Protein kinase A negatively regulates Ca2+ signalling in Toxoplasma gondii

Abstract

The phylum Apicomplexa comprises a group of obligate intracellular parasites that alternate between intracellular replicating stages and actively motile extracellular forms that move through tissue. Parasite cytosolic Ca2+ signalling activates motility, but how this is switched off after invasion is complete to allow for replication to begin is not understood. Here, we show that the cyclic adenosine monophosphate (cAMP)-dependent protein kinase A catalytic subunit 1 (PKAc1) of Toxoplasma is responsible for suppression of Ca2+ signalling upon host cell invasion. We demonstrate that PKAc1 is sequestered to the parasite periphery by dual acylation of PKA regulatory subunit 1 (PKAr1). Upon genetic depletion of PKAc1 we show that newly invaded parasites exit host cells shortly thereafter, in a perforin-like protein 1 (PLP-1)-dependent fashion. Furthermore, we demonstrate that loss of PKAc1 prevents rapid down-regulation of cytosolic [Ca2+] levels shortly after invasion. We also provide evidence that loss of PKAc1 sensitises parasites to cyclic GMP (cGMP)-induced Ca2+ signalling, thus demonstrating a functional link between cAMP and these other signalling modalities. Together, this work provides a new paradigm in understanding how Toxoplasma and related apicomplexan parasites regulate infectivity.

Calcium negatively regulates secretion from dense granules in Toxoplasma gondii

“…We note that phosphodiesterase inhibitors have been reported to elevate cAMP in addition to cGMP in Plasmodium (Howard et al, ). In Toxoplasma , cAMP is implicated in reducing cytosolic Ca 2+ and, although we do not know if any similar elevation of cAMP occurs with BIPPO/zaprinast treatment, the observed increase in cytosolic Ca 2+ in Toxoplasma with these agents suggest that cAMP is unlikely to be a major contributor to this response (Jia et al, ; Stewart et al, ; Uboldi et al, ).…”

“…Upon successful entry of parasites into their host cell, all of this activation for MIC release and motility must then be supressed in order for the rhoptry‐ and dense granule‐mediated intracellular events to progress. Recent work suggests that cAMP‐signalling that activates protein kinase A (PKA) is involved in reducing cytosolic Ca 2+ levels upon host cell entry and, in turn, reversing the processes that led to MIC secretion (Jia et al, ; Uboldi et al, ).…”

Protein O-fucosyltransferase 2–mediated O-glycosylation of the adhesin MIC2 is dispensable for Toxoplasma gondii tachyzoite infection

“…We used [K ϩ ] shift to promote synchronous invasion whereas the Bandini et al study (62) did not. [K ϩ ] shift strongly promotes cytosolic Ca 2ϩ signaling and microneme secretion, resulting in synchronous motility and invasion (47,48). It could be that these differences in conditions are enough to subtly effect invasion efficiency and thus explain the differences in the two studies.…”

Signaling Cascades Governing Entry into and Exit from Host Cells by Toxoplasma gondii

“…gondii encodes three PKAc subunits (PKAc1-3) and only one PKAr subunit. PKAc1 has recently been characterized in T. gondii, and its binding to PKAr has been confirmed (68, 125,129). PKAc1 localizes to the parasite periphery upon binding of the double dually acylated PKAr by myristoylation and palmitoylation, and it is released soluble and active upon increase in cAMP concentration (68).…”