mpermeable estrogen. Phosphorylation of extracellular regulated kinases and the cAMP response element-binding protein SB-1317 web occurred as soon as 5 to 15 min and was reversed by pertussis toxin, a G protein inhibitor. In 2011, both Franco et al. and Rago et al. showed by immunohistochemistry that GPER was present in testicular germ cell tumours, including seminoma. Our study confirmed this expression and demonstrated that GPER was overexpressed in seminomas but not in non seminoma tumours by using a robust quantitative approach of mRNAs and proteins levels by Q-PCR and Western blotting related to b-actin, considered as a housekeeping gene, and compared with the peri-tumoural normal testicular tissue for each patient. Eventhough some abnormal expression could be already observed in peritumoural tissues, this method excludes inter-individual variations of GPER expression as each patient represents its own control. This overexpression was also confirmed by comparison between JKT-1 seminoma cells and NCCIT choriocarcinoma cells. GPER overexpression has been linked to the development of advanced breast cancer, high-grade endometrial tumours and poor prognosis for ovarian cancer. GPER overexpression in seminomas may also represent a prognosis factor for testicular germ cell tumours and perhaps a potential therapeutic target as suggested by the result obtained with a GPER-specific antagonist. These “7851485 different points are now under investigation in our laboratory. The precise mechanisms leading to such a GPER overexpression in seminomas need investigation. In a recent study, three GPER SNPs were correlated with aggressive breast cancers; involvement of GPER SNPs in human testicular germ cell cancers is now under verification in our laboratory. Another explanation Overexpression of GPR30 in Human Seminoma for GPER overexpression could be hypomethylation of the GPER gene promoter region considering that such epigenetic modifications are now largely described in cancer. We have already reported that E2 is able to induce a suppressive effect on JKT-1 cell proliferation. This effect is completely suppressed by a pure ER antagonist, which supports the role of ERb, the only classical ER expressed in JKT-1 cells. In JKT-1 cells, ERb and GPER did not co-localize but induced two opposite pathways. E2, with a well-known high affinity for ERb but a low reported affinity for GPER, inhibited cell proliferation likely through ERb, as already described for other oestrogen-dependent cancers. In contrast, G1, a selective GPER agonist, which had a low affinity for ERb but a high affinity for GPER, and E2-BSA, induced JKT-1 cell proliferation. It has been suggested in some models that GPER and ER or a truncated splice variant of ERs could cooperate or cross-talk. ERa is not express in JKT-1 and ERb does 
not localize in the membrane as shown by western blot analysis after subcellular fractioning. Moreover ERb did not immunoprecipitated with caveolin, a protein of the raft region in the cell membrane. Kang et al. have reported in a human breast cancer model that a truncated variant form of ERa, ER-a36, expressed in the membrane, acted upstream of GPER and was even able to trigger by itself a rapid non genomic estrogenic activation. While we 6 Overexpression of GPR30 in Human Seminoma cannot completely eliminate a truncated splice variant form of ERa or ERb in JKT-1 not recognized by our primers or by our antibodies such as ER-a36, our data support the direct implication of GP
