Cell cycle control in Plasmodium falciparum: a genomics perspective

The molecular mechanisms regulating cell proliferation and development in malaria parasites are still largely unknown. Phenomenological observations, pertaining to the organisation of the cell cycle during schizogony or to the signal transduction pathways whose activation is responsible for the developmental stage transitions, can now be complemented with information gathered from genomic databases. The PlasmoDB database has been used extensively to identify putative homologues of a number of eukaryotic cell cycle regulators such as cyclins, cyclin-dependent kinases, factors involved in the control of DNA synthesis, and components of signal transduction pathways. However, gene identification based on sequence homology is limited by the fact that any Plasmodium-specific functional homologue will be missed by this approach. Furthermore, experimental data indicate that the structure of some regulatory pathways (unlike that of metabolic pathways) cannot be deducted directly from database mining. Because of these limitations in the direct exploitation of genomic database, elucidation of the organisation of the signal transduction and cell cycle machineries requires experimental, proteomics-based approaches such as the characterisation of protein complexes containing cell cycle regulators and the establishment of a map of protein-protein interactions involving these elements

Structure-activity relationships and inhibitory effects of various purine derivatives on the in vitro growth of Plasmodium falciparum

The development of novel chemotherapeutic agents has become an urgent task due to the development and rapid spread of drug resistance in Plasmodium falciparum, the protozoan parasite responsible for cerebral malaria. Cyclin-dependent kinases (CDKs) are essential for the regulation of the eukaryotic cell cycle, and several enzymes of this family have been identified in P. falciparum. In recent years, a number of purine-derived kinase inhibitors have been synthesised, some of which display selective activity against CDKs. This report describes a study in which various purine derivatives were screened for in vitro antimalarial activity. The erythrocytic asexual stages of the chloroquine-resistant P. falciparum strain (FCR-3) were cultivated in vitro in the presence of the various purines, and their effect on parasite proliferation was determined by the [3H]hypoxanthine incorporation assay. Our results show considerable variation in the sensitivity of P. falciparum to the different purines, as well as a general independence from their effect on purified starfish CDK1/cyclin B activity, which has been the standard assay used to identify CDK-specific inhibitors. Two subfamilies of purines with moderate to poor activity against CDK1/cyclin B activity showed submicromolar activity against P. falciparum. Structure-activity analysis indicates that certain structural features are associated with increased activity against P. falciparum. These features can be exploited to synthesise compounds with higher activity and specificity towards P. falciparum.

Elsevier2001

The cell cycle of parasitic protozoa: potential for chemotherapeutic exploitation

There is an urgent need to develop new drugs against eukaryotic parasitic protozoa such as Plasmodium, Trypanosoma and Leishmania, which cause the diseases malaria, trypanosomiasis and the leishmaniases respectively. The biology of these organisms has many unusual facets that might be exploited for drug design, and the recent availability of parasite genome sequence data has facilitated the search for novel drug targets. Here we review current understanding of the cell cycle in these parasites and show that important structural and functional differences exist between parasite and mammalian cell cycle control machineries and signal transduction pathways, which might be utilised for rational drug design. Potential targets include protein kinases from the cyclin-dependent kinase, cAMP-dependent kinase and mitogen activated protein kinase families.

2003

A Plasmodium falciparum transcriptional CDK-related kinase with a crucial role in parasite proliferation associates with histone deacetylase activity

Cyclin-dependent protein kinases (CDKs) are key regulators of the eukaryotic cell cycle and of the eukaryotic transcription machinery. Here, we report the characterisation of Pfcrk-3 (P. falciparum CDK-related kinase 3, PlasmoDB identifier PFD0740w), an unusually large CDK-related protein whose kinase domain displays maximal homology to those CDKs which are, in other eukaryotes, involved in the control of transcription. The closest enzyme in yeast is BUR1 ("Bypass Upstream activating sequence Requirement"), known to control gene expression through interaction with chromatin modification enzymes. Consistent with this, immunofluorescence data show that Pfcrk-3 colocalises with histones. We show that recombinant Pfcrk-3 associates with histone H1 kinase activity in parasite extracts, and that this association is detectable even if the catalytic domain of Pfcrk-3 is rendered inactive by site-directed mutagenesis, indicating that Pfcrk-3 is part of a complex that includes other protein kinases. Immunoprecipitates obtained from extracts of transgenic parasites expressing HA-tagged Pfcrk-3 using a hemagglutinin (HA) antibody displayed both protein kinase and histone deacetylase activities. Reverse genetics data show that the pfcrk-3 locus can be targeted only if the genetic modification does not cause loss-of-function. Taken together, our data strongly suggests that Pfcrk-3 fulfils a crucial role in the intraerythrocytic development of P. falciparum, presumably through chromatin modification-dependent regulation of gene expression.

American Society for Microbiology2010