Human enterovirus A71(EV-A71)is a major causative agent of hand,foot and mouth disease(HFMD),which poses a significant public health threat,particularly among young children.Mitochondrial antiviral signaling protein(M...Human enterovirus A71(EV-A71)is a major causative agent of hand,foot and mouth disease(HFMD),which poses a significant public health threat,particularly among young children.Mitochondrial antiviral signaling protein(MAVS)and interferon regulatory factor 3(IRF3)are vital proteins for the induction of type I interferons(IFN-I)and downstream interferon-stimulated genes(ISGs)during EVA71 infection.While posttranslational modifications are known to critically influence viral infection processes,the mechanisms by which EV-A71 exploits host deubiquitinases(DUBs)for immune evasion remain poorly understood.In this study,we demonstrated that EV-A71 infection upregulated ubiquitinspecific protease 5(USP5)expression.Knockdown of USP5 not only inhibited EV-A71 replication but also observably increased the production of IFN-I and ISGs.Furthermore,USP5 also regulated the replication of EV-D68 and CVA16 and the production of IFN-I and ISGs.Mechanistically,USP5 physically interacted with MAVS and IRF3 and reduced the K63-linked polyubiquitination of MAVS and IRF3.Conversely,USP5 knockdown increased the K63-linked polyubiquitination of MAVS and IRF3,thereby accelerating the phosphorylation of IRF3 and increasing IFN-I production during EV-A71 infection.Furthermore,pharmacological inhibition of USP5 with the small-molecule inhibitor PR-619 significantly potentiated the antiviral effects of IFN against EV-A71.Collectively,our findings reveal a previously unrecognized role of USP5 in facilitating EV-A71 immune evasion by dampening MAVSand IRF3-mediated antiviral signaling.These insights provide a novel therapeutic avenue for combating EV-A71 infection through targeted modulation of the USP5-IRF3 axis.展开更多
本研究利用CRISPR/Cas9基因编辑技术构建MAVS基因稳定敲除的细胞株,对流感病毒的增殖特性进行初步研究。设计、构建靶向MAVS基因sgRNA表达载体,与pCAG-Cas9-EGFP表达载体共转染MDCK细胞,经过流式细胞仪分选、PCR、基因测序筛选MAVS敲除...本研究利用CRISPR/Cas9基因编辑技术构建MAVS基因稳定敲除的细胞株,对流感病毒的增殖特性进行初步研究。设计、构建靶向MAVS基因sgRNA表达载体,与pCAG-Cas9-EGFP表达载体共转染MDCK细胞,经过流式细胞仪分选、PCR、基因测序筛选MAVS敲除细胞系,CCK-8法检测细胞增殖速度;荧光定量PCR方法检测H9N2亚型禽流感病毒(AIV)感染后的TCID_(50)、病毒拷贝数以及IRF3、IFN-β、Mx1基因转录水平变化。结果显示,筛选出1株MAVS基因缺失44 bp的MDCK细胞(MAVS^(-/-)MDCK),其增殖速度与正常细胞相比未观察到显著差异;荧光定量PCR结果表明,TCID_(50)、病毒拷贝数差异最高可分别达到MDCK细胞的4.11倍和1.82倍;MAVS^(-/-)MDCK中IRF3、IFN-β和Mx1 m RNA表达水平显著降低,表明MAVS敲除后抑制了Ⅰ型干扰素信号通路。表明,本研究获得的MAVS^(-/-)MDCK能够促进禽流感病毒的复制,为提高疫苗生产效率和质量提供候选细胞株;该细胞株也为进一步研究MAVS参与抗病毒天然免疫应答奠定基础。展开更多
Although radiation induced bystander effect(RIBE)has been investigated for decades for secondary cancer risk assessment during cancer radiotherapy,the underlying gene regulation remains unclear,especially the roles of...Although radiation induced bystander effect(RIBE)has been investigated for decades for secondary cancer risk assessment during cancer radiotherapy,the underlying gene regulation remains unclear,especially the roles of immune factors in RIBE.Mitochondrial antiviral signaling(MAVS)protein.展开更多
Mitochondrial antiviral signaling(MAVS)protein is signaling adaptor with antiviral feature and locate in the mitochondrial out-membrane.Our study demonstrated that knockdown of MAVS increases the radioresistance and i...Mitochondrial antiviral signaling(MAVS)protein is signaling adaptor with antiviral feature and locate in the mitochondrial out-membrane.Our study demonstrated that knockdown of MAVS increases the radioresistance and irradiation(IR)induced the change of MAVS expression in cells.Knockdown of MAVS alone could decrease the mitochondrial membrane potential,while increase the mitochondrial ATP production and the expressions of apoptosis related proteins.While knockdown of MAVS followed by X-rays increased the cellular mitochondrial membrane potential,ATP production and expression of apoptosis protein,compared to the IR group only.展开更多
