Human herpesvirus 8 (HHV-8) encodes four viral interferon regulatory factors (vIRFs) that target cellular IRFs and/or other innate-immune and stress signaling regulators and suppress the cellular response to viral infection and replication. For vIRF-1, cellular protein targets include IRFs, p53, p53-activating ATM kinase, BH3-only proteins, and antiviral signaling effectors MAVS and STING; vIRF-1 inhibits each, with demonstrated or likely promotion of HHV-8 de novo infection and productive replication. Here, we identify direct interactions of vIRF-1 with STAT3 and STAT-activating Janus kinase TYK2 (the latter reported previously by us to be inhibited by vIRF-1) and suppression by vIRF-1 of cytokine-induced STAT3 activation. Suppression of active, phosphorylated STAT3 (pSTAT3) by vIRF-1 was evident in transfected cells and vIRF-1 ablation in lytically-reactivated recombinant-HHV-8-infected cells led to increased levels of pSTAT3. Using a panel of vIRF-1 deletion variants, regions of vIRF-1 required for interactions with STAT3 and TYK2 were identified, which enabled correlation of STAT3 signaling inhibition by vIRF-1 with TYK2 binding, independently of STAT3 interaction. A viral mutant expressing vIRF-1 deletion-variant Δ198-222 refractory for TYK2 interaction and pSTAT3 suppression was severely compromised for productive replication. Conversely, expression of phosphatase-resistant, protractedly-active STAT3 led to impaired HHV-8 replication. Cells infected with HHV-8 mutants expressing STAT3-refractory vIRF-1 deletion variants or depleted of STAT3 displayed reduced vIRF-1 expression, while custom-peptide-promoted STAT3 interaction could effect increased vIRF-1 expression and enhanced virus replication. Taken together, our data identify vIRF-1 targeting and inhibition of TYK2 as a mechanism of STAT3-signaling suppression and critical for HHV-8 productive replication, the importance of specific pSTAT3 levels for replication, positive roles of STAT3 and vIRF-1-STAT3 interaction in vIRF-1 expression, and significant contributions to lytic replication of STAT3 targeting by vIRF-1.

中文翻译：

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人疱疹病毒 8 干扰素调节因子 1 与 STAT3 和 Janus 激酶 TYK2 的直接且具有生物学意义的相互作用。

人类疱疹病毒 8 (HHV-8) 编码四种病毒干扰素调节因子 (vIRF)，这些因子以细胞 IRF 和/或其他先天免疫和应激信号调节因子为目标，并抑制细胞对病毒感染和复制的反应。对于 vIRF-1，细胞蛋白靶点包括 IRF、p53、p53 激活 ATM 激酶、BH3-only 蛋白以及抗病毒信号效应器 MAVS 和 STING；vIRF-1 抑制每一种，并已证实或可能促进 HHV-8 从头感染和有效复制。在这里，我们确定了 vIRF-1 与 STAT3 和 STAT 激活 Janus 激酶 TYK2（我们之前报道后者被 vIRF-1 抑制）的直接相互作用以及 vIRF-1 对细胞因子诱导的 STAT3 激活的抑制。在转染的细胞中，vIRF-1 对活性磷酸化 STAT3 (pSTAT3) 的抑制是明显的，而在裂解性再激活的重组 HHV-8 感染细胞中，vIRF-1 的消除导致 pSTAT3 水平升高。使用一组 vIRF-1 缺失变体，鉴定了与 STAT3 和 TYK2 相互作用所需的 vIRF-1 区域，这使得 vIRF-1 的 STAT3 信号传导抑制与 TYK2 结合相关联，而与 STAT3 相互作用无关。表达 vIRF-1 缺失变体 Δ198-222 的病毒突变体对 TYK2 相互作用和 pSTAT3 抑制无效，其有效复制受到严重损害。相反，磷酸酶抗性、长期活性的 STAT3 的表达会导致 HHV-8 复制受损。感染表达 STAT3 难治性 vIRF-1 缺失变体或 STAT3 耗尽的 HHV-8 突变体的细胞显示出 vIRF-1 表达降低，而定制肽促进的 STAT3 相互作用可能会增加 vIRF-1 表达并增强病毒复制。综上所述，我们的数据确定了 vIRF-1 靶向和抑制 TYK2 作为 STAT3 信号传导抑制的机制，对 HHV-8 高效复制至关重要、特定 pSTAT3 水平对复制的重要性、STAT3 和 vIRF-1-STAT3 的积极作用vIRF-1 表达中的相互作用，以及对 vIRF-1 靶向的 STAT3 裂解复制的显着贡献。