Autophagic degradation of PML promotes susceptibility to HSV-1 by stress-induced Corticosterone
Autophagic degradation of PML promotes susceptibility to HSV-1 by stress-induced Corticosterone
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PML 的自噬降解通过应激诱导的皮质酮促进对 HSV-1 的易感性
DOI：
10.7150/thno.46921
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发表时间：
2020-01-01
期刊：
影响因子：
12.4
通讯作者：
He, Rong-Rong
中科院分区：
文献类型：
--
作者：
Li, Wen;Luo, Zhuo;He, Rong-Rong
理论基础：单纯疱疹病毒1型(HSV-1)是一种嗜神经性病毒，可引起多种临床症状，包括黏膜皮肤病和HSV-1脑炎(HSE)。在这里，我们描述了在应激条件下对HSV-1易感性的分子机制。方法：分别采用束缚应激和皮质酮(皮质酮，主要应激激素)在体内和体外建立HSV-1易感模型。空斑试验测定病毒滴度。用Western blotting、免疫荧光、透射电子显微镜、qRT-PCR、H&E染色、IHC染色和流式细胞术检测病毒相关蛋白的表达和自噬的激活。在机械实验中应用了丧失和获得功能分析、免疫共沉淀(co-IP)技术和自噬激动剂/拮抗剂治疗。结果：束缚应激增加了小鼠脑对HSV-1的易感性。同样，皮质醇治疗提高了神经细胞对HSV-1的敏感性。体内应激处理或皮质醇处理可显著降低HSV-1感染的脑组织和神经细胞中PML蛋白水平，但对其转录水平无明显影响。值得注意的是，在PML过表达的细胞中观察到ICP27和GB的蛋白表达显著下降，这一点被皮质醇处理逆转。而在病毒感染的SH-SY5Y细胞中，用si-PML基因敲除后，gB蛋白的表达增加。我们进一步发现，皮质醇驱动的PML降解依赖于自噬的激活，这种方式不依赖于ULK1，而不是蛋白酶体途径。BaF1可减弱CORT对HSV-1感染的增强作用。在Lc3缺失的细胞中，病毒蛋白的表达减少，而在Lc3基因敲除的细胞中，RNAi阻止了皮质醇诱导的自噬对PML的降解。有趣的是，PML被发现与自噬货物受体p62和自噬效应蛋白Lc3相互作用。此外，当PML被沉默时，CORT未能增加GB蛋白水平，这为PML的自噬降解与CORT诱导的病毒易感性提供了直接证据。结论：束缚应激/皮质醇通过将PML导入自溶酶体进行降解，增加了HSV-1的易感性。从体外和体内模型获得的结果不仅证明了应激对HSV-1感染的不利影响，而且系统地研究了潜在的分子机制。这些发现拓宽了我们对宿主和病毒之间相互作用的理解，全面了解自噬在病毒感染中的作用将为未来开发抗病毒感染的创新药物提供信息。
Rationale: Herpes simplex virus type 1 (HSV-1) is a neurotropic virus that can cause a variety of clinical syndromes including mucocutaneous disease and HSV-1 encephalitis (HSE). Here, we characterize the molecular mechanisms underlying the susceptibility to HSV-1 under stressful conditions. Methods: Restraint stress and corticosterone (CORT, a primary stress hormone) were respectively used to establish HSV-1 susceptible model in vivo and in vitro. Viral titers were determined by plaque assay. Western blotting, immunofluorescence, transmission electron microscopy (TEM), qRT-PCR, H&E staining, IHC staining and flow cytometry were employed to evaluate virus-related protein expressions and detect the activation of autophagy. Loss- and gain-function assays, co-immunoprecipitation (co-IP) technique and autophagy agonist/antagonist treatments were applied in mechanistic experiments. Results: Restraint stress increased the susceptibility of mouse brain to HSV-1. Similarly, CORT treatment enhanced the susceptibility of neural cells to HSV-1. Furthermore, PML protein level in HSV-1 infected brain tissues and neural cells was remarkably decreased by stress treatment in vivo or CORT treatment in vitro, while its transcriptional level was not affected. Notably, a striking decline in protein expressions of ICP27 and gB was observed in PML-overexpressing cells, which was reversed by CORT treatment. By contrast, protein expression of gB was increased by knockdown with si-PML in virus-infected SH-SY5Y cells. We further discovered that CORT-driven PML degradation was dependent on the activation of autophagy in a ULK1-independent manner, rather than proteasome pathway. Bafilomycin A1 (BaF1) attenuated the augmentation effect of CORT on HSV-1 infection. The expressions of viral proteins were reduced in LC3-depleted cells, and the degradation of PML by CORT-induced autophagy was prevented in cells with LC3 knockdown by RNAi. Interestingly, PML was revealed to interact with the autophagic cargo receptor P62 and the autophagic effector protein LC3. Additionally, CORT failed to increase gB protein level when PML was silenced, providing direct evidence linking autophagic degradation of PML and CORT-induced virus susceptibility. Conclusion: Our results revealed that restraint stress/CORT increased HSV-1 susceptibility by delivering PML into autolysosomes for degradation. The results obtained from in vitro and in vivo models not only demonstrated the adverse effects of stress on HSV-1 infection, but also systematically investigated the underlying molecular mechanisms. These discoveries broaden our understanding of the interplay between host and viruses, and a comprehensive understanding of the role of autophagy in viral infection will provide information for future development of innovative drugs against viral infection.