Secretion modification region-derived peptide disrupts HIV-1 Nef's interaction with mortalin and blocks virus and Nef exosome release.
Nef is secreted from infected cells in exosomes and is found in abundance in the sera of HIV-infected individuals. Secreted exosomal Nef (exNef) induces apoptosis in uninfected CD4⁺ T cells and may be a key component of HIV pathogenesis. The exosomal pathway has been implicated in HIV-1 virus release, suggesting a possible link between these two viral processes. However, the underlying mechanisms and cellular components of exNef secretion have not been elucidated. We have previously described a Nef motif, the secretion modification region (SMR; amino acids 66 to 70), that is required for exNef secretion. In silico modeling data suggest that this motif can form a putative binding pocket. We hypothesized that the Nef SMR binds a cellular protein involved in protein trafficking and that inhibition of this interaction would abrogate exNef secretion. By using tandem mass spectrometry and coimmunoprecipitation with a novel SMR-based peptide (SMRwt) that blocks exNef secretion and HIV-1 virus release, we identified mortalin as an SMR-specific cellular protein. A second set of coimmunoprecipitation experiments with full-length Nef confirmed that mortalin interacts with Nef via Nef's SMR motif and that this interaction is disrupted by the SMRwt peptide. Overexpression and microRNA knockdown of mortalin revealed a positive correlation between exNef secretion levels and mortalin protein expression. Using antibody inhibition we demonstrated that the Nef/mortalin interaction is necessary for exNef secretion. Taken together, this work constitutes a significant step in understanding the underlying mechanism of exNef secretion, identifies a novel host-pathogen interaction, and introduces an HIV-derived peptide with antiviral properties.
Department of Microbiology, Biochemistry, and Immunology, Morehouse School of Medicine, Atlanta, Georgia, USA.
Martin N
MN
Shelton
Ming-Bo
MB
Huang
Syed A
SA
Ali
Michael D
MD
Powell
Vincent C
VC
Bond
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D002460:Cell Line; D055354:Exosomes / Q000235:genetics / Q000378:metabolism; D005110:Extracellular Space / Q000378:metabolism*; D015658:HIV Infections / Q000235:genetics / Q000378:metabolism* / Q000821:virology; D015497:HIV-1 / Q000235:genetics / Q000378:metabolism; D018840:HSP70 Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D021381:Protein Transport; D054311:nef Gene Products, Human Immunodeficiency Virus / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
D018840:HSP70 Heat-Shock Proteins; D010455:Peptides; C080352:mortalin; D054311:nef Gene Products, Human Immunodeficiency Virus