Honeysuckle has been used in the treatment of influenza virus infection for thousands of years in China.However,its main active components and the functional mechanisms remain to be elucidated.Here,four honeysuckle ex...Honeysuckle has been used in the treatment of influenza virus infection for thousands of years in China.However,its main active components and the functional mechanisms remain to be elucidated.Here,four honeysuckle extracts,including acids extract,flavonoids extract,total extract and acids-flavonoids mixture,were prepared to clarify the main active antiviral components.The cytopathic effect reduction assay showed that all the four extracts inhibited the replication of influenza viruses H1N1,H3N2 and the oseltamivir-resistant mutant strain H1N1-H275Y.The acids-flavonoids mixture had the strongest inhibitory effects in vitro with EC_(50) values of 3.8,4.1,and >20 lg/m L against H1N1,H3N2 and H1N1-H275Y,respectively,showing competitive antiviral activity with oseltamivir and ribavirin.Honeysuckle acids extract also showed the most significant antiviral activity in vivo.Oral administration of the acids extract at a dosage of 600 mg/kg/d effectively alleviated viral pneumonia,maintained body weight and improved the survival rate to 30% of the mice infected with a lethal dose of H1N1.The results of time-of-drug addition experiment and neuraminidase (NA) inhibition assay showed that honeysuckle extracts had a broad-spectrum inhibitory effect against influenza virus NAs.The flavonoid extract showed the strongest inhibitory effect on the NA of influenza virus H7N9 with an IC_(50) of 24.7 lg/m L.These results suggested that these extracts might exert their antiviral activity by suppressing the release of influenza viruses.Briefly,our findings demonstrate that acids and flavonoids extracts of honeysuckle are the major antiviral active components,and the acids extract has the potential to be developed into an antiviral agent against influenza virus,especially for oseltamivirresistant viruses.展开更多
Influenza A virus(IAV),responsible for seasonal epidemics and recurring pandemics,represents a global threat to public health.Given the risk of a potential IAV pandemic,it is increasingly important to better understan...Influenza A virus(IAV),responsible for seasonal epidemics and recurring pandemics,represents a global threat to public health.Given the risk of a potential IAV pandemic,it is increasingly important to better understand virushost interactions and develop new anti-viral strategies.Here,we reported nonmuscle myosin IIA(MYH9)-mediated regulation of IAV infection.MYH9 depletion caused a profound inhibition of IAV infection by reducing viral attachment and internalization in human lung epithelial cells.Surprisingly,overexpression of MYH9 also led to a significant reduction in viral productive infection.Interestingly,overexpression of MYH9 retained viral attachment,internalization,or uncoating,but suppressed the viral ribonucleoprotein(vRNP)activity in a minigenome system.Further analyses found that excess MYH9 might interrupt the formation of vRNP by interacting with the viral nucleoprotein(NP)and result in the reduction of the completed vRNP in the nucleus,thereby inhibiting subsequent viral RNA transcription and replication.Together,we discovered that MYH9 can interact with IAV NP protein and engage in the regulation of vRNP complexes,thereby involving viral replication.These findings enlighten new mechanistic insights into the complicated interface of host-IAV interactions,ultimately making it an attractive target for the generation of antiviral drugs.展开更多
The matrix protein 2 (M2) is a preferred target for developing a universal vaccine against the influenza A virus (IAV). This study aimed to develop a method for assessing antibody-dependent cell-mediated cytotoxicity ...The matrix protein 2 (M2) is a preferred target for developing a universal vaccine against the influenza A virus (IAV). This study aimed to develop a method for assessing antibody-dependent cell-mediated cytotoxicity (ADCC) associated with M2-based immunization in mice. We first established a stable cell line derived from mouse lymphoma cells (YAC-1) expressing M2 of H3N2. This cell line, designated as YAC-1-M2, was generated using a second-generation lentiviral tricistronic plasmid system to transduce the M2 gene into YAC-1 cells. The ADCC effect induced by polyclonal antibodies targeting matrix protein 2 ectodomain (M2e) was demonstrated by YAC-1-M2 cell lysis by natural killer cells (NK) derived from mice, in the presence of anti-M2 antibodies obtained from mice immunized with an mRNA vaccine based on M2e. This ADCC effect was found to be stronger compared to the effect induced by monoclonal antibodies (14C2) against M2. Moreover, the ADCC effect was enhanced as the effector-to-target ratio of NK to YAC-1-M2 cells increased. In conclusion, we established a novel method to detect ADCC of M2 of IAV, which paves the way for the development of an M2-based universal vaccine against IAV and an in-depth analysis of its mechanism of broad-spectrum immune protection in mice.展开更多
