α疱疹病毒的被膜蛋白 UL21 通过 TOLLIP 介导的选择性自噬触发 CGAS 降解来抑制先天免疫。
Tegument protein UL21 of alpha-herpesvirus inhibits the innate immunity by triggering CGAS degradation through TOLLIP-mediated selective autophagy.
机构信息
Key Laboratory of Animal Disease Diagnostics and Immunology, Ministry of Agriculture, MOE International Joint Collaborative Research Laboratory for Animal Health & Food Safety, College of Veterinary Medicine, Nanjing Agricultural University, Nanjing 210095, China.
Institute of Laboratory Medicine, Guangdong Provincial Key Laboratory of Medical Molecular Diagnostics, School of Medical Technology, the First Dongguan Affiliated Hospital, Guangdong Medical University, Dongguan, China.
出版信息
Autophagy. 2023 May;19(5):1512-1532. doi: 10.1080/15548627.2022.2139921. Epub 2022 Nov 7.
Alpha-herpesvirus causes lifelong infections and serious diseases in a wide range of hosts and has developed multiple strategies to counteract the host defense. Here, we demonstrate that the tegument protein UL21 (unique long region 21) in pseudorabies virus (PRV) dampens type I interferon signaling by triggering the degradation of CGAS (cyclic GMP-AMP synthase) through the macroautophagy/autophagy-lysosome pathway. Mechanistically, the UL21 protein scaffolds the E3 ligase UBE3C (ubiquitin protein ligase E3C) to catalyze the K27-linked ubiquitination of CGAS at Lys384, which is recognized by the cargo receptor TOLLIP (toll interacting protein) and degraded in the lysosome. Additionally, we show that the N terminus of UL21 in PRV is dominant in destabilizing CGAS-mediated innate immunity. Moreover, viral tegument protein UL21 in herpes simplex virus type 1 (HSV-1) also displays the conserved inhibitory mechanisms. Furthermore, by using PRV, we demonstrate the roles of UL21 in degrading CGAS to promote viral infection . Altogether, these findings describe a distinct pathway where alpha-herpesvirus exploits TOLLIP-mediated selective autophagy to evade host antiviral immunity, highlighting a new interface of interplay between the host and DNA virus.: 3-MA: 3-methyladenine; ACTB: actin beta; AHV-1: anatid herpesvirus 1; ATG7: autophagy related 7; ATG13: autophagy related 13; ATG101: autophagy related 101; BHV-1: bovine alphaherpesvirus 1; BNIP3L/Nix: BCL2 interacting protein 3 like; CALCOCO2/NDP52: calcium binding and coiled-coil domain 2; CCDC50: coiled-coil domain containing 50; CCT2: chaperonin containing TCP1 subunit 2; CGAS: cyclic GMP-AMP synthase; CHV-2: cercopithecine herpesvirus 2; co-IP: co-immunoprecipitation; CQ: chloroquine; CRISPR: clustered regulatory interspaced short palindromic repeat; Cas9: CRISPR-associated system 9; CTD: C-terminal domain; Ctrl: control; DAPI: 4',6-diamidino-2-phenylindole; DBD: N-terminal DNA binding domain; DMSO: dimethyl sulfoxide; DYNLRB1: dynein light chain roadblock-type 1; EHV-1: equine herpesvirus 1; gB: glycoprotein B; GFP: green fluorescent protein; H&E: hematoxylin and eosin; HSV-1: herpes simplex virus 1; HSV-2: herpes simplex virus 2; IB: immunoblotting; IRF3: interferon regulatory factor 3; lenti: lentivirus; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MARCHF9: membrane associated ring-CH-type finger 9; MG132: cbz-leu-leu-leucinal; NBR1: NBR1 autophagy cargo receptor; NC: negative control; NEDD4L: NEDD4 like E3 ubiquitin protein ligase; NHCl: ammonium chloride; OPTN: optineurin; p-: phosphorylated; PFU: plaque-forming unit; Poly(dA:dT): Poly(deoxyadenylic-deoxythymidylic) acid; PPP1: protein phosphatase 1; PRV: pseudorabies virus; RB1CC1/FIP200: RB1 inducible coiled-coil 1; RNF126: ring finger protein 126; RT-PCR: real-time polymerase chain reaction; sgRNA: single guide RNA; siRNA: small interfering RNA; SQSTM1/p62: sequestosome 1; STING1: stimulator of interferon response cGAMP interactor 1; TBK1: TANK binding kinase 1; TOLLIP: toll interacting protein; TRIM33: tripartite motif containing 33; UL16: unique long region 16; UL21: unique long region 21; UL54: unique long region 54; Ub: ubiquitin; UBE3C: ubiquitin protein ligase E3C; ULK1: unc-51 like autophagy activating kinase 1; Vec: vector; VSV: vesicular stomatitis virus; VZV: varicella-zoster virus; WCL: whole-cell lysate; WT: wild-type; Z-VAD: carbobenzoxy-valyl-alanyl-aspartyl-[O-methyl]-fluoromethylketone.
