Gβγ subunits inhibit Epac-induced melanoma cell migration

BMC Cancer

Research Article

Mariana S De Lorenzo¹  Masanari Umemura¹  Erdene Baljinnyam¹  Mizuka Iwatsubo¹  Lai-Hua Xie¹  Kousaku Iwatsubo²  Suzie Chen³  Martha Nowycky⁴  James S Goydos⁵ 

[1] Department of Cell Biology and Molecular Medicine, New Jersey Medical School-University of Medicine and Dentistry of New Jersey, 185 South Orange Avenue, 07103, Newark, USA;Department of Cell Biology and Molecular Medicine, New Jersey Medical School-University of Medicine and Dentistry of New Jersey, 185 South Orange Avenue, 07103, Newark, USA;Cardiovascular Research Institute, Yokohama City University Graduate School of Medicine, 236-0004, Yokohama, 3-9 Fukuura, Japan;Department of Chemical Biology, Susan Lehman Cullen Laboratory of Cancer Research in the Ernest Mario School of Pharmacy, Rutgers University, 60 Frelinghuysen Road, 08854, Piscataway, New Jersey, USA;Department of Pharmacology and Physiology, New Jersey Medical School-University of Medicine and Dentistry of New Jersey, 185 South Orange Avenue, 07103, Newark, USA;Division of Surgical Oncology, Department of Surgery, UMDNJ-Robert Wood Johnson Medical School, 195 Little Albany Street, 08854, New Brunswick, New Jersey, USA;

关键词: Melanoma;    Cell Migration;    Melanoma Cell;    Transient Receptor Potential Channel;    Canonical Transient Receptor Potential;   

DOI  :  10.1186/1471-2407-11-256

 received in 2010-12-24, accepted in 2011-06-17,  发布年份 2011

来源: Springer

【 摘 要 】

BackgroundRecently we reported that activation of Epac1, an exchange protein activated by cAMP, increases melanoma cell migration via Ca 2+ release from the endoplasmic reticulum (ER). G-protein βγ subunits (Gβγ) are known to act as an independent signaling molecule upon activation of G-protein coupled receptor. However, the role of Gβγ in cell migration and Ca 2+ signaling in melanoma has not been well studied. Here we report that there is crosstalk of Ca 2+ signaling between Gβγ and Epac in melanoma, which plays a role in regulation of cell migration.MethodsSK-Mel-2 cells, a human metastatic melanoma cell line, were mainly used in this study. Intracellular Ca 2+ was measured with Fluo-4AM fluorescent dyes. Cell migration was examined using the Boyden chambers.ResultsThe effect of Gβγ on Epac-induced cell migration was first examined. Epac-induced cell migration was inhibited by mSIRK, a Gβγ -activating peptide, but not its inactive analog, L9A, in SK-Mel-2 cells. Guanosine 5', α-β-methylene triphosphate (Gp(CH2)pp), a constitutively active GTP analogue that activates Gβγ, also inhibited Epac-induced cell migration. In addition, co-overexpression of β1 and γ2, which is the major combination of Gβγ, inhibited Epac1-induced cell migration. By contrast, when the C-terminus of β adrenergic receptor kinase (βARK-CT), an endogenous inhibitor for Gβγ, was overexpressed, mSIRK's inhibitory effect on Epac-induced cell migration was negated, suggesting the specificity of mSIRK for Gβγ. We next examined the effect of mSIRK on Epac-induced Ca 2+ response. When cells were pretreated with mSIRK, but not with L9A, 8-(4-Methoxyphenylthio)-2'-O-methyladenosine-3',5'-cyclic monophosphate (8-pMeOPT), an Epac-specific agonist, failed to increase Ca 2+ signal. Co-overexpression of β1 and γ2 subunits inhibited 8-pMeOPT-induced Ca 2+ elevation. Inhibition of Gβγ with βARK-CT or guanosine 5'-O-(2-thiodiphosphate) (GDPβS), a GDP analogue that inactivates Gβγ, restored 8-pMeOPT-induced Ca 2+ elevation even in the presence of mSIRK. These data suggested that Gβγ inhibits Epac-induced Ca 2+ elevation. Subsequently, the mechanism by which Gβγ inhibits Epac-induced Ca 2+ elevation was explored. mSIRK activates Ca 2+ influx from the extracellular space. In addition, W-5, an inhibitor of calmodulin, abolished mSIRK's inhibitory effects on Epac-induced Ca 2+ elevation, and cell migration. These data suggest that, the mSIRK-induced Ca 2+ from the extracellular space inhibits the Epac-induced Ca 2+ release from the ER, resulting suppression of cell migration.ConclusionWe found the cross talk of Ca 2+ signaling between Gβγ and Epac, which plays a major role in melanoma cell migration.

【 授权许可】

Unknown   
© Baljinnyam et al; licensee BioMed Central Ltd. 2011. This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.