Identifying important regulators of transcription in Plasmodium falciparum

© 2014 Dr. Gabrielle JoslingIn the malaria parasite Plasmodium falciparum, many key pathogenic processes are regulated through epigenetic mechanisms. One of the best studied of these mechanisms is the post-translational modification of histones. This project aims to investigate the role of proteins that recognise acetylated histones, as this modification is unusually abundant in P. falciparum and is generally positively correlated with transcription. In the first part of this work, a novel parasite bromodomain protein (Bromodomain protein 1, or BDP1) was investigated. The bromodomain is a domain involved in binding to acetylated histones, and thus bromodomain proteins often play a role in regulating transcription. Conditional knockdown of BDP1 results in a slight lag in growth as well as a dramatic decrease in multiplication rate which is likely due to a defect in invasion. BDP1 knockdown results in downregulation of invasion genes, which likely accounts for this invasion defect. Members of several clonally variant gene families were also deregulated, suggesting that BDP1 also plays a role in regulating these genes. To further characterise BDP1, chromatin immunoprecipitation was performed to identify genes that BDP1 is associated with. This revealed that BDP1 is enriched near the transcriptional start sites of genes involved in invasion when these genes are expressed. It is not enriched at the transcriptional start sites of other expressed genes, indicating BDP1 directly regulates a subset of genes that includes invasion genes. Taken together, these data strongly suggest that BDP1 regulates the expression of invasion genes. The effect of bromodomain inhibitors on parasite growth was also investigated. While an inhibitor of a mammalian bromodomain protein only weakly inhibited parasite growth, a putative inhibitor of a P. falciparum bromodomain protein was much more effective. These data demonstrate that bromodomain inhibitors have potential as a class of novel antimalarial drugs. Finally, a protocol to identify proteins that bind modified histones was developed and used to identify additional proteins that bind acetylated histones. This method identified potential epigenetic regulators, including a second parasite-specific bromodomain protein. Overall, this work demonstrates the important role that proteins that recognise acetylated histones play in gene regulation in P. falciparum, highlights a potential new class of drug targets, and identifies interesting candidates for future investigation