Deciphering essential roles of cAMP signalling during malaria parasite transmission

Abstract

Intracellular messengers such as cyclic guanosine phosphate (cGMP) and cyclic-adenosine phosphate (cAMP), mediate cellular responses to extracellular stimuli in both free-living and pathogenic eukaryotes. During its complex life-cycle, the malaria parasite experiences extensive environmental changes where accurate responses to changing host and vector environments are critical for both establishing infection and successful transmission. Several studies have demonstrated the importance of cGMP signalling during the parasite’s life cycle, including the transition from the mammalian host to the mosquito vector. However, signalling through cAMP is poorly understood during this critical step of parasite transmission. The chief effector of cAMP signalling is Protein Kinase A (PKA), which consists of a regulatory (PKAr) and catalytic subunit (PKAc). Using a range of genetic engineering methods (promoter-swap and the DiCre-loxP system) we generated PKAr-KO parasites across different mosquito life stages revealing the essential role of this enzyme during mitosis. We found that in the absence of PKAr, parasites failed to infect the mosquito as the sexual precursor cells (gametocytes) were infertile.

Additionally, we found a second critical role for PKAr during the process of sporogony where in the absence of PKAr, parasites failed to develop into fully mature mammalian infective forms (sporozoites). High-resolution imaging techniques revealed PKA is involved in the establishment of bipolar mitotic spindles, stability of microtubules and cytokinesis. Furthermore, we characterized the role of a second putative cAMP binding protein, Exchange Protein Activated by cAMP (EPAC), during parasite colonization of the mosquito midgut and we show that its cyclic nucleotide binding capability controls ookinete invasion. This data demonstrates the importance of cAMP signalling across multiple transmission stages of the malaria parasite and increases our understanding of the molecular mechanisms that govern parasite transmission.