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Oncogenic activities of metabotropic glutamate receptor 1 (Grm1) in melanocyte transformation.

ABSTRACT: Previously, we reported a transgenic mouse line, TG-3, that develops spontaneous melanoma with 100% penetrance. We demonstrated that ectopic expression of Grm1 in melanocytes was sufficient to induce melanoma in vivo. In this present study, the transforming properties of Grm1 in two cultured immortalized melanocytes were investigated. We showed that, in contrast to parental melanocytes, these Grm1-clones have lost their requirement of TPA supplement for proliferation and have acquired the ability to form colonies in semi-solid medium. Xenografts of these cells formed robust tumors in both immunodeficient nude and syngeneic mice with a short latency (3-5 days). The malignancy of these cells was demonstrated by angiogenesis and invasion to the muscle and the intestine. The requirement of Grm1 expression for the maintenance of transformation was demonstrated by an inducible siRNA system. Induction of expression of siRNA for Grm1 reduced the number of proliferating/viable cells in vitro and suppressed in vivo xenografted tumor growth in comparison with control. Taken together, these results showed that expression of exogeneously introduced Grm1 is sufficient to induce full transformation of immortalized melanocytes.

SUBMITTER: Shin SS

PROVIDER: S-EPMC2854004 | biostudies-literature | 2008 Jun

REPOSITORIES: biostudies-literature

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Oncogenic activities of metabotropic glutamate receptor 1 (Grm1) in melanocyte transformation.

Shin Seung-Shick SS Namkoong Jin J Wall Brian A BA Gleason Ryan R Lee Hwa Jin HJ Chen Suzie S

Pigment cell & melanoma research 20080423 3

Previously, we reported a transgenic mouse line, TG-3, that develops spontaneous melanoma with 100% penetrance. We demonstrated that ectopic expression of Grm1 in melanocytes was sufficient to induce melanoma in vivo. In this present study, the transforming properties of Grm1 in two cultured immortalized melanocytes were investigated. We showed that, in contrast to parental melanocytes, these Grm1-clones have lost their requirement of TPA supplement for proliferation and have acquired the abilit ...[more]

PMID: 18435704

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Project description:Non-neuronal expression of components of the glutamatergic system has been increasingly observed, and our laboratory previously had demonstrated the etiological role of ectopically expressed metabotropic glutamate receptor 1 (Grm1/mGluR1) in mouse models of melanoma. We hypothesize that inappropriate glutamatergic signaling in other cell types can dysregulate growth leading to transformation and tumorigenesis. As most cancers are carcinomas, we selected an immortalized primary baby mouse kidney (iBMK) cell model to assess whether Grm1 can transform epithelial cells. These iBMK cells, engineered to be immortal yet nontumorigenic and retaining normal epithelial characteristics, were used as recipients for exogenous Grm1 cDNA. Several stable Grm1-expressing clones were isolated and the Grm1-receptors were shown to be functional, as evidenced by the accumulation of second messengers in response to Grm1 agonist. Additionally activated by agonist were mitogen-activated protein kinase (MAPK) and AKT/protein kinase B signaling cascades, the major intracellular pathways shown by many investigators to be critical in melanomagenesis and other neoplasms. These Grm1-iBMK cells exhibited enhanced cell proliferation in in vitro methylthiazolyldiphenyl-tetrazolium bromide (MTT) assays and significant tumorigenicity in in vivo allografts. Persistent Grm1 expression was required for the maintenance of the in vivo tumorigenic phenotype as demonstrated by an inducible Grm1-silencing RNA. These are the first results that indicate that Grm1 can be an oncogene in epithelial cells. In addition, relevance to human disease in the corresponding tumor type of renal cell carcinoma (RCC) may be suggested by observed expression of GRM1/mGluR1 in a number of RCC tumor biopsy samples and cell lines, and the effects of GRM1 modulation on tumorigenicity therein. Moreover, RCC cell lines exhibited elevated levels of extracellular glutamate, and some lines responded to drugs, which modulate the glutamatergic system. These findings imply a possible role for glutamate signaling apparatus in RCC cell growth, and that the glutamatergic system may be a therapeutic target in RCC.

Project description:Metabotropic glutamate receptor 1 (mGluR1/Grm1) is a member of the G-protein-coupled receptor superfamily, which was once thought to only participate in synaptic transmission and neuronal excitability, but has more recently been implicated in non-neuronal tissue functions. We previously described the oncogenic properties of Grm1 in cultured melanocytes in vitro and in spontaneous melanoma development with 100 % penetrance in vivo. Aberrant mGluR1 expression was detected in 60-80 % of human melanoma cell lines and biopsy samples. As most human cancers are of epithelial origin, we utilized immortalized mouse mammary epithelial cells (iMMECs) as a model system to study the transformative properties of Grm1. We introduced Grm1 into iMMECs and isolated several stable mGluR1-expressing clones. Phenotypic alterations in mammary acinar architecture were assessed using three-dimensional morphogenesis assays. We found that mGluR1-expressing iMMECs exhibited delayed lumen formation in association with decreased central acinar cell death, disrupted cell polarity, and a dramatic increase in the activation of the mitogen-activated protein kinase pathway. Orthotopic implantation of mGluR1-expressing iMMEC clones into mammary fat pads of immunodeficient nude mice resulted in mammary tumor formation in vivo. Persistent mGluR1 expression was required for the maintenance of the tumorigenic phenotypes in vitro and in vivo, as demonstrated by an inducible Grm1-silencing RNA system. Furthermore, mGluR1 was found be expressed in human breast cancer cell lines and breast tumor biopsies. Elevated levels of extracellular glutamate were observed in mGluR1-expressing breast cancer cell lines and concurrent treatment of MCF7 xenografts with glutamate release inhibitor, riluzole, and an AKT inhibitor led to suppression of tumor progression. Our results are likely relevant to human breast cancer, highlighting a putative role of mGluR1 in the pathophysiology of breast cancer and the potential of mGluR1 as a novel therapeutic target.

Dataset Information

Oncogenic activities of metabotropic glutamate receptor 1 (Grm1) in melanocyte transformation.

Publications

Oncogenic activities of metabotropic glutamate receptor 1 (Grm1) in melanocyte transformation.

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