Toxoplasma gondii Cyclic AMP-Dependent Protein Kinase Subunit 3 Is Involved in the Switch from Tachyzoite to Bradyzoite Development

Abstract

ABSTRACT Toxoplasma gondii is an obligate intracellular apicomplexan parasite that infects warm-blooded vertebrates, including humans. Asexual reproduction in T. gondii allows it to switch between the rapidly replicating tachyzoite and quiescent bradyzoite life cycle stages. A transient cyclic AMP (cAMP) pulse promotes bradyzoite differentiation, whereas a prolonged elevation of cAMP inhibits this process. We investigated the mechanism(s) by which differential modulation of cAMP exerts a bidirectional effect on parasite differentiation. There are three protein kinase A (PKA) catalytic subunits ( Tg PKAc1 to -3) expressed in T. gondii . Unlike Tg PKAc1 and Tg PKAc2, which are conserved in the phylum Apicomplexa, Tg PKAc3 appears evolutionarily divergent and specific to coccidian parasites. Tg PKAc1 and Tg PKAc2 are distributed in the cytomembranes, whereas Tg PKAc3 resides in the cytosol. Tg PKAc3 was genetically ablated in a type II cyst-forming strain of T. gondii (Pru Δku80Δhxgprt ) and in a type I strain (RH Δku80Δhxgprt ), which typically does not form cysts. The Δ pkac3 mutant exhibited slower growth than the parental and complemented strains, which correlated with a higher basal rate of tachyzoite-to-bradyzoite differentiation. 3-Isobutyl-1-methylxanthine (IBMX) treatment, which elevates cAMP levels, maintained wild-type parasites as tachyzoites under bradyzoite induction culture conditions (pH 8.2/low CO 2 ), whereas the Δ pkac3 mutant failed to respond to the treatment. This suggests that Tg PKAc3 is the factor responsible for the cAMP-dependent tachyzoite maintenance. In addition, the Δ pkac3 mutant had a defect in the production of brain cysts in vivo , suggesting that a substrate of Tg PKAc3 is probably involved in the persistence of this parasite in the intermediate host animals. IMPORTANCE Toxoplasma gondii is one of the most prevalent eukaryotic parasites in mammals, including humans. Parasites can switch from rapidly replicating tachyzoites responsible for acute infection to slowly replicating bradyzoites that persist as a latent infection. Previous studies have demonstrated that T. gondii cAMP signaling can induce or suppress bradyzoite differentiation, depending on the strength and duration of cAMP signal. Here, we report that Tg PKAc3 is responsible for cAMP-dependent tachyzoite maintenance while suppressing differentiation into bradyzoites, revealing one mechanism underlying how this parasite transduces cAMP signals during differentiation.