Galphai protein-dependant extracellular signal-regulated kinase-1/2 activation is required for HIV-1 reverse transcription.
BACKGROUND
HIV-1 triggers infection through interaction with the CD4 receptor and the chemokine receptors, CCR5 or CXCR4, on host cells. The involvement of signaling via the chemokine receptors in viral infection remains an issue of debate. We have previously reported that Galphai1 is involved in the signaling triggered by R5 HIV-1 strains through CCR5 binding to facilitate viral replication in unstimulated peripheral blood mononuclear cells. In this study, we pursued the identification of the downstream signaling molecules in CCR5-mediated infection. We also questioned whether CXCR4 using HIV-1 strains induce the same signaling mechanism.


METHODS
We analyzed by western blotting the coreceptor-mediated activation of various mitogen-acitvated protein kinases, including extracellular signal-regulated kinase (ERK)1/2, p38 and c-jun N-terminal kinase in non-stimulated human peripheral blood mononuclear cells. The involvement of Galphai protein in ERK1/2 activation was tested using pertussis toxin. Using real-time PCR, we studied the role of ERK1/2 in the life cycle of HIV-1.


RESULTS
We found that pertussis toxin inhibited the replication of X4 as well as R5 strains. Furthermore, both strains activated a pertussis toxin-sensitive mitogen-activated protein kinase pathway involving mitogen-activated protein kinase kinases-1/2 and ERK1/2. The inhibition of ERK1/2 activation by U0126 and PD98059 blocked both R5 and X4 HIV-1 replication. Furthermore, ERK1/2 activity was required for the completion of HIV-1 reverse transcription.


CONCLUSIONS
Our results show that R5 and X4 HIV-1 strains induce the same Galphai-dependent ERK pathway that facilitates reverse transcription. The identification of the signaling pathway required for optimal viral replication sheds a new light on HIV physiopathology and opens new therapeutic possibilities.
Mettling
Clément
C
Institut de Génétique Humaine, CNRS-UPR1142, Montpellier, France.
Desmetz
Caroline
C
Fiser
Anne-Laure
AL
Réant
Brigitte
B
Corbeau
Pierre
P
Lin
Yea-Lih
YL
D015153:Blotting, Western; D002478:Cells, Cultured; D004789:Enzyme Activation; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000378:metabolism*; D015658:HIV Infections / Q000821:virology*; D015497:HIV-1 / Q000235:genetics* / Q000502:physiology; D006801:Humans; D007963:Leukocytes, Mononuclear / Q000821:virology*; D020935:MAP Kinase Signaling System; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism*; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D037342:Pertussis Toxin / Q000494:pharmacology; D010766:Phosphorylation; D020133:Reverse Transcriptase Polymerase Chain Reaction; D048348:Reverse Transcription / Q000502:physiology*
D037342:Pertussis Toxin; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; D019206:GTP-Binding Protein alpha Subunits, Gi-Go