Format

Send to

Choose Destination
J Pharmacol Exp Ther. 2018 Apr;365(1):156-164. doi: 10.1124/jpet.117.244483. Epub 2018 Feb 9.

Flavopiridol Inhibits TGF-β-Stimulated Biglycan Synthesis by Blocking Linker Region Phosphorylation and Nuclear Translocation of Smad2.

Author information

1
Kulliyyah of Allied Health Sciences, International Islamic University Malaysia, Kuantan, Pahang, Malaysia (M.A.R.); School of Health and Biomedical Sciences, RMIT University, Bundoora, Victoria, Australia (A.S., T.J.P., R.G., N.O., P.J.L.); School of Pharmacy, Pharmacy Australia Centre of Excellence, The University of Queensland, Woolloongabba, Queensland, Australia (D.K., P.M., P.J.L.); Department of Pharmacy, Xinhua College of Sun Yat-sen University, Tianhe, Guangzhou, China (Y.C., P.J.L.); Faculty of Health Sciences, University of Macau, Taipa, Macau, China (W.Z.); and Monash University, Departments of Medicine and Immunology, Central and Eastern Clinical School, Alfred Health, Melbourne, Victoria, Australia (N.O.).
2
Kulliyyah of Allied Health Sciences, International Islamic University Malaysia, Kuantan, Pahang, Malaysia (M.A.R.); School of Health and Biomedical Sciences, RMIT University, Bundoora, Victoria, Australia (A.S., T.J.P., R.G., N.O., P.J.L.); School of Pharmacy, Pharmacy Australia Centre of Excellence, The University of Queensland, Woolloongabba, Queensland, Australia (D.K., P.M., P.J.L.); Department of Pharmacy, Xinhua College of Sun Yat-sen University, Tianhe, Guangzhou, China (Y.C., P.J.L.); Faculty of Health Sciences, University of Macau, Taipa, Macau, China (W.Z.); and Monash University, Departments of Medicine and Immunology, Central and Eastern Clinical School, Alfred Health, Melbourne, Victoria, Australia (N.O.) P.little@uq.edu.au.

Abstract

Transforming growth factor-β (TGF-β) is a pleiotropic growth factor implicated in the development of atherosclerosis for its role in mediating glycosaminoglycan (GAG) chain hyperelongation on the proteoglycan biglycan, a phenomenon that increases the binding of atherogenic lipoproteins in the vessel wall. Phosphorylation of the transcription factor Smad has emerged as a critical step in the signaling pathways that control the synthesis of biglycan, both the core protein and the GAG chains. We have used flavopiridol, a well-known cyclin-dependent kinase inhibitor, to study the role of linker region phosphorylation in the TGF-β-stimulated synthesis of biglycan. We used radiosulfate incorporation and SDS-PAGE to assess proteoglycan synthesis, real-time polymerase chain reaction to assess gene expression, and chromatin immunoprecipitation to assess the binding of Smads to the promoter region of GAG Synthesizing genes. Flavopiridol blocked TGF-β-stimulated synthesis of mRNA for the GAG synthesizing enzymes, and chondroitin 4-sulfotransferase (C4ST-1), chondroitin sulfate synthase-1 (ChSy-1) and TGF-β-mediated proteoglycans synthesis as well as GAG hyperelongation. Flavopiridol blocked TGF-β-stimulated Smad2 phosphorylation at both the serine triplet and the isolated threonine residue in the linker region. The binding of Smad to the promoter region of the C4ST-1 and ChSy-1 genes was stimulated by TGF-β, and this response was blocked by flavopiridol, demonstrating that linker region phosphorylated Smad can pass to the nucleus and positively regulate transcription. These results demonstrate the validity of the kinases, which phosphorylate the Smad linker region as potential therapeutic target(s) for the development of an agent to prevent atherosclerosis.

PMID:
29438988
DOI:
10.1124/jpet.117.244483

Supplemental Content

Full text links

Icon for HighWire
Loading ...
Support Center