The mitochondrial antiviral signaling protein(MAVS)plays a pivotal role in the interferon(IFN)activation signaling pathway,which culminates in the production of IFN and the establishment of a positive feedback loop.Th...The mitochondrial antiviral signaling protein(MAVS)plays a pivotal role in the interferon(IFN)activation signaling pathway,which culminates in the production of IFN and the establishment of a positive feedback loop.The unrestricted expression of IFN has been demonstrated to result in the development of autoimmune diseases,therefore IFN must be subject to strict regulation.The present study reports a novel mechanism by which IFN expression is negatively regulated in fish by LIM and SH3 protein 1(LASP1).In the aftermath of infection with the spring viremia of carp virus(SVCV),a substantial increase in lasp1 mRNA levels was observed.Furthermore,overexpression of LASP1 led to a reduction in IFN expression,thereby facilitating SVCV replication.Conversely,the knockdown of LASP1 resulted in an augmentation of the ability of cells to produce IFN,thus attenuating SVCV replication.Subsequent research revealed that LASP1 bound and degraded MAVS via autophagic degradation.Subsequent analysis of the mechanism revealed that LASP1 preferentially associated with a selective autophagy receptor,a neighbor of BRCA1(NBR1),which facilitated the autophagy degradation of MAVS.Finally,the degradation cascade mediated by LASP1 facilitates SVCV infection by blocking the MAVS-mediated innate signaling pathway.In conclusion,these data reveal LASP1 to act as a negative regulator of the host's innate immune responses to SVCV infection and provide insights into the selective autophagy and innate signaling pathways.展开更多
Innate immunity in host cells must be rapidly activated to combat invading microbes.Upon RIG-I activation,the transcription of type I interferons is induced within one hour in virus-infected cells.Previous studies hav...Innate immunity in host cells must be rapidly activated to combat invading microbes.Upon RIG-I activation,the transcription of type I interferons is induced within one hour in virus-infected cells.Previous studies have shown that endogenous MAVS spreads signals via aggregation on the mitochondrial membrane,whereas truncated recombinant MAVS forms prion-like filaments in vitro.How MAVS transmits signals so quickly,and the molecular architecture of its membrane aggregates,remains elusive.Here,we report that activated MAVS forms fibrils encircling its resident mitochondrion or connecting neighboring mitochondria with a“ladder-like”structure,allowing the activation of dormant MAVS on encountered mitochondria.This“intermitochondrial activation”process promotes a rapid antiviral response in cells to overcome the immediate danger caused by viruses.Moreover,stuck MAVS fibrils between mitochondria have limited cytosolic protein access and thus relay signals poorly.This study demonstrated that prion-like MAVS fibrils cluster in mitochondria to ensure a rapid antiviral response.展开更多
基金supported by the National Natural Science Foundation of China(32300133 to SZ.and 32100106 to YR)the China Postdoctoral Science Foundation(2023M730965 to SZ.)+3 种基金the Science and Technology Department of Henan Province(232102311103 to SZ.)the Chinese Academy of Sciences(CAS)Youth Innovation Promotion Association(2023351 to YR)the Hubei Province Natural Science Funds(2023AFA008 and 2023AFB582 to YR)the Open project of the State Key Laboratory of Antiviral Drugs,Henan University(FX3020A030002).
文摘Human enterovirus A71(EV-A71)is a major causative agent of hand,foot and mouth disease(HFMD),which poses a significant public health threat,particularly among young children.Mitochondrial antiviral signaling protein(MAVS)and interferon regulatory factor 3(IRF3)are vital proteins for the induction of type I interferons(IFN-I)and downstream interferon-stimulated genes(ISGs)during EVA71 infection.While posttranslational modifications are known to critically influence viral infection processes,the mechanisms by which EV-A71 exploits host deubiquitinases(DUBs)for immune evasion remain poorly understood.In this study,we demonstrated that EV-A71 infection upregulated ubiquitinspecific protease 5(USP5)expression.Knockdown of USP5 not only inhibited EV-A71 replication but also observably increased the production of IFN-I and ISGs.Furthermore,USP5 also regulated the replication of EV-D68 and CVA16 and the production of IFN-I and ISGs.Mechanistically,USP5 physically interacted with MAVS and IRF3 and reduced the K63-linked polyubiquitination of MAVS and IRF3.Conversely,USP5 knockdown increased the K63-linked polyubiquitination of MAVS and IRF3,thereby accelerating the phosphorylation of IRF3 and increasing IFN-I production during EV-A71 infection.Furthermore,pharmacological inhibition of USP5 with the small-molecule inhibitor PR-619 significantly potentiated the antiviral effects of IFN against EV-A71.Collectively,our findings reveal a previously unrecognized role of USP5 in facilitating EV-A71 immune evasion by dampening MAVSand IRF3-mediated antiviral signaling.These insights provide a novel therapeutic avenue for combating EV-A71 infection through targeted modulation of the USP5-IRF3 axis.
