title: Modulation of GSK-3β activity in Venezuelan equine encephalitis virus infection.
abstract: Alphaviruses, including Venezuelan Equine Encephalitis Virus (VEEV), cause disease in both equine and humans that exhibit overt encephalitis in a significant percentage of cases. Features of the host immune response and tissue-specific responses may contribute to fatal outcomes as well as the development of encephalitis. It has previously been shown that VEEV infection of mice induces transcription of pro-inflammatory cytokines genes (e.g., IFN-γ, IL-6, IL-12, iNOS and TNF-α) within 6 h. GSK-3β is a host protein that is known to modulate pro-inflammatory gene expression and has been a therapeutic target in neurodegenerative disorders such as Alzheimer's. Hence inhibition of GSK-3β in the context of encephalitic viral infections has been useful in a neuroprotective capacity. Small molecule GSK-3β inhibitors and GSK-3β siRNA experiments indicated that GSK-3β was important for VEEV replication. Thirty-eight second generation BIO derivatives were tested and BIOder was found to be the most potent inhibitor, with an IC(50) of ∼0.5 µM and a CC(50) of >100 µM. BIOder was a more potent inhibitor of GSK-3β than BIO, as demonstrated through in vitro kinase assays from uninfected and infected cells. Size exclusion chromatography experiments demonstrated that GSK-3β is found in three distinct complexes in VEEV infected cells, whereas GSK-3β is only present in one complex in uninfected cells. Cells treated with BIOder demonstrated an increase in the anti-apoptotic gene, survivin, and a decrease in the pro-apoptotic gene, BID, suggesting that modulation of pro- and anti-apoptotic genes contributes to the protective effect of BIOder treatment. Finally, BIOder partially protected mice from VEEV induced mortality. Our studies demonstrate the utility of GSK-3β inhibitors for modulating VEEV infection.
authors.affiliation: National Center for Biodefense and Infectious Diseases, George Mason University, Manassas, Virginia, United States of America.
authors.forename: Kylene
authors.initials: K
authors.lastname: Kehn-Hall
authors.forename: Aarthi
authors.initials: A
authors.lastname: Narayanan
authors.forename: Lindsay
authors.initials: L
authors.lastname: Lundberg
authors.forename: Gavin
authors.initials: G
authors.lastname: Sampey
authors.forename: Chelsea
authors.initials: C
authors.lastname: Pinkham
authors.forename: Irene
authors.initials: I
authors.lastname: Guendel
authors.forename: Rachel
authors.initials: R
authors.lastname: Van Duyne
authors.forename: Svetlana
authors.initials: S
authors.lastname: Senina
authors.forename: Kimberly L
authors.initials: KL
authors.lastname: Schultz
authors.forename: Eric
authors.initials: E
authors.lastname: Stavale
authors.forename: M Javad
authors.initials: MJ
authors.lastname: Aman
authors.forename: Charles
authors.initials: C
authors.lastname: Bailey
authors.forename: Fatah
authors.initials: F
authors.lastname: Kashanchi
mesh_terms: D000818:Animals; D051023:BH3 Interacting Domain Death Agonist Protein / Q000032:analysis; D004666:Encephalitis Virus, Venezuelan Equine / Q000187:drug effects*; D004685:Encephalomyelitis, Venezuelan Equine / Q000188:drug therapy* / Q000401:mortality; D004791:Enzyme Inhibitors / Q000627:therapeutic use*; D005260:Female; D038362:Glycogen Synthase Kinase 3 / Q000037:antagonists & inhibitors*; D000071679:Glycogen Synthase Kinase 3 beta; D051034:Inhibitor of Apoptosis Proteins / Q000032:analysis; D051379:Mice; D008809:Mice, Inbred C3H; D012097:Repressor Proteins / Q000032:analysis; D000077022:Survivin; D014779:Virus Replication / Q000187:drug effects
chemical_list: D051023:BH3 Interacting Domain Death Agonist Protein; C494815:Bid protein, mouse; C117749:Birc5 protein, mouse; D004791:Enzyme Inhibitors; D051034:Inhibitor of Apoptosis Proteins; D012097:Repressor Proteins; D000077022:Survivin; C000605506:GSK3B protein, human; D000071679:Glycogen Synthase Kinase 3 beta; C000605507:Gsk3b protein, mouse; D038362:Glycogen Synthase Kinase 3