Innate immunity provides immediate defense against viral infection. Influenza A virus(IAV) is able to get past the first line of defense. Elucidation of the molecular interaction between influenza factors and the newl...Innate immunity provides immediate defense against viral infection. Influenza A virus(IAV) is able to get past the first line of defense. Elucidation of the molecular interaction between influenza factors and the newly recognized host players in the innate response might help in our understanding of the root causes of virulence and pathogenicity of IAV. In this study, we show that expression of miR-26 a leads to a significant inhibition of IAV replication. miR-26 a does not directly target IAV genome. Instead, miR-26 a activates the type I interferon(IFN) signaling pathway and promotes the production of IFN-stimulated genes, thus suppressing viral replication. Furthermore,ubiquitin-specific protease 3(USP3), a negative regulator of type I IFN pathway, is targeted by miR-26 a upon IAV challenge. However, miR-26 a is significantly downregulated during IAV infection.Thus, downregulation of miR-26 a is a new strategy evolved by IAV to counteract cellular antiviral responses. Our findings indicate that delivery of miR-26 a may be a potential strategy for anti-IAV therapies.展开更多
Influenza A virus(IAV)genome comprises eight negative-sense RNA segments,of which the replication is well orchestrated and the delicate balance of multiple segments are dynamically regulated throughout IAV life cycle....Influenza A virus(IAV)genome comprises eight negative-sense RNA segments,of which the replication is well orchestrated and the delicate balance of multiple segments are dynamically regulated throughout IAV life cycle.However,previous studies seldom discuss these balances except for functional hemagglutinin-neuraminidase balance that is pivotal for both virus entry and release.Therefore,we attempt to revisit IAV life cycle by highlighting the critical role of“genome balance”.Moreover,we raise a“balance regression”model of IAV evolution that the virus evolves to rebalance its genome after reassortment or interspecies transmission,and direct a“balance compensation”strategy to rectify the“genome imbalance”as a result of artificial modifications during creation of recombinant IAVs.This review not only improves our understanding of IAV life cycle,but also facilitates both basic and applied research of IAV in future.展开更多
Cyclophilin A (CypA) is a peptidyl-prolyl cis/trans isomerase that interacts with the matrix protein (M1) of influenza A virus (IAV) and restricts virus replication by regulating the ubiquitin–proteasome-mediated deg...Cyclophilin A (CypA) is a peptidyl-prolyl cis/trans isomerase that interacts with the matrix protein (M1) of influenza A virus (IAV) and restricts virus replication by regulating the ubiquitin–proteasome-mediated degradation of M1. However,the mechanism by which CypA regulates M1 ubiquitination remains unknown. In this study, we reported that E3 ubiquitin ligase AIP4 promoted K48-linked ubiquitination of M1 at K102 and K104, and accelerated ubiquitin–proteasome-mediated degradation of M1. The recombinant IAV with mutant M1 (K102 R/K104 R) could not be rescued, suggesting that the ubiquitination of M1 at K102/K104 was essential for IAV replication. Furthermore, CypA inhibited AIP4-mediated M1 ubiquitination by impairing the interaction between AIP4 and M1. More importantly, both the mutations of M1 (K102 R/K104 R) and CypA inhibited the nuclear export of M1, indicating that CypA regulates the cellular localization of M1 via inhibition of AIP4-mediated M1 ubiquitination at K102 and K104, which results in the reduced replication of IAV.Collectively, our findings reveal a novel ubiquitination-based mechanism by which CypA regulates the replication of IAV.展开更多
