Role of clathrin-mediated endocytosis in CXCR2 sequestration, resensitization, and signal transduction.
CXCR2 is a seven-transmembrane receptor that transduces intracellular signals in response to the chemokines interleukin-8, melanoma growth-stimulatory activity/growth-regulatory protein, and other ELR motif-containing CXC chemokines by coupling to heterotrimeric GTP-binding proteins. In this study, we explored the mechanism responsible for ligand-induced CXCR2 endocytosis. Here, we demonstrate that dynamin, a component of clathrin-mediated endocytosis, is essential for CXCR2 endocytosis and resensitization. In HEK293 cells, dynamin I K44A, a dominant-negative mutant of dynamin that inhibits the clathrin-mediated endocytosis, blocks the ligand-stimulated CXCR2 sequestration. Furthermore, co-expression of dynamin I K44A significantly delays dephosphorylation of CXCR2 after ligand stimulation, suggesting that clathrin-mediated endocytosis plays an important role in receptor dephosphorylation and resensitization. In addition, ligand-mediated receptor down-regulation is attenuated when receptor internalization is inhibited by dynamin I K44A. Interestingly, inhibition of receptor endocytosis by dynamin I K44A does not affect the CXCR2-mediated stimulation of mitogen-activated protein kinase. Most significantly, our data indicate that the ligand-stimulated receptor endocytosis is required for CXCR2-mediated chemotaxis in HEK293 cells. Taken together, our findings suggest that clathrin-mediated CXCR2 internalization is crucial for receptor endocytosis, resensitization, and chemotaxis.
Yang
W
W
Department of Cell Biology, Vanderbilt University School of Medicine, Nashville, Tennessee 37232-2175, USA.
Wang
D
D
Richmond
A
A
D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism; D002460:Cell Line; D054360:Chemokine CXCL1; D019743:Chemokines, CXC / Q000378:metabolism; D002630:Chemotactic Factors / Q000494:pharmacology; D002633:Chemotaxis; D002966:Clathrin / Q000378:metabolism*; D015536:Down-Regulation; D034284:Dynamin I; D034281:Dynamins; D004705:Endocytosis / Q000187:drug effects; D004789:Enzyme Activation; D020558:GTP Phosphohydrolases / Q000494:pharmacology; D006133:Growth Substances / Q000494:pharmacology; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins; D016209:Interleukin-8 / Q000378:metabolism; D010766:Phosphorylation; D019707:Receptors, Chemokine / Q000378:metabolism*; D018123:Receptors, Interleukin / Q000378:metabolism*; D023063:Receptors, Interleukin-8B; D015398:Signal Transduction
C475829:CXCL1 protein, human; D054360:Chemokine CXCL1; D019743:Chemokines, CXC; D002630:Chemotactic Factors; D002966:Clathrin; D006133:Growth Substances; D036341:Intercellular Signaling Peptides and Proteins; D016209:Interleukin-8; D019707:Receptors, Chemokine; D018123:Receptors, Interleukin; D023063:Receptors, Interleukin-8B; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D034284:Dynamin I; D020558:GTP Phosphohydrolases; D034281:Dynamins