α疱疹病毒在广泛的宿主中引起终身感染和严重疾病，并发展出多种策略来对抗宿主防御。在这里，我们证明了伪狂犬病病毒（PRV）的包膜蛋白 UL21（独特的长区域 21）通过触发 CGAS（环状 GMP-AMP 合酶）的降解来抑制 I 型干扰素信号，通过巨自噬/自噬溶酶体途径。在机制上，UL21 蛋白支架 E3 连接酶 UBE3C（泛素蛋白连接酶 E3C）催化 CGAS 在 Lys384 处的 K27 连接泛素化，该泛素化被货物受体 TOLLIP（toll 相互作用蛋白）识别，并在溶酶体中降解。此外，我们还表明 PRV 的 UL21 N 端在破坏 CGAS 介导的先天免疫方面具有优势。此外，单纯疱疹病毒 1（HSV-1）中的病毒包膜蛋白 UL21 也显示出保守的抑制机制。此外，我们使用 PRV 证明了 UL21 在降解 CGAS 以促进病毒感染中的作用。总之，这些发现描述了一种独特的途径，α疱疹病毒利用 TOLLIP 介导的选择性自噬来逃避宿主抗病毒免疫，强调了宿主和 DNA 病毒之间相互作用的新界面。：3-MA：3-甲基腺嘌呤；ACTB：肌动蛋白 β；AHV-1：鸭疱疹病毒 1；ATG7：自噬相关 7；ATG13：自噬相关 13；ATG101：自噬相关 101；BHV-1：牛疱疹病毒 1；BNIP3L/Nix：Bcl2 相互作用蛋白 3 样；CALCOCO2/NDP52：钙结合和卷曲螺旋域 2；CCDC50：卷曲螺旋结构域包含 50；CCT2：伴侣蛋白包含 TCP1 亚基 2；CGAS：环状 GMP-AMP 合酶；CHV-2：食蟹猴疱疹病毒 2；co-IP：免疫沉淀；CQ：氯喹；CRISPR：成簇规律间隔短回文重复序列；Cas9：CRISPR 相关系统 9；CTD：C 端结构域；Ctrl：对照；DAPI：4'，6-二脒基-2-苯基吲哚；DBD：N 端 DNA 结合域；DMSO：二甲基亚砜；DYNLRB1：动力蛋白轻链路障 1；EHV-1：马疱疹病毒 1；gB：糖蛋白 B；GFP：绿色荧光蛋白；H&E：苏木精和伊红；HSV-1：单纯疱疹病毒 1；HSV-2：单纯疱疹病毒 2；IB：免疫印迹；IRF3：干扰素调节因子 3；lenti：慢病毒；MAP1LC3/LC3：微管相关蛋白 1 轻链 3；MARCHF9：膜相关环-CH 型指 9；MG132：cbz-leu-leu-leucinal；NBR1：NBR1 自噬货物受体；NC：阴性对照；NEDD4L：NEDD4 样 E3 泛素蛋白连接酶；NHCl：氯化铵；OPTN：optineurin；p-：磷酸化；PFU：噬菌斑形成单位；Poly(dA:dT)：Poly（脱氧腺嘌呤-脱氧胸苷）酸；PPP1：蛋白磷酸酶 1；PRV：伪狂犬病病毒；RB1CC1/FIP200：RB1 诱导的卷曲螺旋 1；RNF126：环指蛋白 126；RT-PCR：实时聚合酶链反应；sgRNA：单引导 RNA；siRNA：小干扰 RNA；SQSTM1/p62：自噬体 1；STING1：干扰素反应 cGAMP 相互作用蛋白 1；TBK1：TANK 结合激酶 1；TOLLIP：toll 相互作用蛋白；TRIM33：三部分基序包含 33；UL16：独特的长区域 16；UL21：独特的长区域 21；UL54：独特的长区域 54；Ub：泛素；UBE3C：泛素蛋白连接酶 E3C；ULK1：UNC-51 样自噬激活激酶 1；Vec：载体；VSV：水疱性口炎病毒；VZV：水痘-带状疱疹病毒；WCL：全细胞裂解物；WT：野生型；Z-VAD：carbobenzoxy-valyl-alanyl-aspartyl-[O-methyl]-fluoromethylketone。
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