title: Calyculin A reveals serine/threonine phosphatase protein phosphatase 1 as a regulatory nodal point in canonical signal transducer and activator of transcription 3 signaling of human microvascular endothelial cells.
abstract: Vascular inflammation is initiated by stimuli acting on endothelial cells. A clinical feature of vascular inflammation is increased circulating interleukin 6 (IL-6) type cytokines such as leukemia inhibitory factor (LIF), but their role in vascular inflammation is not fully defined. IL-6 type cytokines activate transcription factor signal transducer and activator of transcription 3 (STAT3), which has a key role in inflammation and the innate immune response. Canonical STAT3 gene induction is due to phosphorylation of (1) Y705, leading to STAT3 dimerization and DNA binding and (2) S727, enhancing homodimerization and DNA binding by recruiting p300/CBP. We asked whether enhancing S727 STAT3 phosphorylation using the protein phosphatase 1 (PP1) inhibitor, calyculin A, would enhance LIF-induced gene expression in human microvascular endothelial cells (HMEC-1). Cotreatment with calyculin A and LIF markedly increased STAT3 S727 phosphorylation, without affecting the increase in the nuclear fraction of STAT3 phosphorylated on Y705. PP2A inhibitors, okadaic acid and fostriecin, did not enhance STAT3 S727 phosphorylation. Surprisingly, calyculin A eliminated LIF-induced gene expression: (1) calyculin A reduced binding of nuclear extracts to a STAT3 consensus site, thereby reducing the overall level of binding observed with LIF; and (2) calyculin A caused p300/CBP phosphorylation, thus resulting in reduced acetylation activity and degradation. Together, these findings reveal a pivotal role of a protein serine/threonine phosphatases that is likely PP1 in HMEC in controlling STAT3 transcriptional activity.
authors.affiliation: Department of Pharmacology and Toxicology, Center for Excellence in Cardiovascular-Renal Research, School of Medicine, The University of Mississippi Medical Center, Jackson, Mississippi 39216-4505, USA.
authors.forename: Carlos
authors.initials: C
authors.lastname: Zgheib
authors.forename: Fouad A
authors.initials: FA
authors.lastname: Zouein
authors.forename: Rony
authors.initials: R
authors.lastname: Chidiac
authors.forename: Mazen
authors.initials: M
authors.lastname: Kurdi
authors.forename: George W
authors.initials: GW
authors.lastname: Booz
mesh_terms: D050882:CREB-Binding Protein; D002273:Carcinogens / Q000494:pharmacology*; D002460:Cell Line; D004727:Endothelium / Q000166:cytology / Q000378:metabolism*; D006801:Humans; D015850:Interleukin-6 / Q000378:metabolism; D053605:Leukemia Inhibitory Factor / Q000378:metabolism / Q000494:pharmacology; D008387:Marine Toxins; D055806:Microvessels / Q000166:cytology / Q000378:metabolism*; D010080:Oxazoles / Q000494:pharmacology*; D010766:Phosphorylation / Q000187:drug effects; D055503:Protein Multimerization / Q000187:drug effects; D054645:Protein Phosphatase 1 / Q000037:antagonists & inhibitors / Q000378:metabolism*; D054648:Protein Phosphatase 2 / Q000378:metabolism; D050796:STAT3 Transcription Factor / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects*; D014158:Transcription, Genetic / Q000187:drug effects
chemical_list: D002273:Carcinogens; C508600:IL6 protein, human; D015850:Interleukin-6; C507837:LIF protein, human; D053605:Leukemia Inhibitory Factor; D008387:Marine Toxins; D010080:Oxazoles; D050796:STAT3 Transcription Factor; C494086:STAT3 protein, human; C059041:calyculin A; D050882:CREB-Binding Protein; C494363:CREBBP protein, human; D054645:Protein Phosphatase 1; D054648:Protein Phosphatase 2