Conventional influenza vaccines need to be designed and manufactured yearly.However,they occasionally provide poor protection owing to antigenic mismatch.Hence,there is an urgent need to develop universal vaccines aga...Conventional influenza vaccines need to be designed and manufactured yearly.However,they occasionally provide poor protection owing to antigenic mismatch.Hence,there is an urgent need to develop universal vaccines against influenza virus.Using nucleoprotein(NP)and extracellular domain of matrix protein 2(M2e)genes from the influenza A virus A/Beijing/30/95(H3N2),we constructed four recombinant vaccinia virus-based influenza vaccines carrying NP fused with one or four copies of M2e genes in different orders.The recombinant vaccinia viruses were used to immunize BALB/C mice.Humoral and cellular responses were measured,and then the immunized mice were challenged with the influenza A virus A/Puerto Rico/8/34(PR8).NP-specific humoral response was elicited in mice immunized with recombinant vaccinia viruses carrying full-length NP,while robust M2e-specific humoral response was elicited only in the mice immunized with recombinant vaccinia viruses carrying multiple copies of M2e.All recombinant viruses elicited NP-and M2e-specific cellular immune responses in mice.Only immunization with RVJ-4M2eNP induced remarkably higher levels of IL-2 and IL-10 cytokines specific to M2e.Furthermore,RVJ-4M2eNP immunization provided the highest cross-protection in mice challenged with 20 MLD5〇of PR8.Therefore,the cross-protection potentially correlates with both NP and M2e-specific humoral and cellular immune responses induced by RVJ-4M2eNP,which expresses a fusion antigen of full-length NP preceded by four M2e repeats.These results suggest that the rational fusion of NP and multiple M2e antigens is critical toward inducing protective immune responses,and the 4M2eNP fusion antigen may be employed to develop a universal influenza vaccine.展开更多
流行性感冒(简称“流感”)是由流感病毒引起的急性呼吸道传染疾病,据世界卫生组织统计,流感每年可导致300万~500万严重病例,其中29万~65万病例死亡,给社会带来沉重的经济负担,是一个世界性的公共卫生难题。研究发现宿主细胞中存在多条...流行性感冒(简称“流感”)是由流感病毒引起的急性呼吸道传染疾病,据世界卫生组织统计,流感每年可导致300万~500万严重病例,其中29万~65万病例死亡,给社会带来沉重的经济负担,是一个世界性的公共卫生难题。研究发现宿主细胞中存在多条信号通路参与对流感病毒感染的应答,越来越多的研究表明宿主miRNAs通过直接或间接的方式,在流感病毒感染、复制的不同阶段发挥着重要调控作用。本文综合分析了目前关于宿主细胞miRNA对流感病毒复制调控的研究进展,对不同的miRNA具体的调控机制进行系统地归类总结后发现:甲型流感病毒(Influenza A virus,IAV)的PB1、PB2、NA、NP、M1基因是宿主miRNA直接抑制病毒复制的主要靶基因,而在间接调控过程中宿主miRNA主要作用在RIG-I样受体信号通路,Jak-STAT信号通路和Toll样受体信号通路三条流感病毒应答信号途径中,以上发现将更有助于全面理解宿主miRNA对于流感病毒调控网络和宿主细胞与流感病毒的互作机制。展开更多
基金supported by the Fundamental Research Funds for the Central Universities (2632017PT03)the National Natural Science Foundation (No. 81700756)。
文摘Honeysuckle has been used in the treatment of influenza virus infection for thousands of years in China.However,its main active components and the functional mechanisms remain to be elucidated.Here,four honeysuckle extracts,including acids extract,flavonoids extract,total extract and acids-flavonoids mixture,were prepared to clarify the main active antiviral components.The cytopathic effect reduction assay showed that all the four extracts inhibited the replication of influenza viruses H1N1,H3N2 and the oseltamivir-resistant mutant strain H1N1-H275Y.The acids-flavonoids mixture had the strongest inhibitory effects in vitro with EC_(50) values of 3.8,4.1,and >20 lg/m L against H1N1,H3N2 and H1N1-H275Y,respectively,showing competitive antiviral activity with oseltamivir and ribavirin.Honeysuckle acids extract also showed the most significant antiviral activity in vivo.Oral administration of the acids extract at a dosage of 600 mg/kg/d effectively alleviated viral pneumonia,maintained body weight and improved the survival rate to 30% of the mice infected with a lethal dose of H1N1.The results of time-of-drug addition experiment and neuraminidase (NA) inhibition assay showed that honeysuckle extracts had a broad-spectrum inhibitory effect against influenza virus NAs.The flavonoid extract showed the strongest inhibitory effect on the NA of influenza virus H7N9 with an IC_(50) of 24.7 lg/m L.These results suggested that these extracts might exert their antiviral activity by suppressing the release of influenza viruses.Briefly,our findings demonstrate that acids and flavonoids extracts of honeysuckle are the major antiviral active components,and the acids extract has the potential to be developed into an antiviral agent against influenza virus,especially for oseltamivirresistant viruses.
基金supported by the National Natural Science Foundation of China(82071788,81901598,81771704,and 82041015)National Key R&D Program of China(2022YFC2604100).
