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Cellular & Molecular Immunology 2022年第10期19卷 1168-1184页

作者： Xuyuan Wang Li Jiang Guangwen Wang Wenjun Shi Yuzhen Hu Bo Wang Xianying Zeng Guobin Tian Guohua Deng Jianzhong Shi Liling Liu Chengjun Li Hualan Chen College of Veterinary Medicine Gansu Agricultural UniversityLanzhou730070China Guangdong Laboratory for Lingnan Modern Agriculture Guangzhou510642China State Key Laboratory of Veterinary Biotechnology Harbin Veterinary Research InstituteChinese Academy of Agricultural SciencesHarbin150069China

As a major component of the viral ribonucleoprotein(vRNP)complex in influenza a virus(IAV),nucleoprotein(NP)interacts with isoforms of importinαfamily members,leading to the import of itself and vRNP complex into the nucleus,a process pivotal in the replication cycle of *** this study,we found that BinCARD1,an isoform of Bcl10-interacting protein with CARD(BinCARD),was leveraged by IAV for efficient viral ***1 promoted the nuclear import of the vRNP complex and newly synthesized NP and thus enhanced vRNP complex ***,we found that BinCARD1 interacted with NP to promote NP binding to importinα7,an adaptor in the host nuclear import ***,we also found that BinCARD1 promoted RIG-I-mediated innate immune signaling by mediating Lys63-linked polyubiquitination of TRAF3,and that TBK1 appeared to degrade *** showed that BinCARD1 was polyubiquitinated at residue K103 through a Lys63 linkage,which was recognized by the TBK1-p62 axis for autophagic ***,our data demonstrate that IAV leverages BinCARD1 as an important host factor that promotes viral replication,and two mechanisms in the host defense system are triggered—innate immune signaling and autophagic degradation—to mitigate the promoting effect of BinCARD1 on the life cycle of IAV.

关键词： influenza a virus BinCARD1 TRAF3 TBK1 p62

Generation and application of two monoclonal antibodies targeting conserved linear epitopes in the NP protein of influenza a virus

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Journal of Integrative Agriculture 2022年第7期21卷 2095-2105页

作者： ZHAO Yu-hui WEN Xia LI Qi-bing JIANG Li WANG Guang-wen LIANG Li-bin WANG Xiu-rong CHEN Hua-lan LI Cheng-jun State Key Laboratory of Veterinary Biotechnology Harbin Veterinary Research InstituteChinese Academy of Agricultural SciencesHarbin 150069P.R.China

Monoclonal antibodies(mAbs) are widely used in virus research and disease diagnosis. The nucleoprotein(NP) of influenza a virus(IAV) plays important roles in multiple stages of the virus life cycle. Therefore, generating conserved mAbs against NP and characterizing their properties will provide useful tools for IAV research. In this study, two mAbs against the NP protein, 10 E9 and 3 F3, were generated with recombinant truncated NP proteins(NP-1 and NP-2) as immunogens. The heavy-chain subclass of both 10 E9 and 3 F3 was determined to be IgG2α, and the light-chain type was κ. Truncation and site-specific mutation analyses showed that the epitopes of mAbs 10 E9 and 3 F3 were located in the N terminal 84–89 amino acids and the C terminal 320–324 amino acids of the NP protein, respectively. We found that mAbs 10 E9 and 3 F3 reacted well with the NP protein of H1–H15 subtypes of IAV. Both 10 E9 and 3 F3 can be used in immunoprecipitation assay, and 10 E9 was also successfully applied in confocal microscopy. Furthermore, we found that the 10 E9-recognized _(84) SAGKDP_(89) epitope and 3 F3-recognized 320 ENPAH324 epitope were highly conserved in NP among all avian and human IAVs. Thus, the two mAbs we developed could be used as powerful tools in the development of diagnostic methods of IAV, and also surely promote the basic research in understanding the replication mechanisms of IAV.

