Correlation between cyclin dependent kinases and artemisinin-induced dormancy in Plasmodium falciparum in vitro

Background Artemisinin-induced dormancy provides a plausible explanation for recrudescence following artemisinin monotherapy. This phenomenon shares similarities with cell cycle arrest where cyclin dependent kinases (CDKs) and cyclins play an important role. Methods Transcription profiles of Plasmodium falciparum CDKs and cyclins before and after dihydroartemisinin (DHA) treatment in three parasite lines, and the effect of CDK inhibitors on parasite recovery from DHA-induced dormancy were investigated. Results After DHA treatment, parasites enter a dormancy phase followed by a recovery phase. During the dormancy phase parasites up-regulate pfcrk1, pfcrk4, pfcyc2 and pfcyc4, and down-regulate pfmrk, pfpk5, pfpk6, pfcrk3, pfcyc1 and pfcyc3. When entering the recovery phase parasites immediately up-regulate all CDK and cyclin genes. Three CDK inhibitors, olomoucine, WR636638 and roscovitine, produced distinct effects on different phases of DHA-induced dormancy, blocking parasites recovery. Conclusions The up-regulation of PfCRK1 and PfCRK4, and down regulation of other CDKs and cyclins correlate with parasite survival in the dormant state. Changes in CDK expression are likely to negatively regulate parasite progression from G1 to S phase. These findings provide new insights into the mechanism of artemisinin-induced dormancy and cell cycle regulation of P. falciparum, opening new opportunities for preventing recrudescence following artemisinin treatment

<i>P</i>. <i>falciparum</i> gene names (abbreviation), ID numbers and primers used for amplification of CDKs and cyclins.

<p>The seryl-tRNA synthetase gene was used as a positive control.</p

Effect of the CDK inhibitors (WR636638, olomoucineon and roscovitine) on untreated and DHA-treated W2 parasites.

<p>Parasites were treated with DHA alone for 6 h, or one of the CDK inhibitors alone for 48 h (at Day 0), or DHA (for 6 h) plus one of the CDK inhibitors (for 48 h). For DHA plus CDK inhibitors, one of the CDK inhibitors was added either together with DHA (Day 0) or after DHA treatment (Days 2, 4 and 6) for 48 h. Two panels (Exp 1 and Exp 2) represent results of 2 independent experiments, each in triplicate. Parasitemia (percentages of parasitized erythrocytes by morphologically normal parasites) was determined by microscopy.</p

Dynamics of parasite density following DHA treatment of <i>P</i>. <i>falciparum</i> lines of W2, D6 and S55.

<p>Parasitemia are determined by SYBR Green staining and FACS analysis. Dotted lines indicate boundaries between dormancy and recovery phase in each parasite line.</p