Extracellular ATP, ecto-ATPase and calcium influx in Dictyostelium discoideum cells

Abstract

There is evidence that a protein kinase is present on the surface of Dictyostelium discoideum cells [l-3] and is capable of phosphorylating certain plasma membrane proteins [3]. Such phosphorylation may be involved in membrane permeability and transport, cell movement, and the activation of cyclic AMP receptors which occurs during early differentiation [4]. External ATP induces cell aggregation, decreases the chemotactic sensitivity of cells to a cyclic AMP gradient and inhibits intercellular contact sites outside the aggregation center [ 1,5]. We found that ATP (0.1-0.8 PM produced at 1.5 X lo7 cells/ml) is present extracellularly in cell suspensions. This reflected a steady state concentration since a Mg”-dependent ecto-ATPase was continually hydrolyzing extracellular ATP. The ectoATPase was inhibited by suramin and I-ethyl-3(3-dimethyl-aminopropyl)carbodiimide (EDAC), but not by oligomycin or ouabain. Ecto-ADPase, -AMPase and nucleotide diphosphokinase activities were also detected. Calcium influx was inhibited by suramin, l-ethyl-3-(3-dimethyl-aminopropyl)carbodiimide (EDAC), KCN, dinitrophenol, carbonyl-mchlorophenylhydrazone (CCCP), adenyl imidodiphosphate, adenyl methylene diphosphate, and by adding hexokinase or apyrase to split extracellular ATP. Addition of ATP (optimum level 10e6 M) to cell suspensions stimulated calcium influx and partially overcame the KCN-induced inhibition of this influx. Glucose uptake was not affected by suramin or apyrase.