title: Estrogen and non-genomic upregulation of voltage-gated Na(+) channel activity in MDA-MB-231 human breast cancer cells: role in adhesion.
abstract: External (but not internal) application of beta-estradiol (E2) increased the current amplitude of voltage-gated Na(+) channels (VGSCs) in MDA-MB-231 human breast cancer (BCa) cells. The G-protein activator GTP-gamma-S, by itself, also increased the VGSC current whilst the G-protein inhibitor GDP-beta-S decreased the effect of E2. Expression of GPR30 (a G-protein-coupled estrogen receptor) in MDA-MB-231 cells was confirmed by PCR, Western blot and immunocytochemistry. Importantly, G-1, a specific agonist for GPR30, also increased the VGSC current amplitude in a dose-dependent manner. Transfection and siRNA-silencing of GPR30 expression resulted in corresponding changes in GPR30 protein expression but only internally, and the response to E2 was not affected. The protein kinase A inhibitor, PKI, abolished the effect of E2, whilst forskolin, an adenylate cyclase activator, by itself, increased VGSC activity. On the other hand, pre-incubation of the MDA-MB-231 cells with brefeldin A (a trans-Golgi protein trafficking inhibitor) had no effect on the E2-induced increase in VGSC amplitude, indicating that such trafficking ('externalisation') of VGSC was not involved. Finally, acute application of E2 decreased cell adhesion whilst the specific VGSC blocker tetrodotoxin increased it. Co-application of E2 and tetrodotoxin inhibited the effect of E2 on cell adhesion, suggesting that the effect of E2 was mainly through VGSC activity. Pre-treatment of the cells with PKI abolished the effect of E2 on adhesion, consistent with the proposed role of PKA. Potential implications of the E2-induced non-genomic upregulation of VGSC activity for BCa progression are discussed.
authors.affiliation: Division of Cell and Molecular Biology, Neuroscience Solutions to Cancer Research Group, Imperial College London, South Kensington Campus, London, UK. s.p.fraser@imperial.ac.uk
authors.forename: Scott P
authors.initials: SP
authors.lastname: Fraser
authors.forename: Iley
authors.initials: I
authors.lastname: Ozerlat-Gunduz
authors.forename: Rustem
authors.initials: R
authors.lastname: Onkal
authors.forename: James K J
authors.initials: JK
authors.lastname: Diss
authors.forename: David S
authors.initials: DS
authors.lastname: Latchman
authors.forename: Mustafa B A
authors.initials: MB
authors.lastname: Djamgoz
mesh_terms: D000262:Adenylyl Cyclases / Q000378:metabolism; D001943:Breast Neoplasms / Q000201:enzymology / Q000378:metabolism* / Q000473:pathology*; D020126:Brefeldin A / Q000494:pharmacology; D002448:Cell Adhesion / Q000187:drug effects; D045744:Cell Line, Tumor; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D004958:Estradiol / Q000494:pharmacology*; D005260:Female; D019204:GTP-Binding Proteins / Q000378:metabolism; D020868:Gene Silencing / Q000187:drug effects; D016678:Genome / Q000235:genetics; D006056:Golgi Apparatus / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D015640:Ion Channel Gating / Q000187:drug effects*; D007700:Kinetics; D008954:Models, Biological; D021381:Protein Transport / Q000187:drug effects; D011960:Receptors, Estrogen; D043562:Receptors, G-Protein-Coupled / Q000235:genetics / Q000378:metabolism; D015222:Sodium Channels / Q000378:metabolism*; D015854:Up-Regulation / Q000187:drug effects*
chemical_list: C109584:GPER1 protein, human; D011960:Receptors, Estrogen; D043562:Receptors, G-Protein-Coupled; D015222:Sodium Channels; D020126:Brefeldin A; D004958:Estradiol; D017868:Cyclic AMP-Dependent Protein Kinases; D019204:GTP-Binding Proteins; D000262:Adenylyl Cyclases