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Heat shock protein 70 enhances vascular bone morphogenetic protein-4 signaling by binding matrix Gla protein.

ABSTRACT: Matrix Gla protein (MGP) is a calcification inhibitor, which binds and inhibits bone morphogenetic protein (BMP)-2 and -4.The objective was to determine whether MGP also binds other proteins, which could interfere with its function.We transfected bovine aortic endothelial cells with N-terminally FLAG-tagged MGP and used immunoprecipitation and liquid chromatographic-tandem mass spectrometric analysis to identify MGP-binding proteins. Heat shock protein (HSP)70, a stress-induced protein expressed in atherosclerotic lesions and soluble in serum, was identified as a novel MGP-binding protein. The interaction between MGP and HSP70 was confirmed by coimmunoprecipitation and chemical crosslinking, and blocked the interaction between MGP and BMP-4. In endothelial cells, HSP70 enhanced BMP-4-induced proliferation and tube formation, and in calcifying vascular cells, HSP70 enhanced BMP-induced calcium deposition. In addition, HSP70 mediated the procalcific effect of interleukin-6 on calcifying vascular cells. In apolipoprotein E-null mice, a model for atherosclerosis, levels of BMP-4, HSP70, MGP, and interleukin-6 were elevated in the aortic wall. Levels of BMP-4, HSP70, and interleukin-6 were also elevated in serum, and anti-HSP70 antibodies diminished its procalcific effect on calcifying vascular cells.HSP70 binds MGP and enhances BMP activity, thereby functioning as a potential link between cellular stress, inflammation, and BMP signaling.

SUBMITTER: Yao Y

PROVIDER: S-EPMC2779117 | biostudies-other | 2009 Sep

REPOSITORIES: biostudies-other

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Heat shock protein 70 enhances vascular bone morphogenetic protein-4 signaling by binding matrix Gla protein.

Yao Yucheng Y Watson Andrew D AD Ji Sheng S Boström Kristina I KI

Circulation research 20090806 6

<h4>Rationale</h4>Matrix Gla protein (MGP) is a calcification inhibitor, which binds and inhibits bone morphogenetic protein (BMP)-2 and -4.<h4>Objective</h4>The objective was to determine whether MGP also binds other proteins, which could interfere with its function.<h4>Methods and results</h4>We transfected bovine aortic endothelial cells with N-terminally FLAG-tagged MGP and used immunoprecipitation and liquid chromatographic-tandem mass spectrometric analysis to identify MGP-binding proteins ...[more]

PMID: 19661459

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Project description:Matrix Gla protein (MGP), a modulator of the BMP-SMAD signals, inhibits arterial calcification in a Glu γ-carboxylation dependent manner but the role of MGP highly expressed in a subset of bone marrow (BM) mesenchymal stem/stromal cells is unknown. Here we provide evidence that MGP might be a niche factor for both normal and malignant myelopoiesis. When mouse BM hematopoietic cells were cocultured with mitomycin C-treated BM stromal cells in the presence of anti-MGP antibody, growth of hematopoietic cells was reduced by half, and maintenance of long-term culture-initiating cells (LTC-ICs) was profoundly attenuated. Antibody-mediated blockage of MGP also inhibited growth (by a fifth) and cobblestone formation (by half) of stroma-dependent MB-1 myeloblastoma cells. MGP was undetectable in normal hematopoietic cells but was expressed in various mesenchymal cells and was aberrantly high in MB-1 cells. MGP and bone morphogenetic protein (BMP)-4 were co-induced in stromal cells cocultured with both normal hematopoietic cells and MB-1 myeloblastoma cells in an oscillating several days-periodic manner. BMP-2 was also induced in stromal cells cocultured with normal hematopoietic cells but was barely expressed when cocultured with MB-1 cells. GST-pulldown and luciferase reporter assays showed that uncarboxylated MGP interacted with BMP-4 and that anti-MGP antibody abolished this interaction. LDN-193189, a selective BMP signaling inhibitor, inhibited growth and cobblestone formation of MB-1 cells. The addition of warfarin, a selective inhibitor of vitamin K-dependent Glu γ-carboxylation, did not affect MB-1 cell growth, suggesting that uncarboxylated MGP has a biological effect in niche. These results indicate that MGP may maintain normal and malignant hematopoietic progenitor cells, possibly by modulating BMP signals independently of Glu γ-carboxylation. Aberrant MGP by leukemic cells and selective induction of BMP-4 relative to BMP-2 in stromal cells might specify malignant niche.

Dataset Information

Heat shock protein 70 enhances vascular bone morphogenetic protein-4 signaling by binding matrix Gla protein.

Publications

Heat shock protein 70 enhances vascular bone morphogenetic protein-4 signaling by binding matrix Gla protein.

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