{"doi":"10.1128/jvi.01864-25","title":"Phosphorylation of Shiftless by casein kinase 1 δ/ε is required for its antiviral activity","abstract":"<jats:title>ABSTRACT</jats:title>\n                  <jats:sec>\n                    <jats:title/>\n                    <jats:p>Shiftless (SHFL) is a host antiviral factor that inhibits the viral −1 programmed ribosomal frameshifting (−1PRF). How the antiviral activity of SHFL is regulated is not known. Here, we report that the phosphorylation of SHFL by the casein kinase 1 delta (CK1δ) and epsilon (CK1ε) regulates its antiviral activity. The casein kinases were identified as SHFL-interacting proteins. Downregulation of the expression of the endogenous casein kinases or pharmacological inhibition of kinase activity significantly impaired the activity of SHFL to inhibit HIV-1 -1PRF and viral production. The residues T250 and T253 were identified as critical phosphorylation sites; substitution of either of the residues with alanine, resulting in the mutants SHFL-T250A or SHFL-T253A, abolished the antiviral activity of SHFL against HIV-1. The T250A mutation did not affect SHFL interaction with target RNA or the ribosomal proteins uL5 and eS31 but significantly reduced its interaction with IGF2BP proteins. Furthermore, SHFL-T250A displayed reduced association with the actively translating polysomes. These results are consistent with the notion that SHFL interaction with ribosomes is critical for its activity to inhibit −1PRF. Taken together, our results indicate that phosphorylation of SHFL by CK1δ/ε is required for its antiviral activity.</jats:p>\n                    <jats:sec>\n                      <jats:title>IMPORTANCE</jats:title>\n                      <jats:p>Innate immune responses and antiviral defense mechanisms are regulated through a variety of mechanisms, among which post-translational modifications, especially phosphorylation, play important roles. Deciphering the regulatory mechanisms helps understand the innate immune responses more comprehensively. SHFL is an interferon-stimulated host antiviral factor that inhibits the replication of multiple viruses. The present study revealed that phosphorylation of SHFL by casein kinase 1 δ/ε is required for its antiviral activity against HIV-1. These results provide an additional example for how immune responses are regulated. Furthermore, given that SHFL inhibits −1PRF, the present studies provide tools for further exploring the mechanisms of −1PRF.</jats:p>\n                    </jats:sec>\n                  </jats:sec>","journal":"Journal of Virology","year":2025,"id":631610,"datarank":0.0,"base_score":0.0,"endowment":0.0,"self_citation_contribution":0.0,"citation_network_contribution":0.0,"self_endowment_contribution":0.0,"citer_contribution":0.0,"corpus_percentile":null,"corpus_rank":null,"citation_count":0,"citer_count":0,"citers_with_citation_signal":0,"citers_with_endowment":0,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":null,"is_data_producer":false,"deposit_databanks":null,"is_oa":false,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":null,"fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":1636906,"name":"Shaozu Fu","orcid":null,"position":1,"is_corresponding":false},{"id":1636907,"name":"Xinlu Wang","orcid":"0000-0001-8887-0550","position":2,"is_corresponding":false},{"id":605866,"name":"Guangxia Gao","orcid":"0000-0001-5284-0472","position":3,"is_corresponding":false},{"id":1041180,"name":"Yongle Wang","orcid":"0009-0001-7108-3902","position":0,"is_corresponding":false}],"reference_count":0,"raw_metadata":{"has_enrichment":true,"resolved":true,"title":"Phosphorylation of Shiftless by casein kinase 1 δ/ε is required for its antiviral activity","abstract":"<jats:title>ABSTRACT</jats:title>\n                  <jats:sec>\n                    <jats:title/>\n                    <jats:p>Shiftless (SHFL) is a host antiviral factor that inhibits the viral −1 programmed ribosomal frameshifting (−1PRF). How the antiviral activity of SHFL is regulated is not known. Here, we