Angiogenesis plays a key role in various physiological and pathological conditions, including inflammation and tumor growth. The bone morphogenetic protein (BMP) antagonist gremlin has been identified as a novel pro-angiogenic factor. Gremlin promotes neovascular responses via a BMP-independent activation of the vascular endothelial growth factor (VEGF) receptor-2 (VEGFR2). BMP antagonists may act as covalent or non-covalent homodimers or in a monomeric form, while VEGFRs ligands are usually dimeric. However, the oligomeric state of gremlin and its role in modulating the biological activity of the protein remain to be elucidated.Here we show that gremlin is expressed in vitro and in vivo both as a monomer and as a covalently linked homodimer. Mutagenesis of amino acid residue Cys141 prevents gremlin dimerization leading to the formation of gremlinC141A monomers. GremlinC141A monomer retains a BMP antagonist activity similar to the wild-type dimer, but is devoid of a significant angiogenic capacity. Notably, we found that gremlinC141A mutant engages VEGFR2 in a non-productive manner, thus acting as receptor antagonist. Accordingly, both gremlinC141A and wild-type monomers inhibit angiogenesis driven by dimeric gremlin or VEGF-A165. Moreover, by acting as a VEGFR2 antagonist, gremlinC141A inhibits the angiogenic and tumorigenic potential of murine breast and prostate cancer cells in vivo.In conclusion, our data show that gremlin exists in multiple forms endowed with specific bioactivities and provide new insights into the molecular bases of gremlin dimerization. Furthermore, we propose gremlin monomer as a new inhibitor of VEGFR2 signalling during tumor growth.
Oncotarget. 2016 May 11 [Epub ahead of print]
Elisabetta Grillo, Cosetta Ravelli, Michela Corsini, Kurt Ballmer-Hofer, Luca Zammataro, Pasqua Oreste, Giorgio Zoppetti, Chiara Tobia, Roberto Ronca, Marco Presta, Stefania Mitola
Experimental Oncology and Immunology, Department of Molecular and Translational Medicine, University of Brescia, Brescia, 25123, Italy., Experimental Oncology and Immunology, Department of Molecular and Translational Medicine, University of Brescia, Brescia, 25123, Italy., Experimental Oncology and Immunology, Department of Molecular and Translational Medicine, National Institute of Neurosciences, University of Brescia, Brescia, 25123, Italy., Biomolecular Research, Molecular Cell Biology, Paul Scherrer Institut, Villigen, 5232, Switzerland., Center of Genomics Science of IIT@SEMM, Milan, 20139, Italy., Glycores2000, Milan, 20155 Italy., Glycores2000, Milan, 20155 Italy., Experimental Oncology and Immunology, Department of Molecular and Translational Medicine, University of Brescia, Brescia, 25123, Italy., Experimental Oncology and Immunology, Department of Molecular and Translational Medicine, University of Brescia, Brescia, 25123, Italy., Experimental Oncology and Immunology, Department of Molecular and Translational Medicine, University of Brescia, Brescia, 25123, Italy., Experimental Oncology and Immunology, Department of Molecular and Translational Medicine, University of Brescia, Brescia, 25123, Italy.