Thomas Nebl, Martha Kotsifas, Pauline Schaap and Paul R. Fisher

J. Muscle Res. & Cell Motility. Dictyostelium Special Issue.

Summary

Dictyostelium mutants expressing aequorin were used to analyze the role of heterotrimeric G-proteins and the second messengers IP₃ and cGMP in regulating chemoreceptor-activated [Ca²⁺]_i increases. It was found that folic acid receptors mediate changes in [Ca²⁺]_i via a Ga4bg-dependent pathway. This was shown by the inhibition of [Ca²⁺]_i responses in Gb (LW6/AEQ) and Ga4 null (ga4-/AEQ) mutants and their restoration with altered kinetics and temperature-sensitivity in Gb null mutants overexpressing wild-type (LW6/Gb+AEQ) and temperature-sensitive Gb isoforms (LW6/Gbts+AEQ). Neither folate nor cAMP-induced [Ca²⁺]_i signals were significantly altered in PLC null transformants (plc-/AEQ), but [Ca²⁺]_i changes elicited by both attractants were significantly prolonged in two stmF mutants lacking cGMP-specific phosphodiesterase activity (NP368/AEQ, NP377/AEQ). This confirms an important role of cGMP in regulating the plasma membrane-associated Ca²⁺ uptake and/or extrusion system of Dictyostelium amoebae. In response to cAMP stimuli this cGMP-dependent part of the Ca²⁺ response appears to be developmentally down-regulated. In contrast to Ca²⁺ responses to folic acid, cAMP receptor-activated [Ca²⁺]_i signals therefore appear to be regulated in a complex manner by two distinct pathways - one G-protein and cGMP-dependent and present at the early aggregation stage (~4 hours post starvation) and the other, G-protein-independent, and dominant once tight aggregates have formed (~8 hours post starvation).