Ramnani Barkha, Devale Trupti, Manivannan Praveen, Haridas Aiswarya, Malathi Krishnamurthy
Department of Biological Sciences, University of Toledo, 2801 West Bancroft Street, Toledo, OH 43606, USA.
Department of Internal Medicine, University of Michigan, Ann Arbor, MI 48109, USA.
Viruses. 2024 Dec 13;16(12):1913. doi: 10.3390/v16121913.
During virus infection, the activation of the antiviral endoribonuclease, ribonuclease L (RNase L), by a unique ligand 2'-5'-oilgoadenylate (2-5A) causes the cleavage of single-stranded viral and cellular RNA targets, restricting protein synthesis, activating stress response pathways, and promoting cell death to establish broad antiviral effects. The immunostimulatory dsRNA cleavage products of RNase L activity (RL RNAs) recruit diverse dsRNA sensors to activate signaling pathways to amplify interferon (IFN) production and activate inflammasome, but the sensors that promote cell death are not known. In this study, we found that DEAH-box polypeptide 15 (DHX15) and retinoic acid-inducible gene I (Rig-I) are essential for apoptosis induced by RL RNAs and require mitochondrial antiviral signaling (MAVS), c-Jun amino terminal kinase (JNK), and p38 mitogen-activated protein kinase (p38 MAPK) for caspase-3-mediated intrinsic apoptosis. In RNase L-activated cells, DHX15 interacts with Rig-I and MAVS, and cells lacking MAVS expression were resistant to apoptosis. RL RNAs induced the transcription of genes for IFN and proinflammatory cytokines by interferon regulatory factor 3 (IRF-3) and nuclear factor kB (NF-kB), while cells lacking both DHX15 and Rig-I showed a reduced induction of cytokines. However, apoptotic cell death is independent of both IRF-3 and NF-kB, suggesting that cytokine and cell death induction by RL RNAs are uncoupled. The RNA binding of both DHX15 and Rig-I is required for apoptosis induction, and the expression of both single proteins in cells lacking both DHX15 and Rig-I is insufficient to promote cell death by RL RNAs. Cell death induced by RL RNAs suppressed Coxsackievirus B3 (CVB3) replication, and inhibiting caspase-3 activity or cells lacking IRF-3 showed that the induction of apoptosis directly resulted in the CVB3 antiviral effect, and the effects were independent of the role of IRF-3.
在病毒感染期间，抗病毒内切核糖核酸酶核糖核酸酶L（RNase L）被独特的配体2'-5'-寡腺苷酸（2-5A）激活，导致单链病毒和细胞RNA靶标的切割，限制蛋白质合成，激活应激反应途径，并促进细胞死亡以建立广泛的抗病毒作用。RNase L活性的免疫刺激双链RNA切割产物（RL RNAs）招募多种双链RNA传感器以激活信号通路，从而放大干扰素（IFN）的产生并激活炎性小体，但促进细胞死亡的传感器尚不清楚。在本研究中，我们发现DEAH盒多肽15（DHX15）和视黄酸诱导基因I（Rig-I）对于RL RNAs诱导的细胞凋亡至关重要，并且对于caspase-3介导的内源性细胞凋亡需要线粒体抗病毒信号（MAVS）、c-Jun氨基末端激酶（JNK）和p38丝裂原活化蛋白激酶（p38 MAPK）。在RNase L激活的细胞中，DHX15与Rig-I和MAVS相互作用，而缺乏MAVS表达的细胞对细胞凋亡具有抗性。RL RNAs通过干扰素调节因子3（IRF-3）和核因子κB（NF-κB）诱导IFN和促炎细胞因子基因的转录，而同时缺乏DHX15和Rig-I的细胞显示细胞因子的诱导减少。然而，凋亡性细胞死亡独立于IRF-3和NF-κB，这表明RL RNAs诱导的细胞因子和细胞死亡是解偶联的。DHX15和Rig-I的RNA结合对于诱导细胞凋亡是必需的，并且在同时缺乏DHX15和Rig-I的细胞中单一蛋白质的表达不足以通过RL RNAs促进细胞死亡。RL RNAs诱导的细胞死亡抑制了柯萨奇病毒B3（CVB3）的复制，并且抑制caspase-3活性或缺乏IRF-3的细胞表明细胞凋亡的诱导直接导致了CVB3的抗病毒作用，并且这些作用独立于IRF-3的作用。