report that the phosphorylation of SHFL by the casein kinase 1 delta (CK1δ) and epsilon (CK1ε) regulates its antiviral activity. The casein kinases were identified as SHFL-interacting proteins. Downregulation of the expression of the endogenous casein kinases or pharmacological inhibition of kinase activity significantly impaired the activity of SHFL to inhibit HIV-1 -1PRF and viral production. The residues T250 and T253 were identified as critical phosphorylation sites; substitution of either of the residues with alanine, resulting in the mutants SHFL-T250A or SHFL-T253A, abolished the antiviral activity of SHFL against HIV-1. The T250A mutation did not affect SHFL interaction with target RNA or the ribosomal proteins uL5 and eS31 but significantly reduced its interaction with IGF2BP proteins. Furthermore, SHFL-T250A displayed reduced association with the actively translating polysomes. These results are consistent with the notion that SHFL interaction with ribosomes is critical for its activity to inhibit −1PRF. Taken together, our results indicate that phosphorylation of SHFL by CK1δ/ε is required for its antiviral activity.</jats:p>\n                    <jats:sec>\n                      <jats:title>IMPORTANCE</jats:title>\n                      <jats:p>Innate immune responses and antiviral defense mechanisms are regulated through a variety of mechanisms, among which post-translational modifications, especially phosphorylation, play important roles. Deciphering the regulatory mechanisms helps understand the innate immune responses more comprehensively. SHFL is an interferon-stimulated host antiviral factor that inhibits the replication of multiple viruses. The present study revealed that phosphorylation of SHFL by casein kinase 1 δ/ε is required for its antiviral activity against HIV-1. These results provide an additional example for how immune responses are regulated. Furthermore, given that SHFL inhibits −1PRF, the present studies provide tools for further exploring the mechanisms of −1PRF.</jats:p>\n                    </jats:sec>\n                  </jats:sec>","is_dataset_classified":null,"base_score":0.0,"endowment":0.0,"datacite_reuse_total":0,"file_count":0,"downloads":0,"views":0,"has_version_chain":false,"is_dataset":false,"is_oa":false,"pmid":"41329002","pmcid":"PMC12724340","openalex_id":null,"authors":[],"funders":[],"total_grants":0,"fwci":null,"citation_percentile":null,"influential_citations":0,"citation_trend":[],"oa_status":"gold","license":"cc-by","oa_locations":[{"url":"https://doi.org/10.1128/jvi.01864-25","host_type":"publisher"},{"url":"https://journals.asm.org/doi/pdf/10.1128/jvi.01864-25","host_type":"publisher"},{"url":"https://doaj.org/article/0d1bed1b50f845c8a4390820324ee561","host_type":"repository"},{"url":"https://pmc.ncbi.nlm.nih.gov/articles/PMC12724340/","host_type":"repository"},{"url":"https://europepmc.org/articles/PMC12724340","host_type":"Europe_PMC"},{"url":"https://europepmc.org/articles/PMC12724340?pdf=render","host_type":"Europe_PMC"}],"fields_of_study":[],"mesh_terms":["Humans","HIV-1","HIV Infections","Casein Kinase Idelta","RNA-Binding Proteins","Antiviral Agents","Virus Replication","Phosphorylation","HEK293 Cells","Casein Kinase 1 epsilon"],"keywords":["Phosphorylation","Casein Kinase 1","Antiviral Factor","Shiftless","-1 Programmed Ribosomal Frameshifting"],"sdg_mappings":[],"linked_datasets":[],"clinical_trials":[],"software_tools":[],"database_accessions":[{"name":"refseq"}],"source":"live","citation_network_status":"fetched"},"created_at":"2026-08-06T00:11:55.971733Z","pmid":null,"pmcid":null,"fwci":null,"citation_percentile":null,"influential_citations":0,"oa_status":null,"license":null,"views":0,"total_file_size_bytes":0,"version_count":0,"fair_f":null,"fair_a":null,"fair_i":null,"fair_r":null,"fair_zscore":null,"fair_rationale":null,"fair_model":null,"fair_agent_version":null,"fair_fulltext_source":null,"fair_has_llm":null,"fair_computed_at":null,"clinical_trials":[],"software_tools":[],"db_accessions":[],"linked_datasets":[],"topics":[]}