关键词： influenza a virus nucleoprotein monoclonal antibody application

Role of feline ANP32 proteins in regulating polymerase activity of influenza a virus

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Journal of Integrative Agriculture 2024年第9期23卷 3145-3158页

作者： Gang Lu Feiyan Zheng Yuqing Xiao Ran Shao Jiajun Ou Xin Yin Shoujun Li Guihong Zhang College of Veterinary Medicine South China Agricultural UniversityGuangzhou 510642China State Key Laboratory for Animal Disease Control and Prevention Harbin Veterinary Research InstituteChinese Academy of Agricultural SciencesHarbin 150000China Maoming Branch Guangdong Laboratory for Lingnan Modern AgricultureMaoming 525000China Guangdong Laboratory for Lingnan Modern Agriculture Guangzhou 510642China

Recently,increasing natural infection cases and experimental animal challenge studies demonstrated domestic cats are susceptible to multiple subtypes influenza a virus(IAV)***,some subtype IAV strains could circulate in domestic cats after cross-species transmission and even infected humans,posing a threat to public *** factors related to viral polymerase activity could determine host range of IAV and acidic nuclear phosphoprotein 32(ANP32)is the most important one among ***,role of cat-derived ANP32 on viral polymerase activity and host range of IAV is still *** the present study,a total of 10 feline ANP32(feANP32)splice variants(including 5 feANP32A,3 feANP32B,and 2 feANP32E)were obtained from domestic cats by *** alignment results demonstrated amino acid deletions and/or insertions occurred among feANP32 variants,but all feANP32 proteins were primarily localized to cell *** replication systems for several representative IAV strains were established and the support ability of feANP32 on IAV polymerase activity was *** results indicated that most feANP32A and feANP32B splice variants were able to support all the tested IAV strains,though the support activity of a single feANP32 protein on polymerase activity varied among different IAV *** addition,the role of feANP32 in supporting H3N2 canine influenza virus was determined by investigating viral replication in ***,our study systematically investigated the support activity of feANP32 on IAV,providing a clue for further exploring the mechanism of susceptibility of cats to IAV.

关键词： ANP32 influenza a virus feline cross-species transmission

Nucleoprotein phosphorylation site(Y385)mutation confers temperature sensitivity to influenza a virus due to impaired nucleoprotein oligomerization at a lower temperature

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Science China(Life Sciences) 2021年第4期64卷 633-643页

作者： Weinan Zheng Liang Cui Minghui Li Yun Li Wenhui Fan Limin Yang Jing Li Lei Sun Wenjun Liu CAS Key Laboralory of Pathogenic Microbiology and Immumology Institute of MicrobiologyChinese Academy of SciencesBejing 100101China University of Chinese Academy of Sciences Bejing 100101China State Key Laboratory for Conservation and Urilization of Subtropical Agro-Bioresourses&Laboratory of Animal Infectious Diseases College of Animal Sciences and Veterinary MedicineGuangxi UniversitNanning 530004China Instinute of Microbiology Center for Biosafety Mega-ScienceChinese Academy of SciencesBeijing 100101China

Mutations in viral proteins can lead to the cold adaption of influenza a virus and the cold-adapted virus is an important vaccination ***,we identify a novel strain of influenza a virus with cold sensitivity conferred by a mutation at a phosphorylation site within the nucleoprotein(NP).The highly conserved tyrosine 385 residue(Y385)of NP was identified as a phosphorylation site by mass *** constructive NP phosphorylation mimicked by Y385 E mutation was fatal for virus replication,while the continuous Y385 dephosphorylation mimicked by Y385 F mutation had little impact on virus replication in ***,the Y385 F virus showed much lower replicative capacity in turbinates of mice compared with the wild type ***,the replication of Y385 F virus was significantly reduced in both A549 and MDCK cells grown at 33℃,when compared to that at 37℃.These results indicated that the Y385 F mutation led to cold sensitivity of *** further found that the cold sensitivity of Y385 F virus could be attributed to diminished NP oligomerization rather than any changes in intracellular *** together,these findings suggest that the phosphorylation of NP may be a critical factor that regulates the temperature sensitivity of influenza a virus.

关键词： influenza a virus nucleoprotein phosphorylation dephosphorylation temperature sensitivity

C1QTNF5 is a novel attachment factor that facilitates the entry of influenza a virus

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Virologica Sinica 2024年第2期39卷 277-289页

