Abstract
The mechanisms of stage-specific gene regulation in the malaria parasite Plasmodium falciparum are largely unclear, with only a small number of specific regulatory transcription factors (AP2 family) having been identified. In particular, the transcription factors that function in the intraerythrocytic stage remain to be elucidated. Previously, as a model case for stage-specific transcription in the P. falciparum intraerythrocytic stage, we analyzed the transcriptional regulation of pf1-cys-prx, a trophozoite/schizont-specific gene, and suggested that some nuclear factors bind specifically to the cis-element of pf1-cys-prx and enhance transcription. In the present study, we purified nuclear factors from parasite nuclear extract by 5 steps of chromatography, and identified a factor termed PREBP. PREBP is not included in the AP2 family, and is a novel protein with four K-homology (KH) domains. The KH domain is known to be found in RNA-binding or single-stranded DNA-binding proteins. PREBP is well conserved in Plasmodium species and partially conserved in phylum Apicomplexa. To evaluate the effects of PREBP overexpression, we used a transient overexpression and luciferase assay combined approach. Overexpression of PREBP markedly enhanced luciferase expression under the control of the pf1-cys-prx cis-element. These results provide the first evidence of a novel transcription factor that activates the gene expression in the malaria parasite intraerythrocytic stage. These findings enhance our understanding of the evolution of specific transcription machinery in Plasmodium and other eukaryotes.
Highlights
Plasmodium falciparum is a significant life-threatening parasitic pathogen that causes falciparum malaria in humans [1]
In the pf1-cys-prx, a trophozoite/schizont stage-specific recruitment of PfGCN5 histone acetyltransferase to the putative enhancer region was observed. These results suggest that P. falciparum possesses a sophisticated system of transcriptional regulation during the intraerythrocytic stages that is managed by coordinated interactions of unique cis-elements, trans-acting factors and chromatin modifications
Purification of Cis-Element Binding Protein In order to identify PRE Binding Protein (PREBP), we set up a huge-scale protein purification scheme using 561011 parasite cells obtained from the culture of 33 liters
Summary
Plasmodium falciparum is a significant life-threatening parasitic pathogen that causes falciparum malaria in humans [1]. In spite of years of intensive research, no effective vaccine is presently available, and existing antimalarial drugs are becoming less effective because of the rapid emergence of drug-resistant parasites [2]. The mechanisms of stage-specific gene regulation remain mostly unclear and only a small number of regulatory transcription factors (TFs) have been predicted in the Plasmodium genome [3,4]. Twenty-six AP2-related genes were predicted in the P. falciparum genome. In a rodent malaria model, it was revealed that some of the AP2 family proteins regulated specific gene expression in the ookinate stage [8], the sporozoite stage [9] and the liver stage [10]. The functions of TF in the intraerythrocytic cell cycle remain to be elucidated at the present time
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