HsN3 proteasomal subunit as a target for human immunodeficiency virus type 1 Nef protein.
HIV-1 Nef protein is important for pathogenicity, but its biochemical function remains obscure. To clarify its role, a yeast two-hybrid system (ths) screening was utilized to identify Nef cellular partners. Of 79 yeast clones harboring cDNAs for putative Nef binding proteins, 27 (34%) contained the coding region for HsN3 proteasomal ... subunit. HsN3 behaved as bona fide Nef partner in ths control crosses. Nef-HsN3 interaction was confirmed by in vitro binding experiments. In particular, recombinant Nef was able to capture the HsN3 subunit from a natural proteasome preparation. In Nef, the interacting region was mapped within aa 34-143, which span the structured portion of the protein, including the SH3-binding domain. In HsN3, Nef-binding portion was restricted to aa 73-249, and the tract 219-249-reminiscent of SH3 domain N-terminal 3/5ths-was shown to be essential, though not sufficient. Attempts to purify a Nef-HsN3 complex from transfected COS7 cells were unsuccessful. However, Nef was found to markedly downregulate intracellular levels of both a coexpressed HsN3 and the endogenous simian homologue. These results suggest that Nef, by binding to a subunit, might alter proteasome function in infected cells.
Mesh Terms:
Amino Acid Sequence, Animals, Binding Sites, COS Cells, Cysteine Endopeptidases, Gene Products, nef, HIV-1, Humans, Molecular Sequence Data, Multienzyme Complexes, Plasmids, Proteasome Endopeptidase Complex, Protein Binding, Saccharomyces cerevisiae, Sequence Alignment, Transfection, Virus Replication, nef Gene Products, Human Immunodeficiency Virus
Amino Acid Sequence, Animals, Binding Sites, COS Cells, Cysteine Endopeptidases, Gene Products, nef, HIV-1, Humans, Molecular Sequence Data, Multienzyme Complexes, Plasmids, Proteasome Endopeptidase Complex, Protein Binding, Saccharomyces cerevisiae, Sequence Alignment, Transfection, Virus Replication, nef Gene Products, Human Immunodeficiency Virus
Virology
Date: Oct. 13, 1997
PubMed ID: 9344905
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