Multiple short windows of calcium-dependent protein kinase 4 activity coordinate distinct cell cycle events during Plasmodium gametogenesis

Abstract

Malaria transmission relies on the production of gametes following ingestion by a mosquito. Here, we show that Ca2+-dependent protein kinase 4 controls three processes essential to progress from a single haploid microgametocyte to the release of eight flagellated microgametes in Plasmodium berghei. A myristoylated isoform is activated by Ca2+ to initiate a first genome replication within twenty seconds of activation. This role is mediated by a protein of the SAPS-domain family involved in S-phase entry. At the same time, CDPK4 is required for the assembly of the subsequent mitotic spindle and to phosphorylate a microtubule-associated protein important for mitotic spindle formation. Finally, a non-myristoylated isoform is essential to complete cytokinesis by activating motility of the male flagellum. This role has been linked to phosphorylation of an uncharacterised flagellar protein. Altogether, this study reveals how a kinase integrates and transduces multiple signals to control key cell-cycle transitions during Plasmodium gametogenesis.DOI: http://dx.doi.org/10.7554/eLife.26524.001

A divergent cyclin/cyclin-dependent kinase complex controls the atypical replication of a malaria parasite during gametogony and transmission

“…By indirect immunofluorescence of fixed cells, we confirmed the pattern of SOC2-HA distribution ( Fig. 3F), as previously described 19 .…”

“…The set of enriched GO terms associated with CRK5-regulation is similar to those associated with calcium dependent protein kinase (CDPK) 4 and CRK4 ( Fig. S2), two kinases previously shown to be crucial for DNA replication and its initiation during gametogony 19 and schizogony 23 , respectively.…”

“…Plasmodium gametogony 19,20 . As multiple dynein-related proteins are phosphorylated in a CRK5dependent manner, we quantified the formation of spindle-like structures and axonemes in CRK5-KO parasites.…”

“…Current evidence suggests that certain canonical cell-cycle checkpoints are absent 16 . For example, compounds that interfere with mitotic spindle formation do not prevent DNA replication proceeding 17,18 , while spindle formation is not affected in a mutant that is unable to replicate DNA 18,19 . Recently, we observed that proteins involved in DNA replication and cytoskeletal reorganisation are phosphorylated during the first seconds of gametogony 20 .…”

“…Analysis of CRK5 immunoprecipitates also identified multiple peptides from PBANKA_082440 (CDK regulatory subunit, CDKrs), a protein related to Cks1 and CksHs2, two CDK-associated proteins in Saccharomyces cerevisiae 24 and human 25 , respectively. To confirm the putative interaction between CRK5, SOC2 and CDKrs, we tagged both SOC2 and CDKrs at the C-terminus by integration of 3xHA 19 or GFP coding sequence into the endogenous gene ( Fig. S3B) and affinity purified the tagged proteins.…”