Abstract
BackgroundThe malaria parasite Plasmodium falciparum has an unusually euchromatic genome with poorly conserved positioning of nucleosomes in intergenic sequences and poorly understood mechanisms of gene regulation. Variant histones and histone modifications determine nucleosome stability and recruit trans factors, but their combinatorial contribution to gene regulation is unclear.ResultsHere, we show that the histone H3 acetylations H3K18ac and H3K27ac and the variant histone Pf H2A.Z are enriched together at regulatory sites upstream of genes. H3K18ac and H3K27ac together dynamically mark regulatory regions of genes expressed during the asexual life cycle. In contrast, H3K4me1 is depleted in intergenic sequence and dynamically depleted upstream of expressed genes. The temporal pattern of H3K27ac and H3K18ac enrichment indicates that they accumulate during S phase and mitosis and are retained at regulatory sequences until at least G1 phase and after cessation of expression of the cognate genes. We integrated our ChIPseq data with existing datasets to show that in schizont stages H3K18ac, H3K27ac and Pf H2A.Z colocalise with the transcription factor PfAP2-I and the bromodomain protein PfBDP1 and are enriched at stably positioned nucleosomes within regions of exposed DNA at active transcriptional start sites. Using transient transfections we showed that sequences enriched with colocalised H3K18ac, H3K27ac and Pf H2A.Z possess promoter activity in schizont stages, but no enhancer-like activity.ConclusionsThe dynamic H3 acetylations define P. falciparum regulatory sequences and contribute to gene activation. These findings expand the knowledge of the chromatin landscape that regulates gene expression in P. falciparum.
Highlights
The malaria parasite Plasmodium falciparum has an unusually euchromatic genome with poorly conserved positioning of nucleosomes in intergenic sequences and poorly understood mechanisms of gene regulation
We demonstrate that dynamic enrichment of H3K18ac and H3K27ac at Pf H2A.Z containing nucleosomes at transcriptional start site (TSS) correlates with gene expression levels, whereas H3K4me1 is generally absent from intergenic sequences and depleted from these putative regulatory elements
Levels of H3K4me1 and Pf H2A.Z remain constant across the intra‐erythrocytic development cycle (IDC) but levels of H3K18ac and H3K27ac vary The antibodies we used to investigate the chromatin marks included a Pf H2A.Z antiserum of confirmed specificity [50] and commercial antibodies to H3K4me1, H3K27ac and H3K18ac that were previously validated for chromatin immunoprecipitation (ChIP) (Antibody Validation Database) [51], (Histone Antibody Specificity Database) [52]
Summary
The malaria parasite Plasmodium falciparum has an unusually euchromatic genome with poorly conserved positioning of nucleosomes in intergenic sequences and poorly understood mechanisms of gene regulation. P. falciparum differs from other eukaryotes in the unusually high AT content of its genome (90% in intergenic regions) [1], its unique variant histones [2] and the high proportion of its genome that it maintains as euchromatin during its pathogenic, intra-erythrocytic lifecycle [3]. The majority of P. falciparum genes are regulated during its haploid, 48-h intra-erythrocytic developmental cycle (IDC) [5, 6] and groups of Plasmodium genes are activated by stage-specific transcription factors [7]. In at least one instance a stage-specific transcription factor [8] requires the presence of the P. falciparum bromodomain protein 1 (PfBDP1) to activate a subset of erythrocyte invasion genes [9]. The many possible combinations between chromatin-binding proteins and specific transcription factors may explain the tightly regulated, dynamic transcription of most P. falciparum genes
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