The HIV-1 Nef protein and phagocyte NADPH oxidase activation.
Nef, a multifunctional HIV protein, activates the Vav/Rac/p21-activated kinase (PAK) signaling pathway. Given the potential role of this pathway in the activation of the phagocyte NADPH oxidase, we have investigated the effect of the HIV-1 Nef protein on the phagocyte respiratory burst. Microglia (cell line and primary culture) were transduced with lentiviral expression vectors. Expression of Nef did not activate the NADPH oxidase by itself but led to a massive enhancement of the responses to a variety of stimuli (Ca(2+) ionophore, formyl peptide, endotoxin). These effects were not caused by up-regulation of phagocyte NADPH oxidase subunits. Nef mutants lacking motifs involved in the interaction with Vav and PAK failed to reproduce the effects of wild type Nef, suggesting a role for the Vav/Rac/PAK signaling pathway. The following results suggest a key role for Rac in the priming effect of Nef. (i) Inactivation of Rac by Clostridium difficile toxin B abolished the Nef effect. (ii) The fraction of activated Rac1 was increased in Nef-transduced cells, and (iii) the dominant positive Rac1(V12) mutant mimicked the effect of Nef. These results are to our knowledge the first analysis of the effect of Rac activation on the NADPH oxidase in intact phagocytes. Rac activation is not sufficient to stimulate the phagocyte NADPH oxidase; however, it markedly enhances the NADPH oxidase response to other stimuli.
Biology of Ageing Laboratory, Department of Geriatrics, Geneva University Hospitals, Switzerland. fvilhardt@hotmail.com
Frederik
F
Vilhardt
Olivier
O
Plastre
Makoto
M
Sawada
Kazuo
K
Suzuki
Maciej
M
Wiznerowicz
Etsuko
E
Kiyokawa
Didier
D
Trono
Karl-Heinz
KH
Krause
D020816:Amino Acid Motifs; D000818:Animals; D002460:Cell Line; D004789:Enzyme Activation; D015693:Gene Products, nef / Q000502:physiology*; D015497:HIV-1 / Q000502:physiology*; D051379:Mice; D017628:Microglia / Q000201:enzymology; D019255:NADPH Oxidases / Q000378:metabolism*; D015513:Oncogene Proteins / Q000502:physiology; D010586:Phagocytes / Q000201:enzymology*; D021122:Protein Subunits; D051567:Proto-Oncogene Proteins c-vav; D013481:Superoxides / Q000378:metabolism; D054311:nef Gene Products, Human Immunodeficiency Virus; D020830:rac1 GTP-Binding Protein / Q000502:physiology; D018909:src Homology Domains
D015693:Gene Products, nef; D015513:Oncogene Proteins; D021122:Protein Subunits; D051567:Proto-Oncogene Proteins c-vav; C496619:Vav1 protein, mouse; D054311:nef Gene Products, Human Immunodeficiency Virus; D013481:Superoxides; D019255:NADPH Oxidases; D020830:rac1 GTP-Binding Protein