Rack1 binds HIV-1 Nef and can act as a Nef-protein kinase C adaptor.
Nef proteins of primate immunodeficiency viruses exert pleiotropic effects, such as enhanced endocytosis of CD4 and MHC-I cell surface molecules, perturbation of signal transduction cascades, and virion infectivity enhancement. Nef function intersects that of a number of cell kinases, including C kinases (PKCs) and Src-family kinases. Here the interaction of HIV-1 Nef with Rack1 (receptor for activated C kinase 1) is reported. Nef binds the Rack1 C-terminal moiety in a yeast two-hybrid system and in cell-free pull-down assays and copurifies with in vitro translated Rack1. Nef and Rack1 partially colocalize on the trans-Golgi network and plasma membranes. The presence of Rack1 doubles Nef phosphorylation by PKCs in vitro. Our data agree with the idea that Rack1 acts as a Nef intracellular docking site, bringing Nef and PKCs together. Other signal transduction or endocytosis proteins, in particular Src-like kinases, might meet Nef by intermediation of the Rack1 adaptor.
Gallina
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A
Department of Medicine and Surgery, Polo Universitario Ospedale San Paolo, University of Milano, via A. di Rudiní 8, I-20142 Milan, Italy.
Rossi
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F
Milanesi
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G
D000818:Animals; D002460:Cell Line; D005455:Fluorescent Antibody Technique; D015693:Gene Products, nef / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D010455:Peptides / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D010957:Plasmids; D011493:Protein Kinase C / Q000378:metabolism*; D051381:Rats; D000075370:Receptors for Activated C Kinase; D014162:Transfection; D020798:Two-Hybrid System Techniques
D015693:Gene Products, nef; D010455:Peptides; D000075370:Receptors for Activated C Kinase; C070301:peptide I; D011493:Protein Kinase C