Additional remarks phenotype | Mutant/mutation
The mutant expresses a C-terminal GFP-tagged version of CDKrs
Protein (function)
CDKrs is a protein related to Cks1 and CksHs2, two CDK-associated proteins in Saccharomyces cerevisiae and and human, respectively. In the paper evidence is presented for an interaction between CRK5, SOC2 and CDKrs
From the Abstract: 'Cell cycle transitions are generally triggered by variation in the activity of cyclindependent kinases (CDKs) bound to cyclins. Malaria-causing parasites have a life cycle with unique cell-division cycles, and a repertoire of divergent CDKs and cyclins of poorly understood function and interdependency. We show that Plasmodium berghei CDK-related kinase 5 (CRK5), is a critical regulator of atypical mitosis in the gametogony and is required for mosquito transmission. It phosphorylates canonical CDK motifs of components in the pre-replicative complex and is essential for DNA replication. During a replicative cycle, CRK5 stably interacts with a single Plasmodium specific cyclin (SOC2), although we obtained no evidence of SOC2 cycling by transcription, translation or degradation. Our results provide evidence that during Plasmodium male gametogony, this divergent cyclin/CDK pair fills the functional space of other eukaryotic cell-cycle kinases controlling DNA replication.'
Phenotype
Not analysed in detail. CDKrs-GFP in the nucleus of non-activated microgametocytes and throughout male gametogony, with a spindle-like localisation during mitoses, as found for CRK5. The mutant is used for mass-spec analyses of co-precipitated proteins in activated male gametocytes.
'To confirm that levels of CRK5 and CDKrs are maintained during gametogony in single cells, we monitored by flow cytometry the fluorescence of gametocytes expressing CRK5-GFP or CDKrs-GFP during gametogony. This analysis confirmed that there was no change in the level of CRK5-GFP or CDKrs-GFP fluorescence following male gametocyte activation and gametogony.'
Additional information
Evidence in this study is presented that:
- CRK5 is a key regulator of gametogony and sporogony in the mosquito
- Phosphoproteome kinetics point to direct phosphorylation of the pre-replicative complex by CRK5
- CRK5 is required for both S- and M- phases during P. berghei gametogony
- CRK5 is part of an atypical nuclear cyclin/CDK complex
- CRK5, SOC2 ((PBANKA_1442200)and CDKrs (PBANKA_0824400; (CDK regulatory subunit, CDKrs) have a similar location and complementary functions during gametogony. CDKrs is a protein related to Cks1 and CksHs2, two CDK-associated proteins in Saccharomyces cerevisiae and and human, respectively.
- CRK5, SOC2 and CDKrs have a similar location and complementary functions during gametogony
- SOC2 expression does not follow a temporal cyclin pattern during gametogony and the CSC complex (CRK5/SOC2/CDKrs complex) is stable during the first round of mitosis
- CSC is dynamically phosphorylated during the first round of replication
- Live fluorescence microscopy identified CDKrs-GFP in the nucleus of non-activated microgametocytes and throughout male gametogony, with a spindle-like localisation during mitoses, as found for CRK5
- Ultrastructural analysis by electron microscopy of SOC2-KO and CDKrs-KO activated gametocytes confirms an early arrest during male gametogony as observed in CRK5-KO parasites.
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