{"title":"Cholesterol-dependent Nsp5-endosomes co-trafficking to lysosomes facilitates porcine reproductive and respiratory syndrome virus replication by activating autophagy","authors":"Haoxin Dong , Qiming Pei , Jiahui Ren , Yaci Zhang , Xuedan Wei , Aijuan Shen , Yunshuo Lu , Ziheng Zhang , Yongkun Du , Guoqing Zhuang , Angke Zhang , Hong Duan","doi":"10.1016/j.vetmic.2025.110507","DOIUrl":null,"url":null,"abstract":"<div><div>Our previous studies showed that intracellular endosomal vesicles participated in PRRS virions trafficking in the early stage of viral infection, and cholesterol retention in endosomal vesicles disturbed viral replication via blocking PRRSV-endosomal vesicles membrane fusion. However, whether endosomal vesicles were associated with PRRSV protein(s) trafficking and the role of cholesterol in this process was still unclarity. In this study, we sought to elucidate the mechanism of cholesterol in endosomal vesicles-mediated viral protein transportation. The results showed that endosomal vesicles participated in trafficking of PRRSV Nsp5 protein. After being synthesized in endoplasmic reticulum (ER) and Golgi apparatus, Nsp5 was trafficked to early endosomes (EEs), but not endocytic recycling compartments (ERCs), then to late endosomes (LEs), and eventually reached lysosomal compartments, whereas disruption of cholesterol flux or LEs function led to the inability of Nsp5 arriving at lysosomes, where Nsp5 activated cellular autophagy to promote PRRSV replication. Molecular docking predictions revealed that cholesterol could form two hydrogen bonds with 74 alanine and 78 asparagine of Nsp5. After mutating the aforementioned binding sites, the replication efficiency of PRRSV decreased. Subsequently, the role of cholesterol in PRRSV replication was explored. Blocking of cholesterol flux significantly inhibited PRRSV replication. Single virus infection cycle analysis showed that cholesterol flux disorder did not affect virus adsorption, but could inhibit virus entry into host cells and block EEs-LEs-lysosomes mediated trafficking of virions, leading to virions retention in endosomal compartments. The present studies suggest that cholesterol and endosomal vesicles synergistically participate in Nsp5 trafficking to promote PRRSV replication, which may provide new insights for the development of novel antiviral drugs targeting cholesterol metabolism pathways or the improvement of commercial vaccines.</div></div>","PeriodicalId":23551,"journal":{"name":"Veterinary microbiology","volume":"305 ","pages":"Article 110507"},"PeriodicalIF":2.7000,"publicationDate":"2025-04-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Veterinary microbiology","FirstCategoryId":"97","ListUrlMain":"https://www.sciencedirect.com/science/article/pii/S0378113525001427","RegionNum":2,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q3","JCRName":"MICROBIOLOGY","Score":null,"Total":0}
引用次数: 0
Abstract
Our previous studies showed that intracellular endosomal vesicles participated in PRRS virions trafficking in the early stage of viral infection, and cholesterol retention in endosomal vesicles disturbed viral replication via blocking PRRSV-endosomal vesicles membrane fusion. However, whether endosomal vesicles were associated with PRRSV protein(s) trafficking and the role of cholesterol in this process was still unclarity. In this study, we sought to elucidate the mechanism of cholesterol in endosomal vesicles-mediated viral protein transportation. The results showed that endosomal vesicles participated in trafficking of PRRSV Nsp5 protein. After being synthesized in endoplasmic reticulum (ER) and Golgi apparatus, Nsp5 was trafficked to early endosomes (EEs), but not endocytic recycling compartments (ERCs), then to late endosomes (LEs), and eventually reached lysosomal compartments, whereas disruption of cholesterol flux or LEs function led to the inability of Nsp5 arriving at lysosomes, where Nsp5 activated cellular autophagy to promote PRRSV replication. Molecular docking predictions revealed that cholesterol could form two hydrogen bonds with 74 alanine and 78 asparagine of Nsp5. After mutating the aforementioned binding sites, the replication efficiency of PRRSV decreased. Subsequently, the role of cholesterol in PRRSV replication was explored. Blocking of cholesterol flux significantly inhibited PRRSV replication. Single virus infection cycle analysis showed that cholesterol flux disorder did not affect virus adsorption, but could inhibit virus entry into host cells and block EEs-LEs-lysosomes mediated trafficking of virions, leading to virions retention in endosomal compartments. The present studies suggest that cholesterol and endosomal vesicles synergistically participate in Nsp5 trafficking to promote PRRSV replication, which may provide new insights for the development of novel antiviral drugs targeting cholesterol metabolism pathways or the improvement of commercial vaccines.
期刊介绍：
Veterinary Microbiology is concerned with microbial (bacterial, fungal, viral) diseases of domesticated vertebrate animals (livestock, companion animals, fur-bearing animals, game, poultry, fish) that supply food, other useful products or companionship. In addition, Microbial diseases of wild animals living in captivity, or as members of the feral fauna will also be considered if the infections are of interest because of their interrelation with humans (zoonoses) and/or domestic animals. Studies of antimicrobial resistance are also included, provided that the results represent a substantial advance in knowledge. Authors are strongly encouraged to read - prior to submission - the Editorials (''Scope or cope'' and ''Scope or cope II'') published previously in the journal. The Editors reserve the right to suggest submission to another journal for those papers which they feel would be more appropriate for consideration by that journal.
Original research papers of high quality and novelty on aspects of control, host response, molecular biology, pathogenesis, prevention, and treatment of microbial diseases of animals are published. Papers dealing primarily with immunology, epidemiology, molecular biology and antiviral or microbial agents will only be considered if they demonstrate a clear impact on a disease. Papers focusing solely on diagnostic techniques (such as another PCR protocol or ELISA) will not be published - focus should be on a microorganism and not on a particular technique. Papers only reporting microbial sequences, transcriptomics data, or proteomics data will not be considered unless the results represent a substantial advance in knowledge.
Drug trial papers will be considered if they have general application or significance. Papers on the identification of microorganisms will also be considered, but detailed taxonomic studies do not fall within the scope of the journal. Case reports will not be published, unless they have general application or contain novel aspects. Papers of geographically limited interest, which repeat what had been established elsewhere will not be considered. The readership of the journal is global.
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