title: Selective Inhibitor of Nuclear Export (SINE) Compounds Alter New World Alphavirus Capsid Localization and Reduce Viral Replication in Mammalian Cells.
abstract: The capsid structural protein of the New World alphavirus, Venezuelan equine encephalitis virus (VEEV), interacts with the host nuclear transport proteins importin α/β1 and CRM1. Novel selective inhibitor of nuclear export (SINE) compounds, KPT-185, KPT-335 (verdinexor), and KPT-350, target the host's primary nuclear export protein, CRM1, in a manner similar to the archetypical inhibitor Leptomycin B. One major limitation of Leptomycin B is its irreversible binding to CRM1; which SINE compounds alleviate because they are slowly reversible. Chemically inhibiting CRM1 with these compounds enhanced capsid localization to the nucleus compared to the inactive compound KPT-301, as indicated by immunofluorescent confocal microscopy. Differences in extracellular versus intracellular viral RNA, as well as decreased capsid in cell free supernatants, indicated the inhibitors affected viral assembly, which led to a decrease in viral titers. The decrease in viral replication was confirmed using a luciferase-tagged virus and through plaque assays. SINE compounds had no effect on VEEV TC83_Cm, which encodes a mutated form of capsid that is unable to enter the nucleus. Serially passaging VEEV in the presence of KPT-185 resulted in mutations within the nuclear localization and nuclear export signals of capsid. Finally, SINE compound treatment also reduced the viral titers of the related eastern and western equine encephalitis viruses, suggesting that CRM1 maintains a common interaction with capsid proteins across the New World alphavirus genus.
authors.affiliation: National Center for Biodefense and Infectious Diseases, School of Systems Biology, George Mason University, Manassas, Virginia, United States of America.
authors.forename: Lindsay
authors.initials: L
authors.lastname: Lundberg
authors.affiliation: National Center for Biodefense and Infectious Diseases, School of Systems Biology, George Mason University, Manassas, Virginia, United States of America.
authors.forename: Chelsea
authors.initials: C
authors.lastname: Pinkham
authors.affiliation: National Center for Biodefense and Infectious Diseases, School of Systems Biology, George Mason University, Manassas, Virginia, United States of America.
authors.forename: Cynthia
authors.initials: C
authors.lastname: de la Fuente
authors.affiliation: National Center for Biodefense and Infectious Diseases, School of Systems Biology, George Mason University, Manassas, Virginia, United States of America.
authors.forename: Ashwini
authors.initials: A
authors.lastname: Brahms
authors.affiliation: National Center for Biodefense and Infectious Diseases, School of Systems Biology, George Mason University, Manassas, Virginia, United States of America.
authors.forename: Nazly
authors.initials: N
authors.lastname: Shafagati
authors.affiliation: Nuclear Signaling Laboratory, Department of Biochemistry and Molecular Biology, Monash University, Clayton, Victoria, Australia.
authors.forename: Kylie M
authors.initials: KM
authors.lastname: Wagstaff
authors.affiliation: Nuclear Signaling Laboratory, Department of Biochemistry and Molecular Biology, Monash University, Clayton, Victoria, Australia.
authors.forename: David A
authors.initials: DA
authors.lastname: Jans
authors.affiliation: Karyopharm Therapeutics Inc., Massachusetts, United States of America.
authors.forename: Sharon
authors.initials: S
authors.lastname: Tamir
authors.affiliation: National Center for Biodefense and Infectious Diseases, School of Systems Biology, George Mason University, Manassas, Virginia, United States of America.
authors.forename: Kylene
authors.initials: K
authors.lastname: Kehn-Hall
mesh_terms: D021581:Active Transport, Cell Nucleus / Q000187:drug effects; D000524:Alphavirus / Q000187:drug effects* / Q000235:genetics / Q000502:physiology; D018354:Alphavirus Infections / Q000821:virology*; D000818:Animals; D000998:Antiviral Agents / Q000494:pharmacology*; D036022:Capsid Proteins / Q000235:genetics / Q000378:metabolism*; D002467:Cell Nucleus / Q000821:virology; D006801:Humans; D028884:Karyopherins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D018160:Receptors, Cytoplasmic and Nuclear / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D019065:Virus Assembly / Q000187:drug effects; D014779:Virus Replication / Q000187:drug effects*; D000097606:Exportin 1 Protein
chemical_list: D000998:Antiviral Agents; D036022:Capsid Proteins; D028884:Karyopherins; D018160:Receptors, Cytoplasmic and Nuclear; D000097606:Exportin 1 Protein