{ "abstract": "Pseudorabies virus (PRV), an alphaherpesvirus, establishes lifelong latency in neurons and requires efficient reactivation for pathogenesis. Thymidine kinase (TK), encoded by UL23, is a key virulence factor essential for viral replication in non-dividing cells and for reactivation from latency; however, the regulatory mechanisms governing TK function remain poorly defined. Here, we identify TK as a substrate for small ubiquitin-like modifier (SUMO) modification and investigate its functional significance during PRV infection. We show that TK undergoes endogenous SUMOylation during viral infection is conjugated by SUMO-1, SUMO-2, and SUMO-3, with lysine residues K242 and K267 serving as the principal SUMO acceptor sites. Disruption of these SUMOylation sites does not affect TK stability and kinase activity but markedly alters its subcellular localization by reducing nuclear accumulation. Functionally, SUMOylation of TK is dispensable for viral replication in epithelial PK15 cells but facilitates efficient replication in neuronal N2a cells. Using an in vitro latency/reactivation model, we further show that SUMOylation-deficient TK does not affect the establishment of latent infection but markedly impairs viral reactivation and subsequent productive replication. In a mouse infection model, disruption of TK SUMOylation attenuates PRV pathogenicity, accompanied by reduced viral loads in the brain and milder histopathological lesions. Collectively, our findings demonstrate that SUMOylation regulates nuclear localization and biological function of PRV TK, thereby facilitating efficient neuronal replication, viral reactivation, and pathogenesis. These findings provide new insights into the post-translational regulation of alphaherpesvirus TK and identify TK SUMOylation as a potential target for controlling PRV infection. Author summary: Pseudorabies virus is an important alphaherpesvirus that causes major economic losses in the swine industry and and has recently emerged as an occasional zoonotic pathogen. A hallmark of PRV infection is its ability to establish lifelong latent infection neurons and subsequently reactivate, making viral eradication difficult. In this study, we found that the viral thymidine kinase (TK) protein is modified by a small ubiquitin-like modifier (SUMO) pathway during infection. This modification promotes nuclear accumulation of TK without affecting its kinase activity. Disruption of TK SUMOylation selectively impairs viral replication and reactivation in neuronal cells while having little effect in epithelial cells. Moreover, viruses carrying SUMOylation-deficient TK exhibit markedly reduced pathogenicity in mice. These findings reveal a role of SUMOylation in regulating PRV TK function and demonstrate how a host post-translational modification contributes to efficient neuronal infection and viral pathogenesis. Our study identifies TK SUMOylation as a potential target for developing antiviral strategies against PRV and related alphaherpesviruses.", "abstract_shown": true, "affiliations": [ "State Key Laboratory of Veterinary Public Health and Safety, College of Veterinary Medicine, China Agricultural University, Beijing, China", "Department of Basic Veterinary, College of Veterinary Medicine, China Agricultural University, Beijing, China", "Department of Preventive Veterinary, College of Veterinary Medicine, China Agricultural University, Beijing, China", "Key Laboratory of Applied Biotechnology on Animal Science & Veterinary Medicine of Zhejiang Province, College of Veterinary Medicine of Zhejiang A&F University, Hangzhou, Zhejiang Province, China", "Ministry of Education Key Laboratory for Animal Pathogens and Biosafety, College of Veterinary Medicine, Henan Agricultural University, Zhengzhou, Henan, China" ], "article_type": "Research Article", "arxiv": null, "author_total": 17, "authors": [ { "affs": [ 0 ], "name": "Xuan Chen" }, { "affs": [ 1 ], "name": "Chuang Li" }, { "affs": [ 2 ], "name": "Dongyue Xing" }, { "affs": [ 3 ], "name": "Jingjing Song" }, { "affs": [ 4 ], "name": "Yun Liu" }, { "name": "Yuxin Liu" }, { "name": "Qin Huang" }, { "name": "Yu Kuang" }, { "name": "Xian Li" }, { "name": "Jinlei Zhang" }, { "name": "Yuqi Zhang" }, { "name": "Nianzhi Zhang" }, { "name": "Rulan Bai" }, { "name": "Changyong Cheng" }, { "name": "Jin Yuan" }, { "name": "Jun Tang" }, { "name": "Rui Zhang" } ], "bibtex": "https://research.draconitic.com/paper/0p0pmsygbgshw.bib", "cited_by": 0, "doi": "10.1371/journal.ppat.1014653", "fulltext_url": "https://journals.plos.org/plospathogens/article?id=10.1371/journal.ppat.1014653", "id": "0p0pmsygbgshw", "issns": [ "1553-7374" ], "landing_url": "https://journals.plos.org/plospathogens/article?id=10.1371/journal.ppat.1014653", "language": null, "licence": "CC-BY-4.0", "licence_label": "CC BY 4.0", "licence_source": "journal", "licence_url": "https://creativecommons.org/licenses/by/4.0/", "metadata_licence": null, "native_subjects": [ [ "openalex", "2713|Herpesvirus Infections and Treatments" ], [ "plos", "/Biology and life sciences/Biochemistry/Proteins/Amino acids/Basic amino acids/Lysine" ], [ "plos", "/Biology and life sciences/Biochemistry/Proteins/Post-translational modification/SUMOylation" ], [ "plos", "/Biology and life sciences/Genetics/Genomics/Microbial genomics/Viral genomics/Viral genome" ], [ "plos", "/Biology and life sciences/Microbiology/Microbial genomics/Viral genomics/Viral genome" ], [ "plos", "/Biology and life sciences/Microbiology/Virology/Viral genomics/Viral genome" ], [ "plos", "/Biology and life sciences/Microbiology/Virology/Viral persistence and latency" ], [ "plos", "/Biology and life sciences/Microbiology/Virology/Viral replication" ], [ "plos", "/Biology and life sciences/Molecular biology/Molecular biology techniques/DNA construction/Plasmid construction" ], [ "plos", "/Biology and life sciences/Molecular biology/Molecular biology techniques/Molecular probe techniques/Immunoblotting" ], [ "plos", "/Physical sciences/Chemistry/Chemical compounds/Organic compounds/Amino acids/Basic amino acids/Lysine" ], [ "plos", "/Physical sciences/Chemistry/Organic chemistry/Organic compounds/Amino acids/Basic amino acids/Lysine" ], [ "plos", "/Research and analysis methods/Molecular biology techniques/DNA construction/Plasmid construction" ], [ "plos", "/Research and analysis methods/Molecular biology techniques/Molecular probe techniques/Immunoblotting" ], [ "plos", "/Research and analysis methods/Precipitation techniques/Immunoprecipitation" ] ], "openalex": "W7220449882", "pdf_url": "https://journals.plos.org/plospathogens/article/file?id=10.1371/journal.ppat.1014653&type=printable", "pmcid": null, "pmid": null, "published": "2026-10-05", "publisher": "Public Library of Science", "ris": "https://research.draconitic.com/paper/0p0pmsygbgshw.ris", "sources": [ { "first_seen": "2026-10-30T12:17:37Z", "native_id": "10.1371/journal.ppat.1014653", "source": "plos", "url": "https://journals.plos.org/plospathogens/article?id=10.1371/journal.ppat.1014653" } ], "subjects": [ { "field": "biochemistry", "origin": "plos:molecular biology", "subfield": "molecular-biology" }, { "field": "biology", "origin": "plos:microbiology", "subfield": "microbiology" } ], "title": "SUMOylation of pseudorabies virus thymidine kinase regulates its nuclear localization and facilitates neuronal replication and reactivation", "title_text": "SUMOylation of pseudorabies virus thymidine kinase regulates its nuclear localization and facilitates neuronal replication and reactivation", "type": "peer-reviewed", "updated_at": "2026-10-30T12:17:37Z", "url": "https://research.draconitic.com/paper/0p0pmsygbgshw", "venue": "PLOS Pathogens", "versions": [] }