title: Negative Feedback Regulation of HIV-1 by Gene Editing Strategy.
abstract: The CRISPR/Cas9 gene editing method is comprised of the guide RNA (gRNA) to target a specific DNA sequence for cleavage and the Cas9 endonuclease for introducing breaks in the double-stranded DNA identified by the gRNA. Co-expression of both a multiplex of HIV-1-specific gRNAs and Cas9 in cells results in the modification and/or excision of the segment of viral DNA, leading to replication-defective virus. In this study, we have personalized the activity of CRISPR/Cas9 by placing the gene encoding Cas9 under the control of a minimal promoter of HIV-1 that is activated by the HIV-1 Tat protein. We demonstrate that functional activation of CRISPR/Cas9 by Tat during the course of viral infection excises the designated segment of the integrated viral DNA and consequently suppresses viral expression. This strategy was also used in a latently infected CD4+ T-cell model after treatment with a variety of HIV-1 stimulating agents including PMA and TSA. Controlled expression of Cas9 by Tat offers a new strategy for safe implementation of the Cas9 technology for ablation of HIV-1 at a very early stage of HIV-1 replication during the course of the acute phase of infection and the reactivation of silent proviral DNA in latently infected cells.
authors.affiliation: Department of Neuroscience Center for Neurovirology, Lewis Katz School of Medicine at Temple University, 3500 N. Broad Street, 7th Floor Philadelphia, PA 19140, USA.
authors.forename: Rafal
authors.initials: R
authors.lastname: Kaminski
authors.affiliation: Department of Neuroscience Center for Neurovirology, Lewis Katz School of Medicine at Temple University, 3500 N. Broad Street, 7th Floor Philadelphia, PA 19140, USA.
authors.forename: Yilan
authors.initials: Y
authors.lastname: Chen
authors.affiliation: Department of Neuroscience Center for Neurovirology, Lewis Katz School of Medicine at Temple University, 3500 N. Broad Street, 7th Floor Philadelphia, PA 19140, USA.
authors.forename: Julian
authors.initials: J
authors.lastname: Salkind
authors.affiliation: Department of Neuroscience Center for Neurovirology, Lewis Katz School of Medicine at Temple University, 3500 N. Broad Street, 7th Floor Philadelphia, PA 19140, USA.
authors.forename: Ramona
authors.initials: R
authors.lastname: Bella
authors.affiliation: Department of Radiology University of Pittsburgh School of Medicine Pittsburgh, PA 15219, USA.
authors.forename: Won-Bin
authors.initials: WB
authors.lastname: Young
authors.affiliation: Department of Neuroscience Center for Neurovirology, Lewis Katz School of Medicine at Temple University, 3500 N. Broad Street, 7th Floor Philadelphia, PA 19140, USA.|Microbiology and Clinical Microbiology, Department of Biomedical, Surgical and Dental Sciences, University of Milan, Milan, Italy.
authors.forename: Pasquale
authors.initials: P
authors.lastname: Ferrante
authors.affiliation: Department of Molecular Biology and Microbiology, Case Western Reserve University, Cleveland, OH 44106, USA.
authors.forename: Jonathan
authors.initials: J
authors.lastname: Karn
authors.affiliation: Excision Biotherapeutics, Inc., 3624 Market Street, #514, Philadelphia, PA 19104, USA.
authors.forename: Thomas
authors.initials: T
authors.lastname: Malcolm
authors.affiliation: Department of Neuroscience Center for Neurovirology, Lewis Katz School of Medicine at Temple University, 3500 N. Broad Street, 7th Floor Philadelphia, PA 19140, USA.
authors.forename: Wenhui
authors.initials: W
authors.lastname: Hu
authors.affiliation: Department of Neuroscience Center for Neurovirology, Lewis Katz School of Medicine at Temple University, 3500 N. Broad Street, 7th Floor Philadelphia, PA 19140, USA.
authors.forename: Kamel
authors.initials: K
authors.lastname: Khalili
mesh_terms: D020413:3' Untranslated Regions; D020121:5' Untranslated Regions; D001483:Base Sequence; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology / Q000821:virology; D064112:Clustered Regularly Interspaced Short Palindromic Repeats; D000072669:Gene Editing; D015967:Gene Expression Regulation, Viral; D005814:Genes, Viral; D016325:HIV Long Terminal Repeat; D015497:HIV-1 / Q000235:genetics* / Q000502:physiology; D006801:Humans; D019169:Jurkat Cells; D011401:Promoter Regions, Genetic; D012689:Sequence Homology, Nucleic Acid; D014779:Virus Replication
chemical_list: D020413:3' Untranslated Regions; D020121:5' Untranslated Regions