HIV-1 Nef, a protein important for the development of AIDS, has well-characterized effects on host membrane trafficking and receptor downregulation. By an unidentified mechanism, Nef increases the intrinsic infectivity of HIV-1 virions in a host-cell-dependent manner. Here we identify the host transmembrane protein SERINC5, and to a lesser extent SERINC3, as a potent inhibitor of HIV-1 particle infectivity that is counteracted by Nef. SERINC5 localizes to the plasma membrane, where it is efficiently incorporated into budding HIV-1 virions and impairs subsequent virion penetration of susceptible target cells. Nef redirects SERINC5 to a Rab7-positive endosomal compartment and thereby excludes it from HIV-1 particles. The ability to counteract SERINC5 was conserved in Nef encoded by diverse primate immunodeficiency viruses, as well as in the structurally unrelated glycosylated Gag from murine leukaemia virus. These examples of functional conservation and convergent evolution emphasize the fundamental importance of SERINC5 as a potent anti-retroviral factor.
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We thank the Centre for AIDS Reagents, NIBSC, and NIH AIDS Research and Reference Reagent Program, Division of AIDS, for cell lines, plasmids and antibodies. We thank V. Adami and the CIBIO high-throughput screening and the Advanced Imaging facilities staff for technical assistance, G. De Silvestro, G. Mattiuzzo, C. Reinhard and L. Conti for reagents, G. Melikian, S. Basmaciogullari, P. Cherepanov, O. Fackler, N. Segata, F. Demichelis, A. Marcello, T. Fedrizzi and A. Helander for critical discussions. This work was supported by the Biotechnology Program of University of Trento, FP7 Marie Curie Career Integration grant number 322130 and Caritro ‘Ricerca Biomedica’ grant number 2013.0248 to M.P., National Institute of Health grant DP1DA034990 to J.L. and European Research Council grant 249968 to S.E.A.
The authors declare no competing financial interests.
Extended data figures and tables
a, Mapping of the INDELS in the genomic locus spanning SERINC5 exon 2 in JTAg cell clonal populations from Fig. 2a. b, Infectivity of HIV-1 from JTAg cells stably transduced with lentiCRISPR targeting GFP or SERINC5 in three different exons (n = 4, experiment replicated twice). c, Relative expression of SERINC5 in primary cells and in cell lines measured by qPCR normalized by expression of ACTB (n = 3). d, Infectivity of HIV-1 from the indicated cell lines expressing SERINC5 (n = 4, experiments were replicated twice). Mean ± s.d., unpaired two-tailed t-test, ***P < 0.001 e, Expression levels of the five SERINC genes in JTAg cells obtained from RNA-seq.
Extended Data Figure 2 Nef and glycoGag expression result in relocalization of SERINC5 to an endosomal compartment and prevent its incorporation into virions.
a, Single round Nef-defective NL4-3 produced by cotransfection of HEK293T cells with plasmids expressing Nef proteins or the empty vector control, and PBJ6-SERINC5–HA: immunoblotting of virions and cell lysates from producer cells. b, Immunofluorescence staining of JTAg cells transfected to express SERINC5–GFP, Nef–HA from HIV-1 isolate 97ZA012 (clade C), from SIVmac239, HA–glycoGag or an empty vector control. Scale bar, 10 μm.
a, Immunodetection of Cre-recombinase (38 kDa) and p24 in HIV-1 particles. b, Effect of 1 μM AZT or 100 nM T20 on Cre-delivery and virus infectivity (TU, transducing units). c, Immunoblotting of HIV-1 virus particles produced from HEK293T expressing increasing levels of SERINC5–HA. d, Effect of SERINC5 on virus fusion measured with BLAM assay T20 served as a negative control. (n = 4, experiment replicated twice). e, Cre delivery by EBOV-GP pseudotyped HIV-1 particles. f, Inhibition of Cre delivery and counteraction by Nef on HIV-1 from HEK293T expressing SERINC5. Mean ± s.d., n = 4, unpaired two-tailed t-test, *P < 0.05, **P < 0.01, ***P < 0.001. Scale bar, 100 μm.
Extended Data Figure 4 SERINC3 and SERINC5 expression is not induced by interferon nor LPS treatments.
a–d, Relative gene expression levels of SERINC3, SERINC5 and CXCL10 in response to treatment with IFN-β and LPS in Jurkat (a), monocyte-derived dendritic cells from two donors (MDDC, b), CD4+ primary T cells unstimulated (c) or stimulated with PHA (d) from two donors. Expression of the housekeeping gene OAZ1 was used as a normalization control. Mean ± s.d., n = 3.
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Rosa, A., Chande, A., Ziglio, S. et al. HIV-1 Nef promotes infection by excluding SERINC5 from virion incorporation. Nature 526, 212–217 (2015). https://doi.org/10.1038/nature15399
Sensitivity to monoclonal antibody 447-52D and an open env trimer conformation correlate poorly with inhibition of HIV-1 infectivity by SERINC5
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