Regulation of Starch Stores by a Ca²⁺-Dependent Protein Kinase Is Essential for Viable Cyst Development in Toxoplasma gondii

Summary Transmissible stages of Toxoplasma gondii store energy in the form of the carbohydrate amylopectin. Here, we show that the Ca²⁺-dependent protein kinase CDPK2 is a critical regulator of amylopectin metabolism. Increased synthesis and loss of degradation of amylopectin in CDPK2 deficient parasites results in the hyperaccumulation of this sugar polymer. A carbohydrate-binding module 20 (CBM20) targets CDPK2 to amylopectin stores, while the EF-hands regulate CDPK2 kinase activity in response to Ca²⁺ to modulate amylopectin levels. We identify enzymes involved in amylopectin turnover whose phosphorylation is dependent on CDPK2 activity. Strikingly, accumulation of massive amylopectin granules in CDPK2-deficient bradyzoite stages leads to gross morphological defects and complete ablation of cyst formation in a mouse model. Together these data show that Ca²⁺ signaling regulates carbohydrate metabolism in Toxoplasma and that the post-translational control of this pathway is required for normal cyst development.