title: Single-Cell Characterization of Viral Translation-Competent Reservoirs in HIV-Infected Individuals.
abstract: HIV cure efforts are hampered by limited characterization of the cells supporting HIV replication in vivo and inadequate methods for quantifying the latent viral reservoir in patients receiving antiretroviral therapy. We combine fluorescent in situ RNA hybridization with detection of HIV protein and flow cytometry, enabling detection of 0.5-1 gag-pol mRNA(+)/Gag protein(+)-infected cells per million. In the peripheral blood of untreated persons, active HIV replication correlated with viremia and occurred in CD4 T cells expressing T follicular helper cell markers and inhibitory co-receptors. In virally suppressed subjects, the approach identified latently infected cells capable of producing HIV mRNA and protein after stimulation with PMA/ionomycin and latency-reversing agents (LRAs). While ingenol-induced reactivation mirrored the effector and central/transitional memory CD4 T cell contribution to the pool of integrated HIV DNA, bryostatin-induced reactivation occurred predominantly in cells expressing effector memory markers. This indicates that CD4 T cell differentiation status differentially affects LRA effectiveness.
authors.affiliation: Research Centre of the Centre Hospitalier de l'Université de Montréal (CRCHUM) and Université de Montréal, Montreal, QC H2X 0A9, Canada; Center for HIV/AIDS Vaccine Immunology and Immunogen Discovery (CHAVI-ID), La Jolla, CA 92037, USA.
authors.forename: Amy E
authors.initials: AE
authors.lastname: Baxter
authors.affiliation: Research Centre of the Centre Hospitalier de l'Université de Montréal (CRCHUM) and Université de Montréal, Montreal, QC H2X 0A9, Canada; Center for HIV/AIDS Vaccine Immunology and Immunogen Discovery (CHAVI-ID), La Jolla, CA 92037, USA.
authors.forename: Julia
authors.initials: J
authors.lastname: Niessl
authors.affiliation: Research Centre of the Centre Hospitalier de l'Université de Montréal (CRCHUM) and Université de Montréal, Montreal, QC H2X 0A9, Canada.
authors.forename: Rémi
authors.initials: R
authors.lastname: Fromentin
authors.affiliation: Research Centre of the Centre Hospitalier de l'Université de Montréal (CRCHUM) and Université de Montréal, Montreal, QC H2X 0A9, Canada.
authors.forename: Jonathan
authors.initials: J
authors.lastname: Richard
authors.affiliation: Center for HIV/AIDS Vaccine Immunology and Immunogen Discovery (CHAVI-ID), La Jolla, CA 92037, USA; Ragon Institute of Massachusetts General Hospital, Massachusetts Institute of Technology and Harvard University, Cambridge, MA 02139, USA.
authors.forename: Filippos
authors.initials: F
authors.lastname: Porichis
authors.affiliation: Research Centre of the Centre Hospitalier de l'Université de Montréal (CRCHUM) and Université de Montréal, Montreal, QC H2X 0A9, Canada.
authors.forename: Roxanne
authors.initials: R
authors.lastname: Charlebois
authors.affiliation: Research Centre of the Centre Hospitalier de l'Université de Montréal (CRCHUM) and Université de Montréal, Montreal, QC H2X 0A9, Canada.
authors.forename: Marta
authors.initials: M
authors.lastname: Massanella
authors.affiliation: Research Centre of the Centre Hospitalier de l'Université de Montréal (CRCHUM) and Université de Montréal, Montreal, QC H2X 0A9, Canada.
authors.forename: Nathalie
authors.initials: N
authors.lastname: Brassard
authors.affiliation: Research Centre of the Centre Hospitalier de l'Université de Montréal (CRCHUM) and Université de Montréal, Montreal, QC H2X 0A9, Canada; Department of Microbiology and Immunology, McGill University, Montreal, QC H3A 2B4, Canada.
authors.forename: Nirmin
authors.initials: N
authors.lastname: Alsahafi
authors.affiliation: Research Centre of the Centre Hospitalier de l'Université de Montréal (CRCHUM) and Université de Montréal, Montreal, QC H2X 0A9, Canada.
authors.forename: Gloria-Gabrielle
authors.initials: GG
authors.lastname: Delgado
authors.affiliation: Chronic Viral Illnesses Service and Division of Hematology, McGill University Health Centre, Montreal, QC H4A 3J1, Canada.
authors.forename: Jean-Pierre
authors.initials: JP
authors.lastname: Routy
authors.affiliation: Center for HIV/AIDS Vaccine Immunology and Immunogen Discovery (CHAVI-ID), La Jolla, CA 92037, USA; Ragon Institute of Massachusetts General Hospital, Massachusetts Institute of Technology and Harvard University, Cambridge, MA 02139, USA; Howard Hughes Medical Institute, Chevy Chase, MD 20815, USA.
authors.forename: Bruce D
authors.initials: BD
authors.lastname: Walker
authors.affiliation: Research Centre of the Centre Hospitalier de l'Université de Montréal (CRCHUM) and Université de Montréal, Montreal, QC H2X 0A9, Canada; Department of Microbiology and Immunology, McGill University, Montreal, QC H3A 2B4, Canada.
authors.forename: Andrés
authors.initials: A
authors.lastname: Finzi
authors.affiliation: Research Centre of the Centre Hospitalier de l'Université de Montréal (CRCHUM) and Université de Montréal, Montreal, QC H2X 0A9, Canada.
authors.forename: Nicolas
authors.initials: N
authors.lastname: Chomont
authors.affiliation: Research Centre of the Centre Hospitalier de l'Université de Montréal (CRCHUM) and Université de Montréal, Montreal, QC H2X 0A9, Canada; Center for HIV/AIDS Vaccine Immunology and Immunogen Discovery (CHAVI-ID), La Jolla, CA 92037, USA; Ragon Institute of Massachusetts General Hospital, Massachusetts Institute of Technology and Harvard University, Cambridge, MA 02139, USA. Electronic address: daniel.kaufmann@umontreal.ca.
authors.forename: Daniel E
authors.initials: DE
authors.lastname: Kaufmann
mesh_terms: D015496:CD4-Positive T-Lymphocytes / Q000821:virology*; D002478:Cells, Cultured; D005434:Flow Cytometry; D015658:HIV Infections / Q000188:drug therapy* / Q000821:virology*; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D012333:RNA, Messenger / Q000032:analysis*; D059010:Single-Cell Analysis; D000072230:Sustained Virologic Response; D054301:gag Gene Products, Human Immunodeficiency Virus / Q000032:analysis*; D054302:pol Gene Products, Human Immunodeficiency Virus / Q000032:analysis*
chemical_list: D012333:RNA, Messenger; D054301:gag Gene Products, Human Immunodeficiency Virus; D054302:pol Gene Products, Human Immunodeficiency Virus