biostudies-literatureLin YBeijing Research Center for Respiratory Infections Diseases ProjectMOST | National Key Research and Development Program of ChinaCAMS Innovation Fund for Medical ScienceBeijing Municipal Administration of Hospitals Incubating ProgramReform and Development of Beijing Municipal Health CommissionMOST | National Natural Science Foundation of Chinae202503320https://www.ebi.ac.uk/biostudies/studies/S-EPMC12179656biostudies-literatureUnknown8(9)Human adenovirus B7 (HAdV-B7) is a significant respiratory pathogen in children, associated with substantial morbidity and mortality. Despite its clinical importance, the molecular mechanisms of HAdV-B7-host interaction remain poorly elucidated. In this study, we performed a high-throughput gain-of-function cDNA library screening and identified glycogen synthase kinase 3α (GSK3A) as a key proviral factor facilitating HAdV-B7 replication. The overexpression of GSK3A enhanced viral replication, whereas knockdown or knockout inhibited it. Furthermore, the kinase-active S21A mutant significantly augmented viral replication, whereas the kinase-inactive Y279A and K148A mutants of GSK3A failed to support it, highlighting the importance of its kinase activity on HAdV-B7 replication. Notably, phospLife science allianceGSK3A promotes human adenovirus replication and phosphorylates viral L4-22K protein.PMC12179656PX2024043BJRID2025-00832470141,820722662019-I2M-5-0262023YFC2306001NoneZhu YJing LLei XXie ZXiao XLin YChen YfalseGSK3A promotes human adenovirus replication and phosphorylates viral L4-22K protein.Human adenovirus B7 (HAdV-B7) is a significant respiratory pathogen in children, associated with substantial morbidity and mortality. Despite its clinical importance, the molecular mechanisms of HAdV-B7-host interaction remain poorly elucidated. In this study, we performed a high-throughput gain-of-function cDNA library screening and identified glycogen synthase kinase 3α (GSK3A) as a key proviral factor facilitating HAdV-B7 replication. The overexpression of GSK3A enhanced viral replication, whereas knockdown or knockout inhibited it. Furthermore, the kinase-active S21A mutant significantly augmented viral replication, whereas the kinase-inactive Y279A and K148A mutants of GSK3A failed to support it, highlighting the importance of its kinase activity on HAdV-B7 replication. Notably, phosp2025-01-01T00:00:00Z2025 Sep2026-06-01T23:22:20.426Z2026-05-23T03:08:23.671ZS-EPMC121796564053728510.26508/lsa.202503320