Expression of an IKKgamma splice variant determines IRF3 and canonical NF-kappaB pathway utilization in ssRNA virus infection.
Single stranded RNA (ssRNA) virus infection activates the retinoic acid inducible gene I (RIG-I)- mitochondrial antiviral signaling (MAVS) complex, a complex that coordinates the host innate immune response via the NF-kappaB and IRF3 pathways. Recent work has shown that the IkappaB kinase (IKK)gamma scaffolding protein is the final common adapter protein required by RIG-I.MAVS to activate divergent rate-limiting kinases downstream controlling the NF-kappaB and IRF3 pathways. Previously we discovered a ubiquitous IKKgamma splice-variant, IKKgammaDelta, that exhibits distinct signaling properties.


RESULTS
We examined the regulation and function of IKKgamma splice forms in response to ssRNA virus infection, a condition that preferentially induces full length IKKgamma-WT mRNA expression. In IKKgammaDelta-expressing cells, we found increased viral translation and cytopathic effect compared to those expressing full length IKKgamma-WT. IKKgammaDelta fails to support viral-induced IRF3 activation in response to ssRNA infections; consequently type I IFN production and the induction of anti-viral interferon stimulated genes (ISGs) are significantly attenuated. By contrast, ectopic RIG-I.MAVS or TNFalpha-induced canonical NF-kappaB activation is preserved in IKKgammaDelta expressing cells. Increasing relative levels of IKKgamma-WT to IKKgammaDelta (while keeping total IKKgamma constant) results in increased type I IFN expression. Conversely, overexpressing IKKgammaDelta (in a background of constant IKKgamma-WT expression) shows IKKgammaDelta functions as a dominant-negative IRF3 signaling inhibitor. IKKgammaDelta binds both IKK-alpha and beta, but not TANK and IKKepsilon, indicating that exon 5 encodes an essential TANK binding domain. Finally, IKKgammaDelta displaces IKKgammaWT from MAVS explaining its domainant negative effect.


CONCLUSIONS
Relative endogenous IKKgammaDelta expression affects cellular selection of inflammatory/anti-viral pathway responses to ssRNA viral infection.
Liu
Ping
P
Department of Medicine, University of Texas Medical Branch (UTMB), Galveston, Texas, United States of America.
Lu
Muping
M
Tian
Bing
B
Li
Kui
K
Garofalo
Roberto P
RP
Prusak
Deborah
D
Wood
Thomas G
TG
Brasier
Allan R
AR
D017398:Alternative Splicing; D000818:Animals; D045744:Cell Line, Tumor; D005347:Fibroblasts / Q000378:metabolism; D005799:Genes, Dominant; D006801:Humans; D051550:I-kappa B Kinase / Q000096:biosynthesis* / Q000737:chemistry*; D050838:Interferon Regulatory Factor-3 / Q000378:metabolism*; D051379:Mice; D008856:Microscopy, Fluorescence / Q000379:methods; D008954:Models, Biological; D016328:NF-kappa B / Q000378:metabolism*; D012327:RNA Virus Infections / Q000378:metabolism; D012328:RNA Viruses / Q000378:metabolism*; D015398:Signal Transduction
C494232:IRF3 protein, human; D050838:Interferon Regulatory Factor-3; C494233:Irf3 protein, mouse; D016328:NF-kappa B; D051550:I-kappa B Kinase