作者： Lei Yu Xinjin Liu Xiaoqin Wei Junrui Ren Xueyun Wang Shuwen Wu Ke Lan State Key Laboratory of Virology Medical Research InstituteCollege of Life SciencesWuhan UniversityWuhan430072China Frontier Science Center for Immunology and Metabolism Wuhan UniversityWuhan430072China Taikang Center for Life and Medical Sciences Wuhan UniversityWuhan430072China

influenza a virus(IAV)binds sialic acid receptors on the cell surface to enter the host cells,which is the key step in initiating infection,transmission and *** the factors that contribute to the highly efficient entry of IAV into human cells will help elucidate the mechanism of viral entry and pathogenicity,and provide new targets for *** the present study,we reported a novel membrane protein,C1QTNF5,which binds to the hemagglutinin protein of IAV and promotes IAV infection in vitro and in *** found that the HA1 region of IAV hemagglutinin is critical for the interaction with C1QTNF5 protein,and C1QTNF5 interacts with hemagglutinin mainly through its N-terminus(1–103 aa).In addition,we further demonstrated that overexpression of C1QTNF5 promotes IAV entry,while blocking the interaction between C1QTNF5 and IAV hemagglutinin greatly inhibits viral ***,C1QTNF5 does not function as a receptor to mediate IAV infection in sialic acid-deficient CHO-Lec2 cells,but promotes IAV to attach to these cells,suggesting that C1QTNF5 is an important attachment factor for *** work reveals C1QTNF5 as a novel IAV attachment factor and provides a new perspective for antiviral strategies.

关键词： C1QTNF5 influenza a virus Viral entry Hemagglutinin Attachment factor

Roles of the hemagglutinin of influenza a virus in viral entry and development of antiviral therapeutics and vaccines

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Protein & Cell 2010年第4期1卷 342-354页

作者： Shibo Jiang Runming Li Lanying Du Shuwen Liu Lindsley F.Kimball Research Institute New York Blood CenterNew YorkNY 10065USA School of Pharmaceutical Sciences Southern Medical UniversityGuangzhou 510515China

Seasonal influenza epidemics and influenza pandemics caused by influenza a virus(IAV)has resulted in millions of deaths in the *** development of anti-IAV vaccines and therapeutics is urgently needed for prevention and treatment of IAV infection and for controlling future influenza ***(HA)of IAV plays a critical role in viral binding,fusion and entry,and contains the major neutralizing ***,HA is an attractive target for developing anti-IAV drugs and *** we have reviewed the recent progress in study of conformational changes of HA during viral fusion process and development of HA-based antiviral therapeutics and vaccines.

关键词： influenza a virus hemagglutinin viral entry antiviral drugs vaccines

Transmission restriction and genomic evolution co-shape the genetic diversity patterns of influenza a virus

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Virologica Sinica 2024年第4期39卷 525-536页

作者： Xiao Ding Jingze Liu Taijiao Jiang Aiping Wu State Key Laboratory of Common Mechanism Research for Major Diseases Suzhou Institute of Systems MedicineChinese Academy of Medical Sciences&Peking Union Medical CollegeSuzhou215123China Key Laboratory of Pathogen Infection Prevention and Control(Peking Union Medical College) Ministry of EducationBeijing100730China Guangzhou National Laboratory Guangzhou510006China State Key Laboratory of Respiratory Disease The Key Laboratory of Advanced Interdisciplinary Studies CenterThe First Affiliated Hospital of Guangzhou Medical UniversityGuangzhou510030China

influenza a virus(IAV)shows an extensive host range and rapid genomic variations,leading to continuous emergence of novel viruses with significant antigenic variations and the potential for cross-species *** causes global pandemics and seasonal flu outbreaks,posing sustained threats ***,studying all IAVs'evolutionary patterns and underlying mechanisms is crucial for effective prevention and *** developed FluTyping to identify IAV genotypes,to explore overall genetic diversity patterns and their restriction *** groups isolates based on genetic distance and phylogenetic relationships using whole genomes,enabling identification of each isolate's *** distinct genetic diversity patterns were observed:one genotype domination pattern comprising only H1N1 and H3N2 seasonal influenza subtypes,multi-genotypes cocirculation pattern including majority avian influenza subtypes and swine influenza H1N2,and hybrid-circulation pattern involving H7N9 and three H5 subtypes of influenza ***,the IAVs in multi-genotypes cocirculation pattern showed region-specific dominant genotypes,implying the restriction of virus transmission is a key factor contributing to distinct genetic diversity patterns,and the genomic evolution underlying different patterns was more influenced by host-specific *** summary,a comprehensive picture of the evolutionary patterns of overall IAVs is provided by the FluTyping's identified genotypes,offering important theoretical foundations for future prevention and control of these viruses.

