Function of RasGRP3 in the formation and progression of human breast cancer

Nagy, Z. and Kovács, I. and Török, M. and Tóth, D. and Vereb, György and Buzás, Krisztina and Bíró, Tamás and Czifra, Gabriella (2014) Function of RasGRP3 in the formation and progression of human breast cancer. MOLECULAR CANCER, 13 (1). pp. 1-17. ISSN 1476-4598


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Introduction: Ras guanine nucleotide exchange factors (RasGEFs) mediate the activation of the Ras signaling pathway that is over activated in many human cancers. The RasGRP3, an activator of H-Ras and R-Ras protein exerts oncogenic effects and the overexpression of the protein is observed in numerous malignant cancer types. Here, we investigated the putative alteration of expression and potential function of RasGRP3 in the formation and progression of human breast cancer.Methods: The RasGRP3 and phosphoRasGRP3 expressions were examined in human invasive ductal adenocarcinoma derived samples and cell lines (BT-474, JIMT-1, MCF7, SK-BR-3, MDA-MB-453, T-47D) both in mRNA (Q-PCR) and protein (Western blot; immunohistochemistry) levels. To explore the biological function of the protein, RasGRP3 knockdown cultures were established. To assess the role of RasGRP3 in the viability of cells, annexin-V/PI staining and MitoProbe™ DilC1 (5) assay were performed. To clarify the function of the protein in cell proliferation and in the development of chemotherapeutic resistance, CyQuant assay was performed. To observe the RasGRP3 function in tumor formation, the Severe combined immunodeficiency (SCID) mouse model was used. To investigate the role of the protein in Ras-related signaling Q-PCR and Western blot experiments were performed.Results: RasGRP3 expression was elevated in human breast tumor tissue samples as well as in multiple human breast cancer cell lines. Down-regulation of RasGRP3 expression in breast cancer cells decreased cell proliferation, induced apoptosis in MCF7 cells, and sensitized T-47D cells to the action of drugs Tamoxifen and trastuzumab (Herceptin). Gene silencing of RasGRP3 reduced tumor formation in mouse xenografts as well. Inhibition of RasGRP3 expression also reduced Akt, ERK1/2 and estrogen receptor alpha phosphorylation downstream from IGF-I insulin like growth factor-I (IGF-I) or epidermal growth factor (EGF) stimulation confirming the functional role of RasGRP3 in the altered behavior of these cells.Conclusions: Taken together, our results suggest that the Ras activator RasGRP3 may have a role in the pathological behavior of breast cancer cells and may constitute a therapeutic target for human breast cancer. © 2014 Nagy et al.; licensee BioMed Central Ltd.

Item Type: Article
Uncontrolled Keywords: Western blotting; tumor xenograft; tumor growth; SIGNAL TRANSDUCTION; real time polymerase chain reaction; protein phosphorylation; PROTEIN FUNCTION; protein expression; nonhuman; MOUSE; mitochondrial membrane potential; MCF 7 cell line; in vivo study; IMMUNOHISTOCHEMISTRY; human tissue; human; gene silencing; down regulation; controlled study; cellular distribution; Cell viability; Cell Survival; cell proliferation; Cell growth; cell function; breast carcinoma; Breast cancer cell line; breast adenocarcinoma; ARTICLE; APOPTOSIS; animal model; animal experiment; unclassified drug; somatomedin C; protein kinase B; progesterone receptor; mitogen activated protein kinase 3; mitogen activated protein kinase 1; messenger rna; loricrin; guanine nucleotide exchange factor 3; guanine nucleotide exchange factor; Estrogen Receptor alpha; epidermal growth factor receptor 2; epidermal growth factor; Cytochrome c; Cathepsin D; Tumorigenesis; trastuzumab; tamoxifen; SIGNALING PATHWAYS; RasGRP3; Ras activator; IGF-I; Human breast cancer; EGF; Chemotherapeutic resistance
Subjects: Q Science / természettudomány > QP Physiology / élettan
Depositing User: MTMT SWORD
Date Deposited: 16 Feb 2015 12:47
Last Modified: 16 Feb 2015 13:06

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