HIV-1 Nef impairs heterotrimeric G-protein signaling by targeting Gα(i2) for degradation through ubiquitination.
The HIV Nef protein is an important pathogenic factor that modulates cell surface receptor trafficking and impairs cell motility, presumably by interfering at multiple steps with chemotactic receptor signaling. Here, we report that a dominant effect of Nef is to trigger AIP4 E3 ligase-mediated Gα(i2) ubiquitination, which leads to Gα(i2) endolysosomal sequestration and destruction. The loss of the Gα(i2) subunit was demonstrable in many cell types in the context of gene transfection, HIV infection, or Nef protein transduction. Nef directly interacts with Gα(i2) and ternary complexes containing AIP4, Nef, and Gα(i2) form. A substantial reversal of Gα(i2) loss and a partial recovery of impaired chemotaxis occurred following siRNA knockdown of AIP4 or NEDD4 or by inhibiting dynamin. The N-terminal myristoyl group, (62)EEEE(65) motif, and (72)PXXP(75) motif of Nef are critical for this effect to occur. Nef expression does not affect a Gq(i5) chimera where the five C-terminal residues of Gq are replaced with those of Gα(i2). Lysine at position 296 of Gα(i2) was identified as the critical determinant of Nef-induced degradation. By specifically degrading Gα(i2), Nef directly subverts leukocyte migration and homing. Impaired trafficking and homing of HIV Nef-expressing lymphocytes probably contributes to early immune dysfunction following HIV infection.
Laboratory of Molecular Immunology, NIAID, National Institutes of Health, Bethesda, Maryland 20892, USA.
Prabha
P
Chandrasekaran
Monica
M
Buckley
Victoria
V
Moore
Long Qin
LQ
Wang
John H
JH
Kehrl
Sundararajan
S
Venkatesan
D018925:Chemokines / Q000378:metabolism; D002633:Chemotaxis; D019281:Dimerization; D011992:Endosomes / Q000378:metabolism; D031541:Fluorescence Resonance Energy Transfer; D051960:GTP-Binding Protein alpha Subunit, Gi2 / Q000378:metabolism*; D019204:GTP-Binding Proteins / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D007107:Immune System / Q000378:metabolism; D019169:Jurkat Cells; D016086:Lentivirus / Q000235:genetics; D008214:Lymphocytes / Q000166:cytology; D008247:Lysosomes / Q000378:metabolism; D018613:Microscopy, Confocal / Q000379:methods; D043562:Receptors, G-Protein-Coupled / Q000378:metabolism; D015398:Signal Transduction; D025801:Ubiquitin / Q000737:chemistry*; D044767:Ubiquitin-Protein Ligases / Q000737:chemistry; D054311:nef Gene Products, Human Immunodeficiency Virus / Q000378:metabolism / Q000502:physiology*
D018925:Chemokines; D043562:Receptors, G-Protein-Coupled; D025801:Ubiquitin; D054311:nef Gene Products, Human Immunodeficiency Virus; C514837:nef protein, Human immunodeficiency virus 1; D044767:Ubiquitin-Protein Ligases; D019204:GTP-Binding Proteins; D051960:GTP-Binding Protein alpha Subunit, Gi2