title: Derivation and Characterization of a CD4-Independent, Non-CD4-Tropic Simian Immunodeficiency Virus.
abstract: CD4 tropism is conserved among all primate lentiviruses and likely contributes to viral pathogenesis by targeting cells that are critical for adaptive antiviral immune responses. Although CD4-independent variants of human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV) have been described that can utilize the coreceptor CCR5 or CXCR4 in the absence of CD4, these viruses typically retain their CD4 binding sites and still can interact with CD4. We describe the derivation of a novel CD4-independent variant of pathogenic SIVmac239, termed iMac239, that was used to derive an infectious R5-tropic SIV lacking a CD4 binding site. Of the seven mutations that differentiate iMac239 from wild-type SIVmac239, a single change (D178G) in the V1/V2 region was sufficient to confer CD4 independence in cell-cell fusion assays, although other mutations were required for replication competence. Like other CD4-independent viruses, iMac239 was highly neutralization sensitive, although mutations were identified that could confer CD4-independent infection without increasing its neutralization sensitivity. Strikingly, iMac239 retained the ability to replicate in cell lines and primary cells even when its CD4 binding site had been ablated by deletion of a highly conserved aspartic acid at position 385, which, for HIV-1, plays a critical role in CD4 binding. iMac239, with and without the D385 deletion, exhibited an expanded host range in primary rhesus peripheral blood mononuclear cells that included CCR5(+) CD8(+) T cells. As the first non-CD4-tropic SIV, iMac239-ΔD385 will afford the opportunity to directly assess the in vivo role of CD4 targeting on pathogenesis and host immune responses.
  
  
  
  CD4 tropism is an invariant feature of primate lentiviruses and likely plays a key role in pathogenesis by focusing viral infection onto cells that mediate adaptive immune responses and in protecting virions attached to cells from neutralizing antibodies. Although CD4-independent viruses are well described for HIV and SIV, these viruses characteristically retain their CD4 binding site and can engage CD4 if available. We derived a novel CD4-independent, CCR5-tropic variant of the pathogenic molecular clone SIVmac239, termed iMac239. The genetic determinants of iMac239's CD4 independence provide new insights into mechanisms that underlie this phenotype. This virus remained replication competent even after its CD4 binding site had been ablated by mutagenesis. As the first truly non-CD4-tropic SIV, lacking the capacity to interact with CD4, iMac239 will provide the unique opportunity to evaluate SIV pathogenesis and host immune responses in the absence of the immunomodulatory effects of CD4(+) T cell targeting and infection.
authors.affiliation: Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, USA.
authors.forename: Adrienne E
authors.initials: AE
authors.lastname: Swanstrom
authors.affiliation: Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, USA.
authors.forename: Beth
authors.initials: B
authors.lastname: Haggarty
authors.affiliation: Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, USA.
authors.forename: Andrea P O
authors.initials: APO
authors.lastname: Jordan
authors.affiliation: Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, USA.
authors.forename: Josephine
authors.initials: J
authors.lastname: Romano
authors.affiliation: Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, USA.
authors.forename: George J
authors.initials: GJ
authors.lastname: Leslie
authors.affiliation: Tulane National Primate Research Center, Covington, and Department of Tropical Medicine, Tulane University, New Orleans, Louisiana, USA.
authors.forename: Pyone P
authors.initials: PP
authors.lastname: Aye
authors.affiliation: Tulane National Primate Research Center, Covington, and Department of Tropical Medicine, Tulane University, New Orleans, Louisiana, USA.
authors.forename: Preston A
authors.initials: PA
authors.lastname: Marx
authors.affiliation: Tulane National Primate Research Center, Covington, and Department of Tropical Medicine, Tulane University, New Orleans, Louisiana, USA.
authors.forename: Andrew A
authors.initials: AA
authors.lastname: Lackner
authors.affiliation: AIDS and Cancer Virus Program, Leidos Biomedical Research, Inc., Frederick National Laboratory for Cancer Research, Frederick, Maryland, USA.
authors.forename: Gregory Q
authors.initials: GQ
authors.lastname: Del Prete
authors.affiliation: Department of Pediatrics, Tulane University School of Medicine, New Orleans, Louisiana, USA.
authors.forename: James E
authors.initials: JE
authors.lastname: Robinson
authors.affiliation: Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, USA.
authors.forename: Michael R
authors.initials: MR
authors.lastname: Betts
authors.affiliation: Department of Surgery, Duke University Medical Center, Durham, North Carolina, USA.
authors.forename: David C
authors.initials: DC
authors.lastname: Montefiori
authors.affiliation: Department of Surgery, Duke University Medical Center, Durham, North Carolina, USA.
authors.forename: Celia C
authors.initials: CC
authors.lastname: LaBranche
authors.affiliation: Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, USA hoxie@mail.med.upenn.edu.
authors.forename: James A
authors.identifier: http://orcid.org/0000-0002-6257-4902
authors.initials: JA
authors.lastname: Hoxie
mesh_terms: D000818:Animals; D057134:Antibodies, Neutralizing / Q000276:immunology; D001665:Binding Sites; D015704:CD4 Antigens / Q000276:immunology / Q000378:metabolism*; D018414:CD8-Positive T-Lymphocytes / Q000821:virology; D002460:Cell Line; D006801:Humans; D007963:Leukocytes, Mononuclear / Q000821:virology; D008253:Macaca mulatta; D016296:Mutagenesis; D019713:Receptors, CCR5 / Q000276:immunology / Q000378:metabolism; D019718:Receptors, CXCR4 / Q000276:immunology / Q000378:metabolism; D015302:Simian Immunodeficiency Virus / Q000235:genetics* / Q000276:immunology / Q000472:pathogenicity / Q000502:physiology*; D014759:Viral Envelope Proteins / Q000235:genetics; D056189:Viral Tropism; D053585:Virus Attachment; D014779:Virus Replication / Q000235:genetics
chemical_list: D057134:Antibodies, Neutralizing; D015704:CD4 Antigens; D019713:Receptors, CCR5; D019718:Receptors, CXCR4; D014759:Viral Envelope Proteins