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Olfactomedin 2, a novel regulator for transforming growth factor-β-induced smooth muscle differentiation of human embryonic stem cell-derived mesenchymal cells.

Shi N, Guo X, Chen SY - Mol. Biol. Cell (2014)

Bottom Line: Olfm2 also inhibited HERP1 expression.Moreover, blockade of Olfm2 expression inhibited TGF-β-induced SRF binding to SM gene promoters in a chromatin setting, whereas overexpression of Olfm2 dose dependently enhanced SRF binding.These results demonstrate that Olfm2 mediates TGF-β-induced SM gene transcription by empowering SRF binding to CArG box in SM gene promoters.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology and Pharmacology, University of Georgia, Athens, GA 30602.

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Olfm2 physically interacted with SRF. (A, C) CoIP with endogenous proteins indicated that Olfm2 physically interacted with SRF. Serum-starved hES-MCs were treated with vehicle (–) or TGF-β (+; 1 ng/ml) for 24 h. Cell lysates were immunoprecipitated with normal IgG, SRF (A), or Olfm2 (C) antibody. The immunoprecipitates were blotted (IB) with Olfm2 (A) or SRF (C) antibody. The interaction between Olfm2 and SRF was enhanced by TGF-β induction. (B, D) Quantifications of Olfm2 immunoprecipitated with SRF (B) and of SRF immunoprecipitated with Olfm2 (D) as shown in A and C, respectively. The proteins coimmunoprecipitated were normalized to α-tubulin. *p < 0.01 compared with IgG-immunoprecipitated group. #p < 0.01 compared with SRF-immunoprecipitated (B) and Olfm2- immunoprecipitated (D) groups with vehicle treatment (n = 3).
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Figure 5: Olfm2 physically interacted with SRF. (A, C) CoIP with endogenous proteins indicated that Olfm2 physically interacted with SRF. Serum-starved hES-MCs were treated with vehicle (–) or TGF-β (+; 1 ng/ml) for 24 h. Cell lysates were immunoprecipitated with normal IgG, SRF (A), or Olfm2 (C) antibody. The immunoprecipitates were blotted (IB) with Olfm2 (A) or SRF (C) antibody. The interaction between Olfm2 and SRF was enhanced by TGF-β induction. (B, D) Quantifications of Olfm2 immunoprecipitated with SRF (B) and of SRF immunoprecipitated with Olfm2 (D) as shown in A and C, respectively. The proteins coimmunoprecipitated were normalized to α-tubulin. *p < 0.01 compared with IgG-immunoprecipitated group. #p < 0.01 compared with SRF-immunoprecipitated (B) and Olfm2- immunoprecipitated (D) groups with vehicle treatment (n = 3).

Mentions: Because Olfm2 function in SM differentiation relies on SRF/CArG, we sought to determine whether Olfm2 interacts with SRF. A coimmunoprecipitation (CoIP) assay using endogenous proteins extracted from hES-MCs treated with vehicle or TGF-β showed that Olfm2 was coimmunoprecipitated with SRF (Figure 5A) and SRF was coimmunoprecipitated with Olfm2 (Figure 5C), demonstrating that Olfm2 physically interacted with SRF in hES-MCs. Their interaction was significantly enhanced by TGF-β, which was likely attributable to the increased expression of Olfm2 by TGF-β (Figure 5, A–D).


Olfactomedin 2, a novel regulator for transforming growth factor-β-induced smooth muscle differentiation of human embryonic stem cell-derived mesenchymal cells.

Shi N, Guo X, Chen SY - Mol. Biol. Cell (2014)

Olfm2 physically interacted with SRF. (A, C) CoIP with endogenous proteins indicated that Olfm2 physically interacted with SRF. Serum-starved hES-MCs were treated with vehicle (–) or TGF-β (+; 1 ng/ml) for 24 h. Cell lysates were immunoprecipitated with normal IgG, SRF (A), or Olfm2 (C) antibody. The immunoprecipitates were blotted (IB) with Olfm2 (A) or SRF (C) antibody. The interaction between Olfm2 and SRF was enhanced by TGF-β induction. (B, D) Quantifications of Olfm2 immunoprecipitated with SRF (B) and of SRF immunoprecipitated with Olfm2 (D) as shown in A and C, respectively. The proteins coimmunoprecipitated were normalized to α-tubulin. *p < 0.01 compared with IgG-immunoprecipitated group. #p < 0.01 compared with SRF-immunoprecipitated (B) and Olfm2- immunoprecipitated (D) groups with vehicle treatment (n = 3).
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Related In: Results  -  Collection

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Figure 5: Olfm2 physically interacted with SRF. (A, C) CoIP with endogenous proteins indicated that Olfm2 physically interacted with SRF. Serum-starved hES-MCs were treated with vehicle (–) or TGF-β (+; 1 ng/ml) for 24 h. Cell lysates were immunoprecipitated with normal IgG, SRF (A), or Olfm2 (C) antibody. The immunoprecipitates were blotted (IB) with Olfm2 (A) or SRF (C) antibody. The interaction between Olfm2 and SRF was enhanced by TGF-β induction. (B, D) Quantifications of Olfm2 immunoprecipitated with SRF (B) and of SRF immunoprecipitated with Olfm2 (D) as shown in A and C, respectively. The proteins coimmunoprecipitated were normalized to α-tubulin. *p < 0.01 compared with IgG-immunoprecipitated group. #p < 0.01 compared with SRF-immunoprecipitated (B) and Olfm2- immunoprecipitated (D) groups with vehicle treatment (n = 3).
Mentions: Because Olfm2 function in SM differentiation relies on SRF/CArG, we sought to determine whether Olfm2 interacts with SRF. A coimmunoprecipitation (CoIP) assay using endogenous proteins extracted from hES-MCs treated with vehicle or TGF-β showed that Olfm2 was coimmunoprecipitated with SRF (Figure 5A) and SRF was coimmunoprecipitated with Olfm2 (Figure 5C), demonstrating that Olfm2 physically interacted with SRF in hES-MCs. Their interaction was significantly enhanced by TGF-β, which was likely attributable to the increased expression of Olfm2 by TGF-β (Figure 5, A–D).

Bottom Line: Olfm2 also inhibited HERP1 expression.Moreover, blockade of Olfm2 expression inhibited TGF-β-induced SRF binding to SM gene promoters in a chromatin setting, whereas overexpression of Olfm2 dose dependently enhanced SRF binding.These results demonstrate that Olfm2 mediates TGF-β-induced SM gene transcription by empowering SRF binding to CArG box in SM gene promoters.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology and Pharmacology, University of Georgia, Athens, GA 30602.

Show MeSH
Related in: MedlinePlus