title: Thrombin stimulation of proteoglycan synthesis in vascular smooth muscle is mediated by protease-activated receptor-1 transactivation of the transforming growth factor beta type I receptor.
abstract: Growth factors modify the structure of the glycosaminoglycan (GAG) chains on biglycan leading to enhanced LDL binding. G-protein receptor-coupled agonists such as thrombin, signal changes the structure of proteoglycans produced by vascular smooth muscle cells (VSMCs). One component of classical G-protein-coupled receptor (GPCR) signaling invokes transactivation of protein tyrosine kinase receptors such as the epidermal growth factor receptor. Serine/threonine receptor growth factors such as transforming growth factor-(TGF)-beta are potent activators of proteoglycan synthesis. We have used the model of proteoglycan synthesis to demonstrate that the signaling paradigm of GPCR signaling can be extended to include the transactivation of serine/threonine receptor, specifically the TGF-beta type I receptor (TbetaRI) also known as activin-like kinase (ALK) V. Thrombin stimulated elongation of GAG chains and increased proteoglycan core protein expression and these responses were blocked by the TbetaRI antagonist, SB431542 and TbetaRI siRNA knockdown, as well as several protease-activated receptor (PAR)-1 antagonists. The canonical downstream response to TGF-beta is increased C-terminal phosphorylation of the transcription factor Smad2 generating phospho-Smad2C (phosphorylation of Smad2 C-terminal region). Thrombin stimulated increased phospho-Smad2C levels, and the response was blocked by SB431542 and JNJ5177094. The proteolytically inactive thrombin mimetic thrombin-receptor activating peptide also stimulated an increase in cytosolic phospho-Smad2C. Signaling pathways for growth factor regulated proteoglycan synthesis represent therapeutic targets for the prevention of atherosclerosis, but the novel finding of a GPCR-mediated transactivation of a serine/threonine growth factor receptor almost certainly has implications well beyond the synthesis of proteoglycans.
authors.affiliation: Diabetes and Cell Biology Laboratory, BakerIDI Heart and Diabetes Institute, Melbourne, 3004 Victoria, Australia; Departments of Medicine and Immunology, Central and Eastern Clinical School, Alfred Hospital, Monash University, Melbourne, 3004 Victoria, Australia.
authors.forename: Micah L
authors.initials: ML
authors.lastname: Burch
authors.affiliation: Diabetes and Cell Biology Laboratory, BakerIDI Heart and Diabetes Institute, Melbourne, 3004 Victoria, Australia.
authors.forename: Mandy L
authors.initials: ML
authors.lastname: Ballinger
authors.affiliation: Diabetes and Cell Biology Laboratory, BakerIDI Heart and Diabetes Institute, Melbourne, 3004 Victoria, Australia.
authors.forename: Sundy N Y
authors.initials: SNY
authors.lastname: Yang
authors.affiliation: Diabetes and Cell Biology Laboratory, BakerIDI Heart and Diabetes Institute, Melbourne, 3004 Victoria, Australia.
authors.forename: Robel
authors.initials: R
authors.lastname: Getachew
authors.affiliation: Departments of Biochemistry & Molecular Biology and Anatomy & Developmental Biology, Monash University, Clayton, Victoria 3800, Australia.
authors.forename: Catherine
authors.initials: C
authors.lastname: Itman
authors.affiliation: Departments of Biochemistry & Molecular Biology and Anatomy & Developmental Biology, Monash University, Clayton, Victoria 3800, Australia.
authors.forename: Kate
authors.initials: K
authors.lastname: Loveland
authors.affiliation: Diabetes and Cell Biology Laboratory, BakerIDI Heart and Diabetes Institute, Melbourne, 3004 Victoria, Australia; Departments of Medicine and Immunology, Central and Eastern Clinical School, Alfred Hospital, Monash University, Melbourne, 3004 Victoria, Australia.
authors.forename: Narin
authors.initials: N
authors.lastname: Osman
authors.affiliation: Diabetes and Cell Biology Laboratory, BakerIDI Heart and Diabetes Institute, Melbourne, 3004 Victoria, Australia; Departments of Medicine and Immunology, Central and Eastern Clinical School, Alfred Hospital, Monash University, Melbourne, 3004 Victoria, Australia. Electronic address: peter.little@bakeridi.edu.au.
authors.forename: Peter J
authors.initials: PJ
authors.lastname: Little
mesh_terms: D001549:Benzamides / Q000494:pharmacology; D002478:Cells, Cultured; D004149:Dioxoles / Q000494:pharmacology; D066246:ErbB Receptors / Q000378:metabolism; D006801:Humans; D008077:Lipoproteins, LDL / Q000378:metabolism; D009130:Muscle, Smooth / Q000378:metabolism*; D032389:Myocytes, Smooth Muscle / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects / Q000502:physiology; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011509:Proteoglycans / Q000096:biosynthesis*; D034741:RNA, Small Interfering / Q000494:pharmacology; D044463:Receptor, PAR-1 / Q000378:metabolism*; D000077293:Receptor, Transforming Growth Factor-beta Type I; D018125:Receptors, Transforming Growth Factor beta / Q000037:antagonists & inhibitors / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*; D051899:Smad2 Protein / Q000378:metabolism; D013917:Thrombin / Q000378:metabolism* / Q000494:pharmacology; D015533:Transcriptional Activation / Q000187:drug effects / Q000502:physiology*; D016212:Transforming Growth Factor beta / Q000378:metabolism
chemical_list: C459179:4-(5-benzo(1,3)dioxol-5-yl-4-pyridin-2-yl-1H-imidazol-2-yl)benzamide; D001549:Benzamides; D004149:Dioxoles; D008077:Lipoproteins, LDL; D011509:Proteoglycans; D034741:RNA, Small Interfering; D044463:Receptor, PAR-1; D018125:Receptors, Transforming Growth Factor beta; C497782:SMAD2 protein, human; D051899:Smad2 Protein; D016212:Transforming Growth Factor beta; C512478:EGFR protein, human; D066246:ErbB Receptors; D017346:Protein Serine-Threonine Kinases; D000077293:Receptor, Transforming Growth Factor-beta Type I; D013917:Thrombin