Extracellular Ca2+ stimulates the activation of mitogen-activated protein kinase and cell growth in human fibroblasts.
In serum-free medium containing serum replacements but totally lacking in protein growth factors, diploid human fibroblasts remained quiescent if the extracellular Ca2+ concentration was only 0.1 mM. However, when the Ca2+ concentration in this medium was increased to 1 mM, the cells replicated as rapidly as they do in medium supplemented with protein growth factors. When quiescent cells in medium with only 0.1 mM Ca2+ were exposed to 1 or 10 mM Ca2+ or 100 ng/ml epidermal growth factor (EGF), the 42 kDa and 44 kDa forms of mitogen-activated protein kinase (MAPK) were rapidly activated, as demonstrated by a characteristic electrophoretic mobility shift of these proteins and by their enhanced ability to phosphorylate myelin basic protein (MBP). Analysis of fractions from Mono Q anion-exchange chromatography of lysates of cells exposed to 10 mM Ca2+ or 100 ng/ml EGF revealed a peak of MBP phosphorylation activity that was coeluted with p42 and p44 MAPK as shown by immunoblot analysis. Activation of MAPK by extracellular Ca2+ was dose-dependent and biphasic, with a peak of activation at 5-10 min after exposure, followed by a period of sustained activation of MAPK at a lower level. This pattern has been shown [Vouret-Craviari, Van Obberghen-Schilling, Scimeca, Van Obberghen and Pouysségur (1993) Biochem J. 289, 209-214] to correlate with the re-entry of mammalian cells into the cell cycle.
Huang
S
S
Department of Microbiology, Michigan State University, East Lansing 48824, USA.
Maher
V M
VM
McCormick
J J
JJ
D002118:Calcium / Q000494:pharmacology*; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism*; D002455:Cell Division / Q000187:drug effects*; D002460:Cell Line; D002852:Chromatography, Ion Exchange; D004789:Enzyme Activation; D004815:Epidermal Growth Factor / Q000494:pharmacology*; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects / Q000201:enzymology; D006801:Humans; D015151:Immunoblotting; D007231:Infant, Newborn; D007700:Kinetics; D008297:Male; D004676:Myelin Basic Protein / Q000302:isolation & purification / Q000378:metabolism; D010750:Phosphoproteins / Q000302:isolation & purification / Q000378:metabolism; D010766:Phosphorylation; D012867:Skin / Q000166:cytology / Q000201:enzymology
D004676:Myelin Basic Protein; D010750:Phosphoproteins; D004815:Epidermal Growth Factor; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D002118:Calcium