title: Inducible STAT3 NH2 terminal mono-ubiquitination promotes BRD4 complex formation to regulate apoptosis.
abstract: Signal Transducers and Activator of Transcription-3 (STAT3) are latent transcription factors that are regulated by post-translational modifications (PTMs) in response to cellular activation by the IL-6 superfamily of cytokines to regulate cell cycle progression and/or apoptosis. Here we observe that STAT3 is inducibly mono-ubiquitinated and investigate its consequences. Using domain mapping and highly specific selected reaction monitoring-mass spectrometric assays, we identify lysine (K) 97 in its NH2-terminal domain as the major mono-ubiquitin conjugation site. We constructed a mono-ubiquitinated mimic consisting of a deubiquitinase-resistant monomeric ubiquitin fused to the NH2 terminus of STAT3 (ubiquitinated-STAT3 FP). In complex assays of ectopically expressed ubi-STAT3-FP, we observed enhanced complex formation with bromodomain-containing protein 4 (BRD4), a component of the activated positive transcriptional elongation factor (P-TEFb) complex. Chromatin immunoprecipitation experiments in STAT3(+/-) and STAT3(-/-) MEFs showed BRD4 recruitment to STAT3-dependent suppressor of cytokine signaling-3 gene (SOCS3). The effect of a selective small molecule inhibitor of BRD4, JQ1, to inhibit SOCS3 expression demonstrated the functional role of BRD4 for STAT3-dependent transcription. Additionally, ectopic ubiquitinated-STAT3 FP expression upregulated BCL2, BCL2L1, APEX1, SOD2, CCND1 and MYC expression indicating the role of ubiquitinated STAT3 in anti-apoptosis and cellular proliferation. Finally we observed that ubiquitinated-STAT3 FP suppressed TNFα-induced apoptotic cell death, indicating the functional importance of mono-ubiquitinated STAT3 in antiapoptotic gene expression. We conclude that STAT3 mono-ubiquitination is a key trigger in BRD4-dependent antiapoptotic and pro-proliferative gene expression programs. Thus, inhibiting the STAT3 mono-ubiquitination-BRD4 pathway may be a novel therapeutic target for the treatment of STAT3-dependent proliferative diseases.
authors.affiliation: Sealy Center for Molecular Medicine, University of Texas Medical Branch, Galveston, TX 77555, United States; Department of Internal Medicine, University of Texas Medical Branch, Galveston, TX 77555, United States.
authors.forename: Sutapa
authors.initials: S
authors.lastname: Ray
authors.affiliation: Sealy Center for Molecular Medicine, University of Texas Medical Branch, Galveston, TX 77555, United States; Department of Internal Medicine, University of Texas Medical Branch, Galveston, TX 77555, United States; Institute for Translational Sciences, University of Texas Medical Branch, Galveston, TX 77555, United States.
authors.forename: Yingxin
authors.initials: Y
authors.lastname: Zhao
authors.affiliation: Institute for Translational Sciences, University of Texas Medical Branch, Galveston, TX 77555, United States.
authors.forename: Mohammad
authors.initials: M
authors.lastname: Jamaluddin
authors.affiliation: Department of Internal Medicine, University of Texas Medical Branch, Galveston, TX 77555, United States.
authors.forename: Chukwudi B
authors.initials: CB
authors.lastname: Edeh
authors.affiliation: Department of Internal Medicine, University of Texas Medical Branch, Galveston, TX 77555, United States.
authors.forename: Chang
authors.initials: C
authors.lastname: Lee
authors.affiliation: Sealy Center for Molecular Medicine, University of Texas Medical Branch, Galveston, TX 77555, United States; Department of Internal Medicine, University of Texas Medical Branch, Galveston, TX 77555, United States; Institute for Translational Sciences, University of Texas Medical Branch, Galveston, TX 77555, United States. Electronic address: arbrasie@utmb.edu.
authors.forename: Allan R
authors.initials: AR
authors.lastname: Brasier
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000235:genetics*; D001381:Azepines / Q000494:pharmacology; D002460:Cell Line; D049109:Cell Proliferation / Q000235:genetics; D057809:HEK293 Cells; D056945:Hep G2 Cells; D006801:Humans; D051379:Mice; D009687:Nuclear Proteins / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000235:genetics; D050796:STAT3 Transcription Factor / Q000096:biosynthesis / Q000235:genetics*; D015398:Signal Transduction; D000071223:Suppressor of Cytokine Signaling 3 Protein; D050826:Suppressor of Cytokine Signaling Proteins / Q000096:biosynthesis; D014157:Transcription Factors / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000378:metabolism*; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D014230:Triazoles / Q000494:pharmacology; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism; D025801:Ubiquitin / Q000378:metabolism; D054875:Ubiquitination; D000092468:Bromodomain Containing Proteins
chemical_list: D001381:Azepines; D009687:Nuclear Proteins; D011993:Recombinant Fusion Proteins; D050796:STAT3 Transcription Factor; D000071223:Suppressor of Cytokine Signaling 3 Protein; D050826:Suppressor of Cytokine Signaling Proteins; D014157:Transcription Factors; D014230:Triazoles; D014409:Tumor Necrosis Factor-alpha; D025801:Ubiquitin; C561695:(+)-JQ1 compound; C486694:Brd4 protein, mouse; D000092468:Bromodomain Containing Proteins; C491411:Socs3 protein, mouse; C494087:Stat3 protein, mouse
keywords: Apoptosis; BRD4; Mono-ubiquitination; Proliferation; STAT3