{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,11]],"date-time":"2026-09-11T15:25:16Z","timestamp":1789140316458,"version":"build-2803163510"},"reference-count":32,"publisher":"Wiley","issue":"1","license":[{"start":{"date-parts":[[2026,5,25]],"date-time":"2026-05-25T00:00:00Z","timestamp":1779667200000},"content-version":"vor","delay-in-days":144,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"},{"start":{"date-parts":[[2026,1,1]],"date-time":"2026-01-01T00:00:00Z","timestamp":1767225600000},"content-version":"tdm","delay-in-days":0,"URL":"http:\/\/doi.wiley.com\/10.1002\/tdm_license_1.1"}],"funder":[{"DOI":"10.13039\/100022963","name":"Key Research and Development Program of Zhejiang Province","doi-asserted-by":"publisher","award":["2025C02065"],"award-info":[{"award-number":["2025C02065"]}],"id":[{"id":"10.13039\/100022963","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001809","name":"National Natural Science Foundation of China","doi-asserted-by":"publisher","award":["81701988"],"award-info":[{"award-number":["81701988"]}],"id":[{"id":"10.13039\/501100001809","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["onlinelibrary.wiley.com"],"crossmark-restriction":true},"short-container-title":["Human Mutation"],"published-print":{"date-parts":[[2026,1]]},"abstract":"<jats:sec>\n                    <jats:title>Background<\/jats:title>\n                    <jats:p>Immunotherapy efficacy in colorectal cancer (CRC) is largely restricted to microsatellite instability\u2010high (MSI\u2010H) tumors, highlighting an urgent need to overcome resistance in microsatellite\u2010stable (MSS) CRC. Mitochondrial DNA (mtDNA) leakage activates the cGAS\u2010STING pathway, a potent inducer of antitumor immunity. However, endogenous regulators constraining mtDNA release in CRC, particularly within the inner mitochondrial membrane (IMM), remain poorly defined.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Methods<\/jats:title>\n                    <jats:p>\n                      In CRC cohorts from TCGA, integrated bioinformatics analysis identified dysregulated mitochondria\u2010associated genes exhibiting significant differential expression and survival outcomes. Phospholipid Scramblase 3 (PLSCR3) emerged as the prime candidate. Functional validation employed siRNA knockdown and CRISPR\/Cas9 knockout in human (HT29) and mouse (CT26) CRC cell lines. Mitochondrial integrity was evaluated via JC\u20101 membrane potential assay, oxygen consumption rate (OCR) measurement, and cytosolic mtDNA quantification. cGAS\u2010STING activation was measured by 2\u2009\n                      <jats:sup>\u2032<\/jats:sup>\n                      3\u2009\n                      <jats:sup>\u2032<\/jats:sup>\n                      \u2010cGAMP ELISA, cytokine (IFN\n                      <jats:italic>\u03b2<\/jats:italic>\n                      and CXCL10) secretion, immunoblotting (p\u2010STING and ISGs), and RT\u2010qPCR. Immune cell cytotoxicity was assessed using ex vivo NK cell killing assays. In vivo antitumor immunity and response to anti\u2010PD\u20101 therapy were evaluated in syngeneic CT26 graft models in BALB\/c mice, with tumor\u2010infiltrating lymphocytes analyzed by flow cytometry.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Results<\/jats:title>\n                    <jats:p>\n                      PLSCR3 was selected for further study because it fulfilled three criteria simultaneously: differential expression in CRC, significant association with survival in univariate analysis, and established mitochondrial localization. PLSCR3 deficiency disrupted mitochondrial integrity, causing membrane depolarization, impaired respiration, and significant cytosolic mtDNA accumulation. This mtDNA leakage robustly activated the cGAS\u2010STING pathway, evidenced by increased 2\u2009\n                      <jats:sup>\u2032<\/jats:sup>\n                      3\u2009\n                      <jats:sup>\u2032<\/jats:sup>\n                      \u2010cGAMP, IFN\n                      <jats:italic>\u03b2<\/jats:italic>\n                      \/CXCL10 secretion, STING phosphorylation (Ser366), and ISG upregulation. Functionally, PLSCR3 knockdown enhanced NK cell\u2013mediated killing of CRC cells in vitro. Critically, PLSCR3 knockout in CT26 tumors significantly potentiated the efficacy of anti\u2010PD\u20101 therapy in vivo, overcoming inherent resistance in this MSS\u2010CRC model. This was associated with increased intratumoral infiltration of CD8+ and CD4+ T cells, elevated Granzyme B levels, and enhanced activation markers on CD8+ T cells within the tumor microenvironment.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Conclusion<\/jats:title>\n                    <jats:p>Our study identifies PLSCR3 as a previously unrecognized negative regulator of mtDNA\u2010associated cGAS\u2010STING activation in CRC. By maintaining mitochondrial homeostasis and limiting mtDNA leakage, PLSCR3 constrains innate immune signaling and reduces sensitivity to anti\u2010PD\u20101 therapy in the CT26 model. These findings establish PLSCR3 as a promising therapeutic target to enhance immunotherapy responses in CRC.<\/jats:p>\n                  <\/jats:sec>","DOI":"10.1155\/humu\/8545428","type":"journal-article","created":{"date-parts":[[2026,5,26]],"date-time":"2026-05-26T05:57:37Z","timestamp":1779775057000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":1,"title":["PLSCR3 Deficiency Triggers mtDNA\u2010Driven cGAS\u2010STING Activation to Potentiate Antitumor Immunity in Colorectal 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