalization with small GTPases Rab11 and Rab5. We measured bioluminescence resonance energy transfer between p50RhoGAP and Rab11, indicating that these proteins form molecular complex in vivo on endosomal membranes. The interaction was mediated by the Sec 14-like domain of p50RhoGAP. Our results indicate that Sec14-like domain, which was previously considered as a phospholipid binding module, may have a role in the mediation of protein-protein interactions. We suggest that p50RhoGAP provides a link between Rab and Rho GTPases in the regulation of receptor-mediated endocytosis.
Sirokmány
Gábor
G
Department of Physiology, Semmelweis University, P. O. Box 259, 1444 Budapest, Hungary.
Szidonya
László
L
Káldi
Krisztina
K
Gáborik
Zsuzsanna
Z
Ligeti
Erzsébet
E
Geiszt
Miklós
M
D000818:Animals; D004705:Endocytosis / Q000502:physiology; D011992:Endosomes / Q000378:metabolism*; D020690:GTPase-Activating Proteins / Q000235:genetics / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D007425:Intracellular Membranes / Q000378:metabolism; D008870:Microtubules / Q000378:metabolism; D049294:Phospholipid Transfer Proteins / Q000235:genetics / Q000378:metabolism; D011990:Receptors, Transferrin / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism; D014168:Transferrin / Q000378:metabolism; D020764:cdc42 GTP-Binding Protein / Q000378:metabolism; D020691:rab GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D020741:rho GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*
D020690:GTPase-Activating Proteins; D049294:Phospholipid Transfer Proteins; D011990:Receptors, Transferrin; D011993:Recombinant Fusion Proteins; C483438:SEC14 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D014168:Transferrin; C085379:rho GTPase-activating protein; D020764:cdc42 GTP-Binding Protein; D020691:rab GTP-Binding Proteins; D020741:rho GTP-Binding Proteins