title: The FACT Complex Promotes Avian Leukosis Virus DNA Integration.
abstract: All retroviruses need to integrate a DNA copy of their genome into the host chromatin. Cellular proteins regulating and targeting lentiviral and gammaretroviral integration in infected cells have been discovered, but the factors that mediate alpharetroviral avian leukosis virus (ALV) integration are unknown. In this study, we have identified the FACT protein complex, which consists of SSRP1 and Spt16, as a principal cellular binding partner of ALV integrase (IN). Biochemical experiments with purified recombinant proteins show that SSRP1 and Spt16 are able to individually bind ALV IN, but only the FACT complex effectively stimulates ALV integration activityin vitroLikewise, in infected cells, the FACT complex promotes ALV integration activity, with proviral integration frequency varying directly with cellular expression levels of the FACT complex. An increase in 2-long-terminal-repeat (2-LTR) circles in the depleted FACT complex cell line indicates that this complex regulates the ALV life cycle at the level of integration. This regulation is shown to be specific to ALV, as disruption of the FACT complex did not inhibit either lentiviral or gammaretroviral integration in infected cells.IMPORTANCEThe majority of human gene therapy approaches utilize HIV-1- or murine leukemia virus (MLV)-based vectors, which preferentially integrate near genes and regulatory regions; thus, insertional mutagenesis is a substantial risk. In contrast, ALV integrates more randomly throughout the genome, which decreases the risks of deleterious integration. Understanding how ALV integration is regulated could facilitate the development of ALV-based vectors for use in human gene therapy. Here we show that the FACT complex directly binds and regulates ALV integration efficiencyin vitroand in infected cells.
authors.affiliation: Department of Biology, Johns Hopkins University, Baltimore, Maryland, USA.
authors.forename: Shelby
authors.initials: S
authors.lastname: Winans
authors.affiliation: Center for Retrovirus Research and Comprehensive Cancer Center, College of Pharmacy, The Ohio State University, Columbus, Ohio, USA.
authors.forename: Ross C
authors.initials: RC
authors.lastname: Larue
authors.affiliation: Center for Retrovirus Research and Comprehensive Cancer Center, College of Pharmacy, The Ohio State University, Columbus, Ohio, USA.
authors.forename: Carly M
authors.initials: CM
authors.lastname: Abraham
authors.affiliation: Center for Retrovirus Research and Comprehensive Cancer Center, College of Pharmacy, The Ohio State University, Columbus, Ohio, USA.
authors.forename: Nikolozi
authors.initials: N
authors.lastname: Shkriabai
authors.affiliation: Department of Biological Sciences, The University of Texas at Dallas, Richardson, Texas, USA.
authors.forename: Amelie
authors.initials: A
authors.lastname: Skopp
authors.affiliation: Department of Biological Sciences, The University of Texas at Dallas, Richardson, Texas, USA.
authors.forename: Duane
authors.initials: D
authors.lastname: Winkler
authors.affiliation: Center for Retrovirus Research and Comprehensive Cancer Center, College of Pharmacy, The Ohio State University, Columbus, Ohio, USA.
authors.forename: Mamuka
authors.initials: M
authors.lastname: Kvaratskhelia
authors.affiliation: Department of Biology, Johns Hopkins University, Baltimore, Maryland, USA KLB@jhu.edu.
authors.forename: Karen L
authors.initials: KL
authors.lastname: Beemon
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001354:Avian Leukosis Virus / Q000201:enzymology / Q000235:genetics*; D018797:Cell Cycle Proteins / Q000502:physiology*; D002642:Chick Embryo; D017124:Conserved Sequence; D004279:DNA, Viral / Q000502:physiology*; D004268:DNA-Binding Proteins / Q000502:physiology*; D057809:HEK293 Cells; D006609:High Mobility Group Proteins / Q000502:physiology*; D006801:Humans; D019426:Integrases / Q000502:physiology; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs; D014157:Transcription Factors / Q000502:physiology*; D035602:Transcriptional Elongation Factors / Q000502:physiology*; D016662:Virus Integration
chemical_list: D018797:Cell Cycle Proteins; D004279:DNA, Viral; D004268:DNA-Binding Proteins; D006609:High Mobility Group Proteins; C073537:SSRP1 protein, human; C489489:SUPT16H protein, human; D014157:Transcription Factors; D035602:Transcriptional Elongation Factors; D019426:Integrases
keywords: ALV; FACT; integration