Department of Medicine, University of Texas Medical Branch (UTMB), Galveston, Texas, United States of America.
PLoS One. 2009 Nov 26;4(11):e8079. doi: 10.1371/journal.pone.0008079.
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.
METHODOLOGY/PRINCIPAL FINDINGS: 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/SIGNIFICANCE: Relative endogenous IKKgammaDelta expression affects cellular selection of inflammatory/anti-viral pathway responses to ssRNA viral infection.
单链 RNA（ssRNA）病毒感染激活了视黄酸诱导基因 I（RIG-I）-线粒体抗病毒信号（MAVS）复合物，该复合物通过 NF-κB 和 IRF3 途径协调宿主固有免疫反应。最近的工作表明，IkappaB 激酶（IKK）γ支架蛋白是 RIG-I.MAVS 激活控制 NF-κB 和 IRF3 途径的不同限速激酶所必需的最终共同衔接蛋白。之前我们发现了一种普遍存在的 IKKγ剪接变异体 IKKγΔ，它表现出不同的信号特性。
方法/主要发现：我们研究了 IKKγ剪接形式在 ssRNA 病毒感染时的调节和功能，这种情况优先诱导全长 IKKγ-WT mRNA 的表达。在表达 IKKγΔ的细胞中，与表达全长 IKKγ-WT 的细胞相比，病毒翻译和细胞病变效应增加。IKKγΔ不能支持 ssRNA 感染诱导的病毒诱导的 IRF3 激活；因此，I 型 IFN 产生和抗病毒干扰素刺激基因（ISGs）的诱导明显减弱。相比之下，在表达 IKKγΔ的细胞中，异位 RIG-I.MAVS 或 TNFα 诱导的典型 NF-κB 激活得到保留。增加 IKKγ-WT 相对于 IKKγΔ的相对水平（同时保持总 IKKγ不变）会导致 I 型 IFN 表达增加。相反，过表达 IKKγΔ（在恒定 IKKγ-WT 表达的背景下）表明 IKKγΔ作为 IRF3 信号抑制剂发挥显性负作用。IKKγΔ结合 IKK-α 和 IKK-β，但不结合 TANK 和 IKK-ε，表明外显子 5 编码一个必需的 TANK 结合结构域。最后，IKKγΔ将 IKKγWT 从 MAVS 上置换出来，解释了其显性负效应。
结论/意义：内源性 IKKγΔ的相对表达水平影响细胞对 ssRNA 病毒感染的炎症/抗病毒途径反应的选择。
