{ "abstract": "Porcine reproductive and respiratory syndrome virus (PRRSV) suppresses innate antiviral immunity in porcine alveolar macrophages, and the PD-1/PD-L1 axis is thought to contribute to this immune dysregulation. This study investigated how PD-L1 knockout (KO) alters the host transcriptional response to PRRSV infection compared to wild-type (WT) immortalized porcine alveolar macrophages (iPAMs). PD-L1 KO iPAMs were generated by CRISPR/Cas9 targeting exon 2 of porcine CD274 locus, and knockout was confirmed in clone #35. After PRRSV-2 infection, KO#35 cells exhibited approximately 5-fold reduced viral RNA accumulation at 48 h post-infection compared with WT cells. Time-course RNA sequencing at 0, 6, and 48 h post-infection was then performed to characterize the transcriptional consequences of PD-L1 KO during acute PRRSV-2 infection. PD-L1 KO iPAMs exhibited downregulation of immune checkpoint and exhaustion-related genes (LAG3, PDCD1, TIGIT, TOX family) and NF-κB inhibitors (SOCS4, NFKBIA), alongside upregulation of antigen presentation (MHC I/II genes like SLA-2, SLA-5, B2M, SLA-DMA, SLA-DMB, HLA-DRA) and co-stimulatory molecules (CD80, CD86, CD40). Cytokine profiling indicated a reprogrammed response favoring antiviral signaling, including sustained elevation of CXCL10/11, ISGs (MX1/2, ISG15, IFIT1–3), and select interleukins (IL15, IL6), while suppressing pro-inflammatory TNF and CXCL8 across time points. Pathway enrichment analysis revealed significant enrichment of the cytokine–receptor interaction pathway. Weighted gene co-expression network analysis (WGCNA) further identified a key co-expression module containing 1,763 DEGs that was associated with innate immune activation. Moreover, protein-protein interaction (PPI) network analysis detected PXDN as a key hub gene connected to the PI3K-Akt signaling and natural killer cytotoxicity pathways. These findings provide transcriptomic insight into the association between PD-L1 deficiency and altered macrophage transcriptional responses to PRRSV infection in an in-vitro iPAM model and provide a basis for future mechanistic investigation of PD-L1 in PRRSV pathogenesis.", "abstract_shown": true, "affiliations": [ "Animal Biotechnology and Genomics Division, National Institute of Animal Science, Rural Development Administration, Wanju, Republic of Korea.", "College of Veterinary Medicine, Jeonbuk National University, Iksan, Republic of Korea.", "Division of Animal Diseases & Health, National Institute of Animal Science, Rural Development Administration, Wanju, Republic of Korea." ], "article_type": "Research Article", "arxiv": null, "author_total": 7, "authors": [ { "affs": [ 0 ], "name": "Sunirmal Sheet", "orcid": "0000-0001-6989-1292" }, { "affs": [ 1 ], "name": "Hwan-Ju Kim" }, { "affs": [ 1 ], "name": "Won-Il Kim" }, { "affs": [ 0 ], "name": "Seokho Kim" }, { "affs": [ 0 ], "name": "Poongyeon Lee" }, { "affs": [ 2 ], "name": "Ara Cho" }, { "affs": [ 0 ], "name": "Dahye Kim" } ], "bibtex": "https://research.draconitic.com/paper/fv0n60jn7rz47.bib", "cited_by": 0, "doi": "10.1371/journal.pone.0355142", "fulltext_url": "https://europepmc.org/articles/PMC13750538", "id": "fv0n60jn7rz47", "issns": [ "1932-6203" ], "landing_url": "https://doi.org/10.1371/journal.pone.0355142", "language": "eng", "licence": "CC-BY", "licence_label": "CC BY", "licence_source": "article", "licence_url": "https://creativecommons.org/licenses/by/4.0/", "metadata_licence": null, "native_subjects": [ [ "plos", "/Biology and life sciences/Cell biology/Signal transduction/Cell signaling/Membrane receptor signaling/Immune receptor signaling" ], [ "plos", "/Biology and life sciences/Computational biology/Genome analysis/Transcriptome analysis" ], [ "plos", "/Biology and life sciences/Developmental biology/Molecular development/Cytokines" ], [ "plos", "/Biology and life sciences/Genetics/Genomics/Genome analysis/Transcriptome analysis" ], [ "plos", "/Biology and life sciences/Immunology/Clinical immunology/Major histocompatibility complex" ], [ "plos", "/Biology and life sciences/Immunology/Immune response" ], [ "plos", "/Biology and life sciences/Immunology/Immune response/Antiviral immune response" ], [ "plos", "/Biology and life sciences/Immunology/Immune system/Innate immune system/Cytokines" ], [ "plos", "/Biology and life sciences/Immunology/Major histocompatibility complex" ], [ "plos", "/Biology and life sciences/Molecular biology/Molecular biology techniques/Cloning" ], [ "plos", "/Biology and life sciences/Physiology/Immune physiology/Cytokines" ], [ "plos", "/Medicine and health sciences/Clinical medicine/Clinical immunology/Major histocompatibility complex" ], [ "plos", "/Medicine and health sciences/Immunology/Clinical immunology/Major histocompatibility complex" ], [ "plos", "/Medicine and health sciences/Immunology/Immune response" ], [ "plos", "/Medicine and health sciences/Immunology/Immune response/Antiviral immune response" ], [ "plos", "/Medicine and health sciences/Immunology/Immune system/Innate immune system/Cytokines" ], [ "plos", "/Medicine and health sciences/Immunology/Major histocompatibility complex" ], [ "plos", "/Medicine and health sciences/Medical conditions/Infectious diseases/Respiratory infections" ], [ "plos", "/Medicine and health sciences/Medical conditions/Respiratory disorders/Respiratory infections" ], [ "plos", "/Medicine and health sciences/Pulmonology/Respiratory disorders/Respiratory infections" ], [ "plos", "/Research and analysis methods/Molecular biology techniques/Cloning" ] ], "openalex": null, "pdf_url": "https://europepmc.org/articles/PMC13750538?pdf=render", "pmcid": "PMC13750538", "pmid": "43008001", "published": "2026-11-10", "publisher": "Public Library of Science", "ris": "https://research.draconitic.com/paper/fv0n60jn7rz47.ris", "sources": [ { "first_seen": "2026-11-11T12:21:00Z", "native_id": "10.1371/journal.pone.0355142", "source": "plos", "url": "https://doi.org/10.1371/journal.pone.0355142" }, { "first_seen": "2026-11-12T15:21:47Z", "native_id": "MED:43008001", "source": "europepmc", "url": "https://europepmc.org/article/MED/43008001" } ], "subjects": [ { "field": "biology", "origin": "plos:immunology", "subfield": "immunology" } ], "title": "CRISPR/Cas9-mediated PD-L1 knockout reshapes temporal transcriptomic responses to PRRSV infection in porcine alveolar macrophages", "title_text": "CRISPR/Cas9-mediated PD-L1 knockout reshapes temporal transcriptomic responses to PRRSV infection in porcine alveolar macrophages", "type": "peer-reviewed", "updated_at": "2026-11-13T03:20:43Z", "url": "https://research.draconitic.com/paper/fv0n60jn7rz47", "venue": "PLOS One", "versions": [] }