关键词： influenza a virus Genetic diversity Transmission Genetic pattern Genotype

M6PR interacts with the HA2 subunit of influenza a virus to facilitate the fusion of viral and endosomal membranes

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Science China(Life Sciences) 2024年第3期67卷 579-595页

作者： Yuzhen Hu Li Jiang Guangwen Wang Yangming Song Zhibo Shan Xuyuan Wang Guohua Deng Jianzhong Shi Guobin Tian Xianying Zeng Liling Liu Hualan Chen Chengjun Li State Key Laboratory for Animal Disease Control and Prevention Harbin Veterinary Research InstituteChinese Academy of Agricultural SciencesHarbin 150069China

influenza a virus(IAV) commandeers numerous host cellular factors for successful replication. However, very few host factors have been revealed to be involved in the fusion of viral envelope and late endosomal membranes. In this study, we identified cation-dependent mannose-6-phosphate receptor(M6PR) as a crucial host factor for the replication of IAV. We found that siRNA knockdown of M6PR expression significantly reduced the growth titers of different subtypes of IAV, and that the inhibitory effect of M6PR siRNA treatment on IAV growth was overcome by the complement of exogenously expressed M6PR. When A549 cells were treated with siRNA targeting M6PR,the nuclear accumulation of viral nucleoprotein(NP) was dramatically inhibited at early timepoints post-infection, indicating that M6PR engages in the early stage of the IAV replication cycle. By investigating the role of M6PR in the individual entry and post-entry steps of IAV replication, we found that the downregulation of M6PR expression had no effect on attachment, internalization, early endosome trafficking,or late endosome acidification. However, we found that M6PR expression was critical for the fusion of viral envelope and late endosomal membranes. Of note, M6PR interacted with the hemagglutinin(HA) protein of IAV, and further studies showed that the lumenal domain of M6PR and the ectodomain of HA2 mediated the interaction and directly promoted the fusion of the viral and late endosomal membranes,thereby facilitating IAV replication. Together, our findings highlight the importance of the M6PR–HA interaction in the fusion of viral and late endosomal membranes during IAV replication.

关键词： influenza a virus M6PR HA membrane fusion late endosome

Antibodies induced by multi-epitope vaccine showed inhibitory activity against heterologous influenza a virus (H3N2)

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Progress in Natural Science:Materials International 2006年第1期16卷 105-108页

作者： DING Jian, WU Fan, WEI Wei and CHEN Yinghua (Laboratory of Molecular Immunology, Department of Biology, Tsinghua University Protein Science Laboratory of the Ministry of Education, Beijing 100084, China) [a]Laboratory of Molecular Immunology Department of Biology Tsinghua University Protein Science Laboratory of Ministry of Education Beijing China

In this study, recognition of 4 recombinant viral proteins (GST-NHA1) by the antibodies induced by multi-epitope vaccine was testified. Inhibitory activities of these antibodies were also investigated in vitro against four heterologous influenza a viruses (H3N2). Three epitope-specific antibodies purified by affinity chromatography could reduce the plaque formation. Interestingly, the three neutralizing antibodies in combination showed obvious enhancement of inhibitory activity, suggesting that the development of recombinant multi-epitope vaccine might be an effective way against viral mutation.

关键词： influenza a virus recombinant multi-epitope vaccine neutralizing antibody plaque reduction assay.

Discovery of the anti-influenza a virus activity of SB216763 and cyclosporine A by mining infected cells and compound cellular signatures

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Chinese Chemical Letters 2022年第5期33卷 2541-2544页

作者： Ke Tang You Wu Shubing Chen Yijing Xin Ying Guo State Key Laboratory of Bioactive Substance and Function of Natural Medicines Institute of Materia MedicaChinese Academy of Medical Sciences and Peking Union Medical CollegeBeijing 100050China Department of Pharmacology Institute of Materia MedicaChinese Academy of Medical Sciences and Peking Union Medical CollegeBeijing 100050China

In this study,SB216763 and cyclosporine A were identified as anti-influenza a virus(IAV)agents by tran-scriptome signature reversion(TSR)analysis through deep mining of the cellular transcriptome of hu-man airway and lung cell lines infected with 3 strains of IAV and the chemical perturbations library.A synergistic effect of SB216763 and cyclosporine A against influenza A was disclosed by quantification of the network-based relationship,which was validated in *** with burgeoning omics approaches,transcriptome-based drug development is flourishing,which provides a novel insight into antivirals dis-covery with comprehensive cellular transcriptional information of disease and chemical perturbations in multicomponent *** strategy can be applied as a new approach in discovering multitar-get antiviral agents from approved drugs,clinical compounds,natural products or other known bioactive compounds.

关键词： Chemical perturbagen signature Host factor Transcriptome signature reversion influenza a virus SB216763 Cyclosporine A Drug combination