Transcriptional analysis of genes encoding enzymes of the folate pathway in the human malaria parasite Plasmodium falciparum.
Folate metabolism in Plasmodium falciparum is essential for cell growth and replication, and the target of important antimalarial agents. The pathway comprises a series of enzymes that convert GTP to derivatives of tetrahydrofolate, which are cofactors in one-carbon transfer reactions. We investigated the expression of five of the genes encoding these enzymes by quantitative reverse transcription-polymerase chain reaction (qRT-PCR) using a threshold detection technique. We followed changes in mRNA levels as parasites progress through the erythrocytic cell cycle and examined this process in two cloned lines of diverse origins, as well as under stress conditions, induced by either removal of important metabolites or challenge by folate enzyme inhibitors. Although conventionally regarded as performing housekeeping functions, these genes show disparate levels of and changes in expression through the cell cycle, but respond quite uniformly to folate pathway-specific stress factors, with no evidence of feedback at the transcriptional level. Overall, the two genes involved in the thymidylate cycle (encoding dihy-drofolate reductase-thymidylate synthase, dhfr-ts, and serine hydroxymethyltransferase, shmt) gave the most abundant transcripts. However, only the latter showed major variation across the cell cycle, with a peak around the time of onset of DNA replication, possibly indicative of a regulatory function.
D000818:Animals; D000962:Antimalarials / Q000494:pharmacology; D004338:Drug Combinations; D004912:Erythrocytes / Q000469:parasitology; D005492:Folic Acid / Q000378:metabolism*; D012696:Glycine Hydroxymethyltransferase / Q000235:genetics / Q000378:metabolism; D006801:Humans; D016778:Malaria, Falciparum / Q000469:parasitology; D009097:Multienzyme Complexes / Q000235:genetics / Q000378:metabolism; D010963:Plasmodium falciparum / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development; D015800:Protozoan Proteins / Q000235:genetics* / Q000378:metabolism*; D011739:Pyrimethamine / Q000494:pharmacology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013413:Sulfadoxine / Q000494:pharmacology; D013762:Tetrahydrofolate Dehydrogenase / Q000235:genetics / Q000378:metabolism; D013940:Thymidylate Synthase / Q000235:genetics / Q000378:metabolism; D014158:Transcription, Genetic
D000962:Antimalarials; D004338:Drug Combinations; D009097:Multienzyme Complexes; D015800:Protozoan Proteins; C056930:thymidylate synthase-dihydrofolate reductase; C001205:fanasil, pyrimethamine drug combination; D013413:Sulfadoxine; D005492:Folic Acid; D013762:Tetrahydrofolate Dehydrogenase; D013940:Thymidylate Synthase; D012696:Glycine Hydroxymethyltransferase; D011739:Pyrimethamine