The genetic Ca2+ sensor GCaMP3 reveals multiple Ca2+ stores differentially coupled to Ca2+ entry in the human malaria parasite plasmodium falciparum

Abstract

Cytosolic Ca21 regulates multiple steps in the host-cell invasion, growth, proliferation, and egress of blood-stage Plasmodium falciparum, yet our understanding of Ca21 signaling in this endemic malaria parasite is incomplete. By using a newly generated transgenic line of P. falciparum (PfGCaMP3) that expresses constitutively the genetically encoded Ca21 indicator GCaMP3, we have investigated the dynamics of Ca21 release and influx elicited by inhibitors of the sarcoplasmic/endoplasmic reticulum Ca21-ATPase pumps, cyclopiazonic acid (CPA), and thapsigargin (Thg). Here we show that in isolated trophozoite phase parasites: (i) both CPA and Thg release Ca21 from intracellular stores in P. falciparum parasites; (ii) Thg is able to induce Ca21 release from an intracellular compartment insensitive to CPA; (iii) only Thg is able to activate Ca21 influx from extracellular media, through a mechanism resembling store-operated Ca21 entry, typical of mammalian cells; and (iv) the Thg-sensitive Ca21 pool is unaffected by collapsing the mitochondria membrane potential with the uncoupler carbonyl cyanide m-chlorophenyl hydrazone or the release of acidic Ca21 stores with nigericin. These data suggest the presence of two Ca21 pools in P. falciparum with differential sensitivity to the sarcoplasmic/endoplasmic reticulum Ca21-ATPase pump inhibitors, and only the release of the Thg-sensitive Ca21 store induces Ca21 influx. Activation of the store-operated Ca21 entry-like Ca21 influx may be relevant for controlling processes such as parasite invasion, egress, and development mediated by kinases, phosphatases, and proteases that rely on Ca21 levels for their activation.