文摘本研究利用CRISPR/Cas9基因编辑技术构建MAVS基因稳定敲除的细胞株,对流感病毒的增殖特性进行初步研究。设计、构建靶向MAVS基因sgRNA表达载体,与pCAG-Cas9-EGFP表达载体共转染MDCK细胞,经过流式细胞仪分选、PCR、基因测序筛选MAVS敲除细胞系,CCK-8法检测细胞增殖速度;荧光定量PCR方法检测H9N2亚型禽流感病毒(AIV)感染后的TCID_(50)、病毒拷贝数以及IRF3、IFN-β、Mx1基因转录水平变化。结果显示,筛选出1株MAVS基因缺失44 bp的MDCK细胞(MAVS^(-/-)MDCK),其增殖速度与正常细胞相比未观察到显著差异;荧光定量PCR结果表明,TCID_(50)、病毒拷贝数差异最高可分别达到MDCK细胞的4.11倍和1.82倍;MAVS^(-/-)MDCK中IRF3、IFN-β和Mx1 m RNA表达水平显著降低,表明MAVS敲除后抑制了Ⅰ型干扰素信号通路。表明,本研究获得的MAVS^(-/-)MDCK能够促进禽流感病毒的复制,为提高疫苗生产效率和质量提供候选细胞株;该细胞株也为进一步研究MAVS参与抗病毒天然免疫应答奠定基础。
文摘Although radiation induced bystander effect(RIBE)has been investigated for decades for secondary cancer risk assessment during cancer radiotherapy,the underlying gene regulation remains unclear,especially the roles of immune factors in RIBE.Mitochondrial antiviral signaling(MAVS)protein.
文摘Mitochondrial antiviral signaling(MAVS)protein is signaling adaptor with antiviral feature and locate in the mitochondrial out-membrane.Our study demonstrated that knockdown of MAVS increases the radioresistance and irradiation(IR)induced the change of MAVS expression in cells.Knockdown of MAVS alone could decrease the mitochondrial membrane potential,while increase the mitochondrial ATP production and the expressions of apoptosis related proteins.While knockdown of MAVS followed by X-rays increased the cellular mitochondrial membrane potential,ATP production and expression of apoptosis protein,compared to the IR group only.
基金supported by the National Key Research and Development Program of China(2023YFD2400201)National Excellent Youth Science Fund(32322086)+1 种基金Youth Innovation Promotion Association(S.L.)National Natural Science Foundation of China(31930111and 32173023).
文摘The mitochondrial antiviral signaling protein(MAVS)plays a pivotal role in the interferon(IFN)activation signaling pathway,which culminates in the production of IFN and the establishment of a positive feedback loop.The unrestricted expression of IFN has been demonstrated to result in the development of autoimmune diseases,therefore IFN must be subject to strict regulation.The present study reports a novel mechanism by which IFN expression is negatively regulated in fish by LIM and SH3 protein 1(LASP1).In the aftermath of infection with the spring viremia of carp virus(SVCV),a substantial increase in lasp1 mRNA levels was observed.Furthermore,overexpression of LASP1 led to a reduction in IFN expression,thereby facilitating SVCV replication.Conversely,the knockdown of LASP1 resulted in an augmentation of the ability of cells to produce IFN,thus attenuating SVCV replication.Subsequent research revealed that LASP1 bound and degraded MAVS via autophagic degradation.Subsequent analysis of the mechanism revealed that LASP1 preferentially associated with a selective autophagy receptor,a neighbor of BRCA1(NBR1),which facilitated the autophagy degradation of MAVS.Finally,the degradation cascade mediated by LASP1 facilitates SVCV infection by blocking the MAVS-mediated innate signaling pathway.In conclusion,these data reveal LASP1 to act as a negative regulator of the host's innate immune responses to SVCV infection and provide insights into the selective autophagy and innate signaling pathways.
基金supported by the National Natural Science Foundation of China(32130038)the Chinese Ministry of Science and Technology(2024YFA1306501 and 2022YFC2303700)+1 种基金the China Postdoctoral Science Foundation(2021M700242 and 2022T150017)the Young Elite Scientists Sponsorship Program by CAST(2022QNRC001).
文摘Innate immunity in host cells must be rapidly activated to combat invading microbes.Upon RIG-I activation,the transcription of type I interferons is induced within one hour in virus-infected cells.Previous studies have shown that endogenous MAVS spreads signals via aggregation on the mitochondrial membrane,whereas truncated recombinant MAVS forms prion-like filaments in vitro.How MAVS transmits signals so quickly,and the molecular architecture of its membrane aggregates,remains elusive.Here,we report that activated MAVS forms fibrils encircling its resident mitochondrion or connecting neighboring mitochondria with a“ladder-like”structure,allowing the activation of dormant MAVS on encountered mitochondria.This“intermitochondrial activation”process promotes a rapid antiviral response in cells to overcome the immediate danger caused by viruses.Moreover,stuck MAVS fibrils between mitochondria have limited cytosolic protein access and thus relay signals poorly.This study demonstrated that prion-like MAVS fibrils cluster in mitochondria to ensure a rapid antiviral response.
