Calf signalling governs stimulated exocytosis and exocytosis-coupled endocytosis also in Paramecium cells. Upon stimulation, the ::s10 3 dense-core exocytotic organelles (trichocysts) can be synchronously (80 ms) released, followed by endocytotic membrane resealing (350 ms) and retrieval. Paramecium is the most synchronous dense-core exocytotic system known, allowing to dissect rapidly reversible Calf -dependent phenomena. This holds for the reversible de-/re-phosphorylation cycle of a 63 kD phosphoprotein, pp63/parafusin (pf), which we have cloned, immuno-Iocalised, and characterised as phosphoglucomutase. the enzyme funneling glucose into the glycolytic pathway. It was isolated ex vivo. followed by MALDl analysis, while X-ray structure analysis was performed after heterologous expression. We found multiple phosphorylation of superficial SerlThr residues. Although present also in exo-mutants. pp63/pf is selectively de-phosphorylated only in exo+ strains during synchronous exocytosis (80 ms) and re-phosphorylated within~20 s, Le., the time required to re-establish [Calf] homeostasis. We have isolated relevant protein phosphatases and kinases and probed their activity on pp63/pf in vitro. We consider CalfIcalmodulin-activated PP2B (calcineurin, whose subunits have been cloned) relevant for de-phosphorylation. Re-phosphorylation can be achieved by two protein kinases that also have been cloned. One is activated by cGMP (PKG) which in turn is formed by Calf-activated guanylate cyclase. Another kinase, casein kinase 2. is inhibited by Calf and. hence, activated with some delay in parallel to decreasing [Calf] after exocytosis. In total, several Calf -sensitive cycles cooperate whose protein components have been localised to the cell cortex. Regulation of the phosphorylation degree of pp63/pf may affect structure binding on a microscale and/or its enzymatic activity. All this may serve fueling substrate into glycolysis with increased ATP re-formation (compromised in exo-mutants) and NADH formation. with effects on Calf signalling including mobilisation from cortical stores (alveolar sacs) and overall effects on ATP and Ca 2 + dynamics during synchronous exo-and endocytosis.