HIV-1 Nef disrupts the podocyte actin cytoskeleton by interacting with diaphanous interacting protein.
The ability of the human immunodeficiency virus, type 1 (HIV-1) protein Nef to induce cytoskeleton changes in infected host cells is a key event in viral replication. In renal podocytes, we found that Nef induced loss of stress fibers and increased lamellipodia, pathological changes leading to proteinuria in HIV-associated nephropathy. These morphological changes were mediated by Nef-induced Rac1 activation and RhoA inhibition. We identified a new interaction between Nef and diaphanous interacting protein (DIP), a recently described regulator of Rho and Rac signaling. We found that the Src homology 3 binding domain of DIP and the Nef PXXP motif were required for this interaction. Nef also interacts with Vav2 in podocytes. DIP and Vav2 both interact directly with Nef in a competitive manner. DIP interacts with p190RhoGAP, and intact DIP was required for Nef-induced phosphorylation of p190RhoGAP. DIP also interacts with Vav2, and although DIP enhanced baseline phosphorylation of Vav2, it was not required for Nef-induced Vav2 activation. In Nef-infected podocytes, Src kinase induces phosphorylation of DIP, p190RhoGAP, and Vav2, leading to RhoA inhibition and Rac1 activation. Inhibition of the Nef-induced signaling pathway by using a dominant negative of either Src or DIP or siRNA for DIP or p190RhoAGAP restored RhoA activity and stress fiber formation in Nef-infected podocytes, whereas siRNA for Vav2 reduced Rac1 activity and formation of lamellipodia. We conclude that in HIV-infected podocytes, Nef, through the recruitment of DIP and p190RhoAGAP to Nef-Src complex, activates p190RhoAGAP and down-regulates RhoA activity.
Lu
Ting-Chi
TC
Department of Medicine, Mount Sinai School of Medicine, New York, NY 10029, USA.
He
John Cijiang
JC
Wang
Zhao-Hui
ZH
Feng
Xiaobei
X
Fukumi-Tominaga
Tomoko
T
Chen
Nan
N
Xu
Jin
J
Iyengar
Ravi
R
Klotman
Paul E
PE
D000199:Actins / Q000378:metabolism*; D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism*; D000818:Animals; D003599:Cytoskeleton / Q000378:metabolism*; D004789:Enzyme Activation; D005786:Gene Expression Regulation; D015497:HIV-1 / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D009124:Muscle Proteins / Q000378:metabolism*; D010766:Phosphorylation; D050199:Podocytes / Q000378:metabolism*; D011485:Protein Binding; D051567:Proto-Oncogene Proteins c-vav / Q000378:metabolism; D015398:Signal Transduction; D054311:nef Gene Products, Human Immunodeficiency Virus / Q000235:genetics / Q000378:metabolism*; D020830:rac1 GTP-Binding Protein / Q000378:metabolism; D020742:rhoA GTP-Binding Protein / Q000378:metabolism
D000199:Actins; D048868:Adaptor Proteins, Signal Transducing; D009124:Muscle Proteins; C426229:Nckipsd protein, mouse; D051567:Proto-Oncogene Proteins c-vav; C490565:Vav2 protein, mouse; D054311:nef Gene Products, Human Immunodeficiency Virus; C514837:nef protein, Human immunodeficiency virus 1; D020830:rac1 GTP-Binding Protein; D020742:rhoA GTP-Binding Protein