Sustained activation of Erk1/2 MAPK and cell growth suppression by the insert-negative, but not the insert-positive isoform of the human calcitonin receptor.
Recently we reported that calcitonin (CT) induces growth arrest at the G2 stage of the cell cycle in HEK-293 cell lines expressing the most abundant, insert-negative, isoform of the human CT receptor (insert -ve hCTR). The present study investigates the involvement of the MAPK signalling pathway in the anti-proliferative actions of CT and compares the activity of an isoform of the hCTR that contains a 16 amino acid insert in the first putative intracellular loop (insert +ve hCTR). Comparison of HEK-293 cells stably transfected with the insert -ve or the insert +ve hCTR, showed that accumulation of cAMP and intracellular free calcium in response to CT were specific for the insert -ve receptor isoform. However, a novel acidification of the extracellular medium was mediated by both isoforms. Treatment with CT of cells expressing the insert -ve hCTR, caused a decrease in cell growth associated with an induction of p21(WAF1/CIP1). Analysis by fluorescence-activated cell scanning showed that growth inhibition was associated with an accumulation of cells in G2. CT treatment of cells expressing the insert -ve, but not insert +ve hCTR, induced the phosphorylation of Erk1/2 MAPK, which persisted for at least 72 h. Treatment of cells expressing the insert -ve hCTR with the MAPK kinase (MEK) inhibitor, PD-98059, inhibited the phosphorylation of Erk1/2 and abrogated the growth inhibitory effects of salmon CT, the accumulation of cells in G2, and the associated induction of p21(WAF1/CIP1). These data suggest that activation of Erk1/2 are downstream effectors of the insert -ve hCTR in modulating cell cycle progression.
Raggatt
L J
LJ
Department of Orthopaedics and Trauma, University of Adelaide, The Royal Adelaide Hospital, Adelaide 5000, South Australia, Australia.
Evdokiou
A
A
Findlay
D M
DM
D001667:Binding, Competitive; D002116:Calcitonin / Q000494:pharmacology*; D002118:Calcium / Q000378:metabolism; D018929:Cell Culture Techniques; D002453:Cell Cycle / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects; D000242:Cyclic AMP / Q000378:metabolism; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D006801:Humans; D006863:Hydrogen-Ion Concentration; D019950:Mitogen-Activated Protein Kinase 1 / Q000187:drug effects / Q000378:metabolism*; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases / Q000187:drug effects / Q000378:metabolism*; D020033:Protein Isoforms / Q000502:physiology; D018003:Receptors, Calcitonin / Q000502:physiology*; D015398:Signal Transduction / Q000502:physiology
C493956:CDKN1A protein, human; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins; D020033:Protein Isoforms; D018003:Receptors, Calcitonin; D002116:Calcitonin; D000242:Cyclic AMP; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; D002118:Calcium