Interaction of HIV-1 Nef with the Cellular Dileucine-Based Sorting Pathway is Required for CD4 Down-Regulation and Optimal Viral Infectivity

• Published in: Proceedings of the National Academy of Sciences - PNAS Vol. 95; no. 19; pp. 11229 - 11234
• Main Authors: Craig, Heather M., Pandori, Mark W., Guatelli, John C.
• Format: Journal Article
• Language: English
• Published: United States National Academy of Sciences of the United States of America 15-09-1998; National Acad Sciences; National Academy of Sciences; The National Academy of Sciences
• Subjects: Biochemistry; Biological Sciences; CD4 Antigens - immunology; Cell Line; Cell lines; Cellular biology; Down regulation; Down-Regulation - physiology; Endocytosis; Endocytosis - physiology; Flow Cytometry; Gene Products, nef - chemistry; Gene Products, nef - physiology; HIV; HIV 1; HIV-1 - chemistry; Human immunodeficiency virus; Human immunodeficiency virus 1; Human immunodeficiency virus 2; Membrane proteins; Membrane Proteins - chemistry; Mutation; nef Gene Products, Human Immunodeficiency Virus; Plasmids; Proteins; Signal Transduction - physiology; Simian immunodeficiency virus; T lymphocytes; Transfection; Transfection - genetics; Transmembrane proteins; Viral Proteins - metabolism
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.95.19.11229

The HIV-1 Nef protein is important for pathogenesis, enhances viral infectivity, and regulates the sorting of at least two cellular transmembrane proteins, CD4 and major histocompatibility complex (MHC) class I. Although several lines of evidence support the hypothesis that the Nef protein interacts directly with the cellular protein sorting machinery, the sorting signal in HIV-1 Nef has not been identified. By using a competition assay that functionally discriminates between dileucine-based and tyrosine-based sorting signals, we have categorized the motif through which Nef interacts with the sorting machinery as dileucine-based. Inspection of diverse Nef proteins from HIV-1, HIV-2, and simian immunodeficiency virus revealed a well-conserved sequence in the central region of the C-terminal, solvent-exposed loop of Nef (E/DXXXLφ ) that conforms to the consensus sequence of the dileucine-based sorting motifs found in cellular transmembrane proteins. This sequence in NefNL4-3, ENTSLL, functioned as an endocytosis signal when appended to the cytoplasmic tail of a heterologous protein. The leucine residues in this motif were required for the interaction of full-length Nef with the dileucine-based sorting pathway and were required for Nef-mediated down-regulation of CD4. These leucine residues were also required for optimal viral infectivity. These data indicate that a dileucine-based sorting signal in Nef is utilized to address the cellular sorting machinery. The data also suggest that an influence on the distribution of cellular transmembrane proteins may mechanistically unite two previously distinct properties of Nef: down-regulation of CD4 and enhancement of viral infectivity.