{"doi":"10.1152/ajpcell.00053.2003","title":"Phosphorylation of threonine 276 in Smad4 is involved in transforming  growth factor-β-induced nuclear accumulation","abstract":"<jats:p> Smad4, the common Smad, is central for transforming growth factor (TGF)-β superfamily ligand signaling. Smad4 has been shown to be constitutively phosphorylated (Nakao A, Imamura T, Souchelnytskyi S, Kawabata M, Ishisaki A, Oeda E, Tamaki K, Hanai J, Heldin C-H, Miyazono K, and ten Dijke P. EMBO J 16: 5353-5362, 1997), but the site(s) of phosphorylation, the kinase(s) that performs this phosphorylation, and the significance of the phosphorylation of Smad4 are currently unknown. This report describes the identification of a consensus ERK phosphorylation site in the linker region of Smad4 at Thr<jats:sup>276</jats:sup>. Our data show that ERK can phosphorylate Smad4 in vitro but not Smad4 with mutated Thr<jats:sup>276</jats:sup>. Flag-tagged Smad4-T276A mutant protein accumulates less efficiently in the nucleus after stimulation by TGF-β and is less efficient in generating a transcriptional response than Smad4 wild-type protein. Tryptic phosphopeptide mapping identified a phosphopeptide in Smad4 wild-type protein that was absent in phosphorylated Smad4-T276A mutant protein. Our results suggest that MAP kinase can phosphorylate Thr<jats:sup>276</jats:sup> of Smad4 and that phosphorylation can lead to enhanced TGF-β-induced nuclear accumulation and, as a consequence, enhanced transcriptional activity of Smad4. </jats:p>","journal":"American Journal of Physiology-Cell Physiology","year":2003,"id":655326,"datarank":0.6307038929086449,"base_score":4.204692619390966,"endowment":4.204692619390966,"self_citation_contribution":0.6307038929086449,"citation_network_contribution":0.0,"self_endowment_contribution":0.6307038929086449,"citer_contribution":0.0,"corpus_percentile":null,"corpus_rank":null,"citation_count":66,"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":739241,"name":"Ori S. Cohen","orcid":null,"position":1,"is_corresponding":false},{"id":1710645,"name":"Malay K. Raychowdhury","orcid":null,"position":2,"is_corresponding":false},{"id":1710646,"name":"Deborah N. Chadee","orcid":null,"position":3,"is_corresponding":false},{"id":832049,"name":"Ying Zhang","orcid":"0000-0003-2194-7116","position":4,"is_corresponding":false},{"id":1710647,"name":"John M. Kyriakis","orcid":null,"position":5,"is_corresponding":false},{"id":1710648,"name":"Alessandro A. Alessandrini","orcid":null,"position":6,"is_corresponding":false},{"id":185493,"name":"Herbert Y. Lin","orcid":null,"position":7,"is_corresponding":false},{"id":1710644,"name":"Bernard A. J. Roelen","orcid":null,"position":0,"is_corresponding":false}],"reference_count":0,"raw_metadata":{"has_enrichment":true,"resolved":true,"title":"Phosphorylation of threonine 276 in Smad4 is involved in transforming  growth factor-β-induced nuclear accumulation","abstract":"<jats:p> Smad4, the common Smad, is central for transforming growth factor (TGF)-β superfamily ligand signaling. Smad4 has been shown to be constitutively phosphorylated (Nakao A, Imamura T, Souchelnytskyi S, Kawabata M, Ishisaki A, Oeda E, Tamaki K, Hanai J, Heldin C-H, Miyazono K, and ten Dijke P. EMBO J 16: 5353-5362, 1997), but the site(s) of phosphorylation, the kinase(s) that performs this phosphorylation, and the significance of the phosphorylation of Smad4 are currently unknown. This report describes the identification of a consensus ERK phosphorylation site in the linker region of Smad4 at Thr<jats:sup>276</jats:sup>. Our data show that ERK can phosphorylate Smad4 in vitro but not Smad4 with mutated Thr<jats:sup>276</jats:sup>. Flag-tagged Smad4-T276A mutant protein accumulates less efficiently in the nucleus after stimulation by TGF-β and is less efficient in generating a transcriptional response than Smad4 wild-type protein. Tryptic phosphopeptide mapping identified a phosphopeptide in Smad4 wild-type protein that was absent in phosphorylated Smad4-T276A mutant protein. Our results suggest that MAP kinase can phosphorylate Thr<jats:sup>276</jats:sup> of Smad4 and that phosphorylation can lead to enhanced TGF-β-induced nuclear accumulation and, as a consequence, enhanced transcriptional activity of Smad4. </jats:p>","is_dataset_classified":null,"base_score":4.204692619390966,"endowment":4.204692619390966,"datacite_reuse_total":0,"file_count":0,"downloads":0,"views":0,"has_version_chain":false,"is_dataset":false,"is_oa":false,"pmid":"12801888","pmcid":null,"openalex_id":"https://openalex.org/W2102095720","authors":[],"funders":[{"funder_name":"NIDDK NIH HHS","grant_id":"DK-02716","title":null},{"funder_name":"NIDDK NIH HHS","grant_id":"DK-19406","title":null}],"total_grants":2,"fwci":0.868,"citation_percentile":0.7104159,"influential_citations":0,"citation_trend":[{"year":2012,"count":3},{"year":2013,"count":2},{"year":2014,"count":2},{"year":2015,"count":2},{"year":2016,"count":2},{"year":2017,"count":2},{"year":2018,"count":4},{"year":2019,"count":2},{"year":2020,"count":4},{"year":2021,"count":1},{"year":2022,"count":4},{"year":2023,"count":2},{"year":2024,"count":4},{"year":2025,"count":2}],"oa_status":"closed","license":null,"oa_locations":[{"url":"https://journals.physiology.org/doi/pdf/10.1152/ajpcell.00053.2003","host_type":"publisher"},{"url":"https://doi.org/10.1152/ajpcell.00053.2003","host_type":"journal"},{"url":"https://pubmed.ncbi.nlm.nih.gov/12801888","host_type":"repository"}],"fields_of_study":["TGF-β signaling in diseases","Genetic factors in colorectal cancer","Cancer-related gene regulation"],"mesh_terms":["Amino Acid Sequence","Animals","Cell Line","Cell Nucleus","DNA-Binding Proteins","Genetic Linkage","Molecular Sequence Data","Mutation","Phosphoproteins","Phosphorylation","Swine","Threonine","Transcription, Genetic","Trans-Activators","Transforming Growth Factor beta","Consensus Sequence","Protein Structure, Tertiary","LLC-PK1 Cells","Mitogen-Activated Protein Kinases"],"keywords":["Phosphopeptide","Phosphorylation","SMAD","MAPK/ERK pathway","Kinase","Mutant","Threonine","Cell biology","Biology","Protein phosphorylation","Protein kinase A","Nuclear localization sequence","Molecular biology","Serine","Biochemistry","Nucleus","Gene"],"sdg_mappings":[],"linked_datasets":[],"clinical_trials":[],"software_tools":[],"database_accessions":[],"source":"live","citation_network_status":"fetched"},"created_at":"2026-08-11T10:27:05.415467Z","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":[]}