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Here, we perform a genome-wide CRISPR knockout screen to identify host restriction factors for SARS-CoV-2, revealing DAZAP2 as a potent antiviral gene. DAZAP2, previously implicated in SARS-CoV-2 restriction, is found to inhibit viral entry by blocking virion fusion with both endolysosomal and plasma membranes. Additionally, DAZAP2 suppresses genomic RNA replication without affecting the primary translation of viral replicases. We demonstrate that DAZAP2 functions as a pan-coronavirus restriction factor across four genera of coronaviruses. Importantly, knockout of\n                      <jats:italic toggle=\"yes\">DAZAP2<\/jats:italic>\n                      enhances SARS-CoV-2 infection in mouse models and in human primary airway epithelial cells, confirming its physiological relevance. Mechanistically, the antiviral activity of DAZAP2 appears to be indirect, potentially through the regulation of host gene expression, as it primarily localizes to the nucleus. Our findings provide new insights into the host defense system against coronaviruses and highlight DAZAP2 as a potential target for host-directed antiviral therapies.\n                    <\/jats:p>\n                    <jats:sec>\n                      <jats:title>IMPORTANCE<\/jats:title>\n                      <jats:p>\n                        During viral infection, the host defense response is mediated by a variety of host factors through distinct mechanisms that have yet to be fully elucidated. Although\n                        <jats:italic toggle=\"yes\">DAZAP2<\/jats:italic>\n                        was previously implicated in SARS-CoV-2 restriction, its mechanisms of action and\n                        <jats:italic toggle=\"yes\">in vivo<\/jats:italic>\n                        relevance remain unclear. In this study, we identify DAZAP2 as a potent pan-coronavirus restriction factor that inhibits viral infection through dual mechanisms: blocking virion fusion with both endolysosomal and plasma membranes, and suppressing genomic RNA replication. We confirm its physiological relevance in host defense using mouse models and primary cell cultures. This study advances our understanding of host-pathogen interactions. Targeting DAZAP2 or its regulatory pathways could provide a new approach to enhance host defense against current and future coronavirus threats.\n                      <\/jats:p>\n                    <\/jats:sec>\n                  <\/jats:sec>","DOI":"10.1128\/mbio.00385-25","type":"journal-article","created":{"date-parts":[[2025,8,20]],"date-time":"2025-08-20T13:01:40Z","timestamp":1755694900000},"update-policy":"https:\/\/doi.org\/10.1128\/asmj-crossmark-policy-page","source":"Crossref","is-referenced-by-count":3,"title":["DAZAP2 functions as a pan-coronavirus restriction factor by inhibiting viral entry and genomic replication"],"prefix":"10.1128","volume":"16","author":[{"ORCID":"https:\/\/orcid.org\/0009-0002-7647-861X","authenticated-orcid":false,"given":"Fei","family":"Feng","sequence":"first","affiliation":[{"id":[{"id":"https:\/\/ror.org\/013q1eq08","id-type":"ROR","asserted-by":"publisher"}],"name":"Key Laboratory of Medical Molecular Virology (MOE\/NHC\/CAMS), Shanghai 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