TY - JOUR
T1 - ERK activation and cell growth require CaM kinases in MCF-7 breast cancer cells
AU - Schmitt, John M.
AU - Abell, Ellen
AU - Wagner, Andrea
AU - Davare, Monika A.
N1 - Funding Information:
Acknowledgments The authors would like to thank Tom Soderling for providing reagents and helpful discussions and Luke Fletcher as well as Sean Nygaard for technical assistance. This work was supported in part by the Paul K. and Evalyn E. C. Richter Memorial Fund and the Holman Endowment for the Sciences at George Fox University to Andrea Wagner and Ellen Abell.
PY - 2010/2
Y1 - 2010/2
N2 - Previous studies on MCF-7 breast cancer cells have shown that the G-protein coupled receptor (GPCR) agonist carbachol increases intracellular calcium levels and the activation of extracellular signal-regulated kinase (ERK). Calcium and calmodulin regulate the calcium/calmodulin-dependent kinase (CaM kinase) family of proteins that have been proposed to regulate ERK and gene transcription. Our results suggest that both estrogen (E2) and carbachol treatment of MCF-7 breast cancer cells trigger phosphorylation of ERK1/2 and the transcription factor Elk-1. Carbachol and estrogen triggered nearly a four- to sixfold increase in MCF-7 cell proliferation by 96 h, respectively. Carbachol-stimulated ERK activation and cell growth was completely blocked by the Muscarinic M3-subtype GPCR inhibitor, 4-DAMP, and siRNA against the M3-subtype GPCR. Interestingly, blockade of CaM KK with the selective inhibitor STO-609 prevented carbachol activation CaM KI, ERK, Elk-1, and cell growth. Consistent with these observations, knockdown of CaM KKα and CaM KIγ with shRNA-containing plasmids blocked ERK activation by carbachol. In addition, Elk-1 phosphorylation and luciferase activity in response to carbachol treatment was also dependent upon CaM kinases and was inhibited by U0126, STO-609, and siRNA knockdown of CaM kinases and ERK2. Finally, blockade of either CaM KK (with STO-609) or ERK (with U0126) activities resulted in the inhibition of carbachol- and estrogen-mediated cyclin D1 expression and MCF-7 cell growth. Taken together, our results suggest that carbachol treatment of MCF-7 cells activates CaM KI, ERK, the transcription factor Elk-1, cyclin D1, and cell growth through CaM KK.
AB - Previous studies on MCF-7 breast cancer cells have shown that the G-protein coupled receptor (GPCR) agonist carbachol increases intracellular calcium levels and the activation of extracellular signal-regulated kinase (ERK). Calcium and calmodulin regulate the calcium/calmodulin-dependent kinase (CaM kinase) family of proteins that have been proposed to regulate ERK and gene transcription. Our results suggest that both estrogen (E2) and carbachol treatment of MCF-7 breast cancer cells trigger phosphorylation of ERK1/2 and the transcription factor Elk-1. Carbachol and estrogen triggered nearly a four- to sixfold increase in MCF-7 cell proliferation by 96 h, respectively. Carbachol-stimulated ERK activation and cell growth was completely blocked by the Muscarinic M3-subtype GPCR inhibitor, 4-DAMP, and siRNA against the M3-subtype GPCR. Interestingly, blockade of CaM KK with the selective inhibitor STO-609 prevented carbachol activation CaM KI, ERK, Elk-1, and cell growth. Consistent with these observations, knockdown of CaM KKα and CaM KIγ with shRNA-containing plasmids blocked ERK activation by carbachol. In addition, Elk-1 phosphorylation and luciferase activity in response to carbachol treatment was also dependent upon CaM kinases and was inhibited by U0126, STO-609, and siRNA knockdown of CaM kinases and ERK2. Finally, blockade of either CaM KK (with STO-609) or ERK (with U0126) activities resulted in the inhibition of carbachol- and estrogen-mediated cyclin D1 expression and MCF-7 cell growth. Taken together, our results suggest that carbachol treatment of MCF-7 cells activates CaM KI, ERK, the transcription factor Elk-1, cyclin D1, and cell growth through CaM KK.
KW - CaM kinase
KW - Carbachol
KW - Cell growth
KW - Cyclin D1
KW - ERK
KW - Elk-1
KW - Estrogen
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U2 - 10.1007/s11010-009-0252-9
DO - 10.1007/s11010-009-0252-9
M3 - Article
C2 - 19763792
AN - SCOPUS:75949110752
SN - 0300-8177
VL - 335
SP - 155
EP - 171
JO - Molecular and Cellular Biochemistry
JF - Molecular and Cellular Biochemistry
IS - 1-2
ER -