Release of Ca 2+ from the Endoplasmic Reticulum Contributes to Ca 2+ Signaling in Dictyostelium discoideum

ABSTRACT Ca 2+ responses to two chemoattractants, folate and cyclic AMP (cAMP), were assayed in Dictyostelium D. discoideum mutants deficient in one or both of two abundant Ca 2+ -binding proteins of the endoplasmic reticulum (ER), calreticulin and calnexin. Mutants deficient in either or both proteins exhibited enhanced cytosolic Ca 2+ responses to both attractants. Not only were the mutant responses greater in amplitude, but they also exhibited earlier onsets, faster rise rates, earlier peaks, and faster fall rates. Correlations among these kinetic parameters and the response amplitudes suggested that key events in the Ca 2+ response are autoregulated by the magnitude of the response itself, i.e., by cytosolic Ca 2+ levels. This autoregulation was sufficient to explain the altered kinetics of the mutant responses: larger responses are faster in both mutant and wild-type cells in response to both folate (vegetative cells) and cAMP (differentiated cells). Searches of the predicted D. discoideum proteome revealed three putative Ca 2+ pumps and four putative Ca 2+ channels. All but one contained sequence motifs for Ca 2+ - or calmodulin-binding sites, consistent with Ca 2+ signals being autoregulatory. Although cytosolic Ca 2+ responses in the calnexin and calreticulin mutants are enhanced, the influx of Ca 2+ from the extracellular medium into the mutant cells was smaller. Compared to wild-type cells, Ca 2+ release from the ER in the mutants thus contributes more to the total cytosolic Ca 2+ response while influx from the extracellular medium contributes less. These results provide the first molecular genetic evidence that release of Ca 2+ from the ER contributes to cytosolic Ca 2+ responses in D. discoideum .