ERK activation and cell growth require CaM kinases in MCF-7 breast cancer cells.
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 M(3)-subtype GPCR inhibitor, 4-DAMP, and siRNA against the M(3)-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 KKalpha and CaM KIgamma 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.
Schmitt
John M
JM
The Biology Department, George Fox University, 414 N Meridian St, Newberg, OR 97132, USA. jschmitt@georgefox.edu
Abell
Ellen
E
Wagner
Andrea
A
Davare
Monika A
MA
D001943:Breast Neoplasms / Q000201:enzymology*; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism*; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism*; D005260:Female; D006801:Humans; D050807:ets-Domain Protein Elk-1 / Q000378:metabolism
D050807:ets-Domain Protein Elk-1; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D048049:Extracellular Signal-Regulated MAP Kinases