Published in:
Open Access
01-12-2012 | Research
Plasmodium serine hydroxymethyltransferase: indispensability and display of distinct localization
Authors:
Wichai Pornthanakasem, Darin Kongkasuriyachai, Chairat Uthaipibull, Yongyuth Yuthavong, Ubolsree Leartsakulpanich
Published in:
Malaria Journal
|
Issue 1/2012
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Abstract
Background
Serine hydroxymethyltransferase (SHMT), a pyridoxal phosphate-dependent enzyme, plays a vital role in the de novo pyrimidine biosynthesis pathway in malaria parasites. Two genes have been identified in Plasmodium spp. encoding a cytosolic SHMT (cSHMT) and putative mitochondria SHMT (mSHMT), but their roles have not been fully investigated.
Methods
The presence of Plasmodium SHMT isoforms in the intra-erythrocytic stage was assessed based on their gene expression using reverse transcription PCR (RT-PCR). Localization studies of Plasmodium SHMT isoforms were performed by transfection of fluorescent-tagged gene constructs into P. falciparum and expressions of fluorescent fusion proteins in parasites were observed using a laser scanning confocal microscope. Genetic targeting through homologous recombination was used to study the essentiality of SHMT in Plasmodium spp.
Results
Semi-quantitative RT-PCR revealed the expression of these two genes throughout intra-erythrocytic development. Localization studies using P. falciparum expressing fluorescent-tagged SHMT showed that Pf cSHMT-red fluorescent fusion protein (Pf cSHMT-DsRed) is localized in the cytoplasm, while Pf mSHMT-green fluorescent fusion protein (Pf mSHMT-GFP) co-localized with Mitotracker™-labelled mitochondria as predicted. The essentiality of plasmodial cSHMT was inferred from transfection experiments where recovery of viable knock-out parasites was not achieved, unless complemented with a functional equivalent copy of shmt.
Conclusions
Distinct compartment localizations of Pf SHMT were observed between cytoplasmic and mitochondrial isoforms, and evidence was provided for the indispensable role of plasmodial cSHMT indicating it as a valid target for development of novel anti-malarials.