Interferon-γ Interferes with Transforming Growth Factor-β Signaling through Direct Interaction of YB-1 with Smad3

Abstract

Transforming growth factor-␤ (TGF-␤) and interferon-␥ (IFN-␥Physical interaction between Smad3 and YB-1 was demonstrated by immunoprecipitation-Western blot analyses, and electrophoretic mobility shift assays using the recombinant Smad3 and YB-1 proteins indicated that YB-1 forms a complex with Smad3 bound to the Smadbinding element. Glutathione S-transferase pull-down assays showed that YB-1 binds to the MH1 domain of Smad3, whereas the central and carboxyl-terminal regions of YB-1 were required for its interaction with Smad3. YB-1 also interferes with the Smad3-p300 interaction by its preferential binding to p300. Altogether, the results provide a novel insight into the mechanism by which IFN-␥/YB-1 counteracts TGF-␤/Smad3. They also indicate that IFN-␥/YB-1 inhibits COL1A2 transcription by dual actions: via the IFN-␥ response element and through a cross-talk with the TGF-␤/Smad signaling pathway.

Transforming Growth Factor β1-mediated Activation of the Smooth Muscle α-Actin Gene in Human Pulmonary Myofibroblasts Is Inhibited by Tumor Necrosis Factor-α via Mitogen-activated Protein Kinase Kinase 1-dependent Induction of the Egr-1 Transcriptional Repressor

“…In nuclear extracts prepared from hPFBs treated with TNF-␣ alone (c), binding of YB-1 repressor protein to the reverse strand of DNA encompassing the THR region of the SM␣A promoter increased by ϳ60% over baseline levels (p Ͻ 0.05). type I collagen promoter by TGF␤1 through the formation of a sequestering complex with Smad3 (Higashi et al, 2003). In addition, our previous studies on hPFBs showed that TGF␤1 displaced YB-1 from the reverse strand of THR site in the SM␣A promoter (Zhang et al, 2005).…”

“…Similarly, mononuclear cell release of IFN␥, a T-helper 1 cytokine that often functions in tandem with TNF-␣, suppresses fibroblast proliferation and collagen deposition and has long been associated with classic type I, innate-immune responses (Kunkel et al, 2003). Notably, the antifibrotic activity of IFN␥ seems to be partly based on its ability to functionally sequester TGF␤1 receptor-regulated Smad3 protein as a heteromeric complex with the YB-1 transcriptional repressor protein thus preventing binding of activated Smads to their cognate CAGA cis-activation sequence motifs in the type I collagen promoter (Higashi et al, 2003;Dooley et al, 2006). Aside from this one mechanistic detail, the molecular pathways governing termination of TGF␤1-mediated myofibroblast differentiation are not well understood but theoretically important in the search for new therapeutic targets to prevent chronic fibroproliferative diseases of the heart, lung, liver, and kidney.…”

“…Transforming growth factor (TGF) ␤1) is the principle mediator of myofibroblast differentiation in healing wounds where it accumulates in granulation tissue in activated form during leukocyte infiltration and potently stimulates transcription of the type I␣2 collagen subunit (COL1␣2) and SM␣A genes (Becker et al, 2000;Massague and Wotton, 2000;Cogan et al, 2002;Higashi et al, 2003;Grotendorst et al, 2004;Leask and Abraham, 2004;Subramanian et al, 2004;Zhang et al, 2005). Smad proteins 2 and 3 are activated by TGF␤1 receptor engagement and enter the nucleus in which they directly bind SM␣A and COL1␣2 promoter DNA.…”

“…Smad proteins 2 and 3 are activated by TGF␤1 receptor engagement and enter the nucleus in which they directly bind SM␣A and COL1␣2 promoter DNA. Smad3 also reportedly forms additional off-promoter complexes with DNA-binding repressor proteins to govern transcriptional output of the COL1␣2 gene in TGF␤1-activated myofibroblasts (Higashi et al, 2003). Poor termination of TGF␤1-mediated myofibroblast differentiation could contribute to chronic fibroproliferative disease and excessive scar formation in the injured lung, heart, liver, kidney, and skin potentially causing pathobiologic phenomena referred to as "endless healing" (Tomasek et al, 2002).…”

Increased Expression of Y-Box-Binding Protein-1 in Hind-Limb Muscles During Regeneration from Ischemic Injury in Mice

“…In addition, several genes important for wound healing and cellular proliferation and survival are regulated by YB-1, including collagen alpha1 and alpha2 (Norman et al 2001;Higashi et al 2003), matrix metalloproteinase 2 (Mertens et al 1997), the B-chain isoform of plateletderived growth factor (Stenina et al 2000), vascular endothelial growth factor (Coles et al 2004), granulocyte-macrophage-colony stimulating factor (Coles et al 2000), and Fas death receptor (Lasham et al 2000); moreover, YB-1 can also regulate αSMA and human pulmonary myofibroblasts (Zhang et al 2005). Takahashi et al (2010) reported that YB-1 is increased in the angiogenic endothelial cells of various tumors and is involved in the growth of endothelial cells.…”