文摘Influenza A virus(IAV),responsible for seasonal epidemics and recurring pandemics,represents a global threat to public health.Given the risk of a potential IAV pandemic,it is increasingly important to better understand virushost interactions and develop new anti-viral strategies.Here,we reported nonmuscle myosin IIA(MYH9)-mediated regulation of IAV infection.MYH9 depletion caused a profound inhibition of IAV infection by reducing viral attachment and internalization in human lung epithelial cells.Surprisingly,overexpression of MYH9 also led to a significant reduction in viral productive infection.Interestingly,overexpression of MYH9 retained viral attachment,internalization,or uncoating,but suppressed the viral ribonucleoprotein(vRNP)activity in a minigenome system.Further analyses found that excess MYH9 might interrupt the formation of vRNP by interacting with the viral nucleoprotein(NP)and result in the reduction of the completed vRNP in the nucleus,thereby inhibiting subsequent viral RNA transcription and replication.Together,we discovered that MYH9 can interact with IAV NP protein and engage in the regulation of vRNP complexes,thereby involving viral replication.These findings enlighten new mechanistic insights into the complicated interface of host-IAV interactions,ultimately making it an attractive target for the generation of antiviral drugs.
基金the National Key Research and Development Program of China(2021YFC2300101).
文摘The matrix protein 2 (M2) is a preferred target for developing a universal vaccine against the influenza A virus (IAV). This study aimed to develop a method for assessing antibody-dependent cell-mediated cytotoxicity (ADCC) associated with M2-based immunization in mice. We first established a stable cell line derived from mouse lymphoma cells (YAC-1) expressing M2 of H3N2. This cell line, designated as YAC-1-M2, was generated using a second-generation lentiviral tricistronic plasmid system to transduce the M2 gene into YAC-1 cells. The ADCC effect induced by polyclonal antibodies targeting matrix protein 2 ectodomain (M2e) was demonstrated by YAC-1-M2 cell lysis by natural killer cells (NK) derived from mice, in the presence of anti-M2 antibodies obtained from mice immunized with an mRNA vaccine based on M2e. This ADCC effect was found to be stronger compared to the effect induced by monoclonal antibodies (14C2) against M2. Moreover, the ADCC effect was enhanced as the effector-to-target ratio of NK to YAC-1-M2 cells increased. In conclusion, we established a novel method to detect ADCC of M2 of IAV, which paves the way for the development of an M2-based universal vaccine against IAV and an in-depth analysis of its mechanism of broad-spectrum immune protection in mice.
基金supported by grants from the National Basic Research Program of China(973 Program,No.2012CB518900)the Beijing Natural Science Foundation(No.7122109)
文摘Innate immunity provides immediate defense against viral infection. Influenza A virus(IAV) is able to get past the first line of defense. Elucidation of the molecular interaction between influenza factors and the newly recognized host players in the innate response might help in our understanding of the root causes of virulence and pathogenicity of IAV. In this study, we show that expression of miR-26 a leads to a significant inhibition of IAV replication. miR-26 a does not directly target IAV genome. Instead, miR-26 a activates the type I interferon(IFN) signaling pathway and promotes the production of IFN-stimulated genes, thus suppressing viral replication. Furthermore,ubiquitin-specific protease 3(USP3), a negative regulator of type I IFN pathway, is targeted by miR-26 a upon IAV challenge. However, miR-26 a is significantly downregulated during IAV infection.Thus, downregulation of miR-26 a is a new strategy evolved by IAV to counteract cellular antiviral responses. Our findings indicate that delivery of miR-26 a may be a potential strategy for anti-IAV therapies.
基金supported by National Natural Science Foundation of China(No.82104134)Key Technology Research and Development Program of Shandong,China(No.2020CXGC010505)The Social Benefiting Technology Program of Qingdao,China(No.21-1-4-rkjk-15-nsh).
文摘Influenza A virus(IAV)genome comprises eight negative-sense RNA segments,of which the replication is well orchestrated and the delicate balance of multiple segments are dynamically regulated throughout IAV life cycle.However,previous studies seldom discuss these balances except for functional hemagglutinin-neuraminidase balance that is pivotal for both virus entry and release.Therefore,we attempt to revisit IAV life cycle by highlighting the critical role of“genome balance”.Moreover,we raise a“balance regression”model of IAV evolution that the virus evolves to rebalance its genome after reassortment or interspecies transmission,and direct a“balance compensation”strategy to rectify the“genome imbalance”as a result of artificial modifications during creation of recombinant IAVs.This review not only improves our understanding of IAV life cycle,but also facilitates both basic and applied research of IAV in future.
基金supported by grants from the National Natural Science Foundation of China (31630079,31672531,31572526,and 31802164)the National Key R&D Program of China (2016YFD0500206,2015BAD11B02)+3 种基金the Strategic Priority Research Program of Chinese Academy of Sciences (XDB29010000)the National Science and Technology Major Project (2018ZX10101004)the Emergency Technology Research Issue on Prevention and Control for Human Infection with A (H7N9) Avian Influenza Virus (10600100000015001206)Wenjun Liu is the principal investigator of the Innovative Research Group of National Natural Science Foundation of China (Grant No.81621091)
文摘Cyclophilin A (CypA) is a peptidyl-prolyl cis/trans isomerase that interacts with the matrix protein (M1) of influenza A virus (IAV) and restricts virus replication by regulating the ubiquitin–proteasome-mediated degradation of M1. However,the mechanism by which CypA regulates M1 ubiquitination remains unknown. In this study, we reported that E3 ubiquitin ligase AIP4 promoted K48-linked ubiquitination of M1 at K102 and K104, and accelerated ubiquitin–proteasome-mediated degradation of M1. The recombinant IAV with mutant M1 (K102 R/K104 R) could not be rescued, suggesting that the ubiquitination of M1 at K102/K104 was essential for IAV replication. Furthermore, CypA inhibited AIP4-mediated M1 ubiquitination by impairing the interaction between AIP4 and M1. More importantly, both the mutations of M1 (K102 R/K104 R) and CypA inhibited the nuclear export of M1, indicating that CypA regulates the cellular localization of M1 via inhibition of AIP4-mediated M1 ubiquitination at K102 and K104, which results in the reduced replication of IAV.Collectively, our findings reveal a novel ubiquitination-based mechanism by which CypA regulates the replication of IAV.
基金supported by grant from the National Key Plan for Scientific Research and Development of China (2016YFC1200200)
文摘Conventional influenza vaccines need to be designed and manufactured yearly.However,they occasionally provide poor protection owing to antigenic mismatch.Hence,there is an urgent need to develop universal vaccines against influenza virus.Using nucleoprotein(NP)and extracellular domain of matrix protein 2(M2e)genes from the influenza A virus A/Beijing/30/95(H3N2),we constructed four recombinant vaccinia virus-based influenza vaccines carrying NP fused with one or four copies of M2e genes in different orders.The recombinant vaccinia viruses were used to immunize BALB/C mice.Humoral and cellular responses were measured,and then the immunized mice were challenged with the influenza A virus A/Puerto Rico/8/34(PR8).NP-specific humoral response was elicited in mice immunized with recombinant vaccinia viruses carrying full-length NP,while robust M2e-specific humoral response was elicited only in the mice immunized with recombinant vaccinia viruses carrying multiple copies of M2e.All recombinant viruses elicited NP-and M2e-specific cellular immune responses in mice.Only immunization with RVJ-4M2eNP induced remarkably higher levels of IL-2 and IL-10 cytokines specific to M2e.Furthermore,RVJ-4M2eNP immunization provided the highest cross-protection in mice challenged with 20 MLD5〇of PR8.Therefore,the cross-protection potentially correlates with both NP and M2e-specific humoral and cellular immune responses induced by RVJ-4M2eNP,which expresses a fusion antigen of full-length NP preceded by four M2e repeats.These results suggest that the rational fusion of NP and multiple M2e antigens is critical toward inducing protective immune responses,and the 4M2eNP fusion antigen may be employed to develop a universal influenza vaccine.
文摘流行性感冒(简称“流感”)是由流感病毒引起的急性呼吸道传染疾病,据世界卫生组织统计,流感每年可导致300万~500万严重病例,其中29万~65万病例死亡,给社会带来沉重的经济负担,是一个世界性的公共卫生难题。研究发现宿主细胞中存在多条信号通路参与对流感病毒感染的应答,越来越多的研究表明宿主miRNAs通过直接或间接的方式,在流感病毒感染、复制的不同阶段发挥着重要调控作用。本文综合分析了目前关于宿主细胞miRNA对流感病毒复制调控的研究进展,对不同的miRNA具体的调控机制进行系统地归类总结后发现:甲型流感病毒(Influenza A virus,IAV)的PB1、PB2、NA、NP、M1基因是宿主miRNA直接抑制病毒复制的主要靶基因,而在间接调控过程中宿主miRNA主要作用在RIG-I样受体信号通路,Jak-STAT信号通路和Toll样受体信号通路三条流感病毒应答信号途径中,以上发现将更有助于全面理解宿主miRNA对于流感病毒调控网络和宿主细胞与流感病毒的互作机制。
