Abstract
Lentiviral Nef increases T cell signaling activity, but the molecular nature of the stimulus involved is incompletely described. We explored CD4 T cell lipid raft composition in the presence and absence of Nef. Here, the E2 ubiquitin-conjugating enzyme UbcH7, which acts in conjunction with c-Cbl, is absent from lipid rafts. This Nef-mediated exclusion is associated with failure of ubiquitination of activated Vav. In the presence of Nef, lipid raft Cdc42 is activated and forms a ternary complex between the c-Cbl-interacting protein p85Cool-1/betaPix and c-Cbl, displacing UbcH7 from rafts. Suppression of p85Cool-1/betaPix expression restores UbcH7 raft localization and Vav ubiquitination and diminishes Cdc42 activity. Moreover, p85Cool-1/betaPix knockdown attenuates HIV replication. Thresholds for activation of signaling involve the intricate balance of positive and negative regulators. Here we provide evidence for Nef disruption of a negative regulator of T cell signaling in promoting HIV replication.
Publication types
-
Comparative Study
-
Research Support, Non-U.S. Gov't
MeSH terms
-
Blotting, Western
-
CD4-Positive T-Lymphocytes / cytology
-
CD4-Positive T-Lymphocytes / immunology*
-
Cell Cycle Proteins / metabolism
-
Cell Line
-
Enzyme-Linked Immunosorbent Assay
-
Gene Products, nef / immunology
-
Gene Products, nef / metabolism*
-
Guanine Nucleotide Exchange Factors / metabolism
-
HIV*
-
Humans
-
Immunoprecipitation
-
Membrane Microdomains / immunology
-
Membrane Microdomains / metabolism*
-
Microscopy, Confocal
-
Proto-Oncogene Proteins c-cbl / immunology
-
Proto-Oncogene Proteins c-cbl / metabolism*
-
RNA, Small Interfering / genetics
-
Rho Guanine Nucleotide Exchange Factors
-
Signal Transduction / immunology*
-
Ubiquitin-Conjugating Enzymes / immunology
-
Ubiquitin-Conjugating Enzymes / metabolism*
-
Virus Replication / immunology*
-
cdc42 GTP-Binding Protein / metabolism
-
nef Gene Products, Human Immunodeficiency Virus
Substances
-
Cell Cycle Proteins
-
Gene Products, nef
-
Guanine Nucleotide Exchange Factors
-
RNA, Small Interfering
-
Rho Guanine Nucleotide Exchange Factors
-
nef Gene Products, Human Immunodeficiency Virus
-
UBE2L3 protein, human
-
Ubiquitin-Conjugating Enzymes
-
Proto-Oncogene Proteins c-cbl
-
cdc42 GTP-Binding Protein
-
CBL protein, human