C3G promotes a selective release of angiogenic factors from activated mouse platelets to regulate angiogenesis and tumor metastasis

// Victor Martin-Granado 1, 2 , Sara Ortiz-Rivero 1, 2 , Rita Carmona 3 , Sara Gutierrez-Herrero 1, 2 , Mario Barrera 1 , Laura San-Segundo 1, 2 , Celia Sequera 4 , Pedro Perdiguero 2, 5 , Francisco Lozano 2, 6 , Francisco Martin-Herrero 2, 5 , Jose Ramon Gonzalez-Porras 2, 7 , Ramon Munoz-Chapuli 3 , Almudena Porras 4 and Carmen Guerrero 1, 2, 8 1 Instituto de Biologia Molecular y Celular del Cancer, USAL-CSIC, Salamanca, Spain 2 Instituto de Investigacion Biomedica de Salamanca (IBSAL), Salamanca, Spain 3 Departamento de Biologia Animal, Universidad de Malaga, Malaga, Spain 4 Departamento de Bioquimica y Biologia Molecular II, Facultad de Farmacia, Universidad Complutense de Madrid, Instituto de Investigacion Sanitaria del Hospital Clinico San Carlos (IdISSC), Madrid, Spain 5 Departamento de Cardiologia, Hospital Universitario de Salamanca, Universidad de Salamanca, Salamanca, Spain 6 Departamento de Angiologia y Cirugia Vascular, Hospital Universitario de Salamanca, Universidad de Salamanca, Salamanca, Spain 7 Departamento de Hematologia, Hospital Universitario de Salamanca, Universidad de Salamanca, Salamanca, Spain 8 Departamento de Medicina, Universidad de Salamanca, Salamanca, Spain Correspondence to: Carmen Guerrero, email: cguerrero@usal.es Almudena Porras, email: maporras@ucm.es Keywords: C3G; platelet secretome; angiogenesis; Vamp-7; metastasis Received: July 05, 2017      Accepted: October 25, 2017      Published: November 06, 2017 ABSTRACT Previous observations indicated that C3G (RAPGEF1) promotes α-granule release, evidenced by the increase in P-selectin exposure on the platelet surface following its activation. The goal of the present study is to further characterize the potential function of C3G as a modulator of the platelet releasate and its implication in the regulation of angiogenesis. Proteomic analysis revealed a decreased secretion of anti-angiogenic factors from activated transgenic C3G and C3GΔCat platelets. Accordingly, the secretome from both transgenic platelets had an overall pro-angiogenic effect as evidenced by an in vitro capillary-tube formation assay with HUVECs (human umbilical vein endothelial cells) and by two in vivo models of heterotopic tumor growth. In addition, transgenic C3G expression in platelets greatly increased mouse melanoma cells metastasis. Moreover, immunofluorescence microscopy showed that the pro-angiogenic factors VEGF and bFGF were partially retained into α-granules in thrombin- and ADP-activated mouse platelets from both, C3G and C3GΔCat transgenic mice. The observed interaction between C3G and Vesicle-associated membrane protein (Vamp)-7 could explain these results. Concomitantly, increased platelet spreading in both transgenic platelets upon thrombin activation supports this novel function of C3G in α-granule exocytosis. Collectively, our data point out to the co-existence of Rap1GEF-dependent and independent mechanisms mediating C3G effects on platelet secretion, which regulates pathological angiogenesis in tumors and other contexts. The results herein support an important role for platelet C3G in angiogenesis and metastasis.