Musabyimana, J.-P. | bioRxiv Science

bioRxiv2024

Plasmodium falciparum SET10 is a histone H3 lysine K18 methyltransferase that participates in a chromatin modulation network crucial for intraerythrocytic development

Lifecycle progression of the malaria parasite Plasmodium falciparum requires precise tuning of gene expression including histone methylation. The histone methyltransferase PfSET10 was previously described as a H3K4 methyltransferase involved in var gene regulation, making it a prominent antimalarial target. In this study, we investigate the role of PfSET10 in the blood stages of P. falciparum in more detail, using tagged PfSET10-knockout (KO) and -knockdown (KD) lines. We demonstrate a nuclear localization of PfSET10 with peak protein levels in schizonts. PfSET10 deficiency results in reduced intraerythrocytic growth, but has no effect on gametocyte formation. When the PfSET10-KO line is screened for histone methylation variations, lack of PfSET10 renders the parasites unable to mark H3K18me1, while no significant changes in the H3K4 methylation status are observed. Comparative transcriptomic profiling of PfSET10-KO schizonts demonstrates the upregulation of transcripts particularly encoding proteins linked to erythrocyte invasion and multigene family proteins, suggesting a repressive function of the histone methylation mark. TurboID coupled with mass spectrometry further reveals an extensive nuclear PfSET10 interaction network with roles in transcriptional regulation, DNA replication and repair, chromatin remodeling and mRNA processing. