RACK1 binds to Smad3 to modulate transforming growth factor-β1- stimulated α2(I) collagen transcription in renal tubular epithelial cells

Kazuhiro Okano, H. William Schnaper*, Karol Bomsztyk, Tomoko Hayashida

*Corresponding author for this work

Research output: Contribution to journalArticle

16 Scopus citations

Abstract

Although it is clear that transforming growth factor-β1 (TGF-β1) is critical for renal fibrogenesis, the complexity of the involved mechanisms is increasingly apparent. TGF-β1 stimulates phosphorylation of Smad2/3 and activates other signaling molecules as well. The molecular link between these other kinases and Smads is not known. We sought new binding partners for Smad3 in renal cells and identified receptor for activated protein kinase C 1 (RACK1) as a novel binding partner of Smad3. The linker region of Smad3 and the tryptophan-aspartic acid repeat 6 and 7 of RACK1 are sufficient for the association. RACK1 also interacts with Smad3 in the human kidney epithelial cell line, HKC. Silencing RACK1 increases transcriptional activity of TGF-β1-responsive promoter sequences of the Smad binding element (SBE), p3TP-Lux, and α2(I) collagen. Conversely, overexpressed RACK1 negatively modulates α2(I) collagen transcriptional activity in TGF-β1- stimulated cells. RACK1 did not affect phosphorylation of Smad3 at the C terminus or in the linker region. However, RACK1 reduced direct binding of Smad3 to the SBE motif. Mutating a RACK1 tyrosine at residue 246, but not at 228, decreased the inhibitory effect of RACK1 on both α2(I) collagen promoter activity and Smad binding to SBE induced by TGF-β1. These results suggest that RACK1 modulates transcription of α2(I) collagen by TGF-β1 through interference with Smad3 binding to the gene promoter.

Original languageEnglish (US)
Pages (from-to)26196-26204
Number of pages9
JournalJournal of Biological Chemistry
Volume281
Issue number36
DOIs
StatePublished - Sep 8 2006

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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