{ "abstract": "African swine fever virus (ASFV), a large double-stranded DNA virus, poses one of the most significant infectious disease threats to the global swine industry. ASFV transcription is particularly complex because of its unique host-independent transcription system. Using nanopore sequencing technology, we accurately identified the viral transcription start and termination sites. Combining direct RNA sequencing with experimental verification strongly supports the presence of long multicistronic RNAs. By combining this information with the viral genome annotation, we identified six novel transcriptional genes with potential functions. Furthermore, we observed transcriptional “read-through” events, in which transcription initiated at a common start site continues beyond typical termination sites, producing multiple RNA transcripts of different lengths. Additionally, via direct RNA nanopore sequencing, we identified m6A and m5C methylation modifications on viral transcripts, which are predominantly distributed at both termini of the viral genome. Compared to unmodified transcripts, modified RNAs have shorter poly(A) tails, indicating a correlation between RNA modification and poly(A) tail length. Further investigation of ASFV transcriptional architecture, “read-through” events, RNA modifications, and candidate genes may improve our understanding of the viral life cycle and pathogenicity and inform the identification of potential antiviral targets. Author summary: African swine fever virus (ASFV) causes a highly lethal disease in pigs, posing a devastating threat to the global agricultural economy. Despite its impact, how this large DNA virus orchestrates the production of its proteins during infection has remained incompletely understood. In this study, we utilized advanced sequencing technologies to comprehensively map the genetic blueprint—or “transcriptional architecture”—of ASFV. We discovered 26 potential new viral genes and revealed that the virus frequently uses an unexpectedly efficient mechanism known as “multicistronic transcription”. This indicates that a single viral RNA molecule can carry the instructions to sequentially produce multiple distinct proteins. By engineering specialized recombinant viruses with fluorescent tags, we provided strong evidence that these complex transcripts are actively translated. Furthermore, we directly detected chemical modifications (m6A and m5C) on the viral RNAs, which are concentrated in genomic regions associated with host immune evasion. Ultimately, our findings fundamentally reshape our understanding of ASFV gene expression, providing a vital foundation for discovering new antiviral targets and rationally designing vaccines.", "abstract_shown": true, "affiliations": [ "National Key Laboratory of Agricultural Microbiology, Hainan Research Institute, Hubei Hongshan Laboratory, College of Animal Sciences & Technology and College of Veterinary Medicine, Huazhong Agricultural University, Wuhan, Hubei, China", "Key Laboratory of Preventive Veterinary Medicine in Hubei Province, The Cooperative Innovation Centre for Sustainable Pig Production, Wuhan, China", "Virology and Cell Technology Research Team, National Center for Genetic Engineering and Biotechnology (BIOTEC), National Science and Technology Development Agency (NSTDA), Pathum Thani, Thailand", "Hubei Jiangxia Laboratory, Wuhan, Hubei, China" ], "article_type": "Research Article", "arxiv": null, "author_total": 14, "authors": [ { "affs": [ 0 ], "name": "Guiqian Wang" }, { "affs": [ 1 ], "name": "Pengfei Li" }, { "affs": [ 2 ], "name": "Xinglin He" }, { "affs": [ 3 ], "name": "Hua Cao" }, { "name": "Ahmed H. Ghonaim" }, { "name": "Shengnan Ruan" }, { "name": "Xuexiang Yu" }, { "name": "Jiaru Zhou" }, { "name": "Hongmei Zhu" }, { "name": "Mengjia Zhang" }, { "name": "Anan Jongkaewwattana" }, { "name": "Guiqing Peng" }, { "name": "Qigai He" }, { "name": "Wentao Li" } ], "bibtex": "https://research.draconitic.com/paper/3m8rrejcfbgk8.bib", "cited_by": 0, "doi": "10.1371/journal.ppat.1014617", "fulltext_url": "https://journals.plos.org/plospathogens/article?id=10.1371/journal.ppat.1014617", "id": "3m8rrejcfbgk8", "issns": [ "1553-7374" ], "landing_url": "https://journals.plos.org/plospathogens/article?id=10.1371/journal.ppat.1014617", "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", "1102|Animal Disease Management and Epidemiology" ], [ "plos", "/Biology and life sciences/Biochemistry/Proteins/Recombinant proteins" ], [ "plos", "/Biology and life sciences/Genetics/Gene expression/DNA transcription" ], [ "plos", "/Biology and life sciences/Genetics/Gene expression/Viral gene expression" ], [ "plos", "/Biology and life sciences/Genetics/Genomics" ], [ "plos", "/Biology and life sciences/Genetics/Genomics/Microbial genomics/Viral genomics" ], [ "plos", "/Biology and life sciences/Genetics/Microbial genetics/Viral genetics/Viral gene expression" ], [ "plos", "/Biology and life sciences/Microbiology/Microbial genomics/Viral genomics" ], [ "plos", "/Biology and life sciences/Microbiology/Virology/Viral genetics/Viral gene expression" ], [ "plos", "/Biology and life sciences/Microbiology/Virology/Viral genomics" ], [ "plos", "/Biology and life sciences/Molecular biology/Molecular biology techniques/Artificial gene amplification and extension/Polymerase chain reaction" ], [ "plos", "/Biology and life sciences/Molecular biology/Molecular biology techniques/Sequencing techniques/RNA sequencing" ], [ "plos", "/Research and analysis methods/Database and informatics methods/Bioinformatics/Sequence analysis/Sequence alignment" ], [ "plos", "/Research and analysis methods/Molecular biology techniques/Artificial gene amplification and extension/Polymerase chain reaction" ], [ "plos", "/Research and analysis methods/Molecular biology techniques/Sequencing techniques/RNA sequencing" ] ], "openalex": "W7214084218", "pdf_url": "https://journals.plos.org/plospathogens/article/file?id=10.1371/journal.ppat.1014617&type=printable", "pmcid": null, "pmid": null, "published": "2026-09-23", "publisher": "Public Library of Science", "ris": "https://research.draconitic.com/paper/3m8rrejcfbgk8.ris", "sources": [ { "first_seen": "2026-10-30T12:17:43Z", "native_id": "10.1371/journal.ppat.1014617", "source": "plos", "url": "https://journals.plos.org/plospathogens/article?id=10.1371/journal.ppat.1014617" } ], "subjects": [ { "field": "biochemistry", "origin": "plos:gene expression", "subfield": "molecular-biology" }, { "field": "biology", "origin": "plos:genomics", "subfield": "genetics-genomics" }, { "field": "biology", "origin": "plos:microbiology", "subfield": "microbiology" } ], "title": "Transcriptional architecture of African swine fever virus", "title_text": "Transcriptional architecture of African swine fever virus", "type": "peer-reviewed", "updated_at": "2026-10-30T12:17:43Z", "url": "https://research.draconitic.com/paper/3m8rrejcfbgk8", "venue": "PLOS Pathogens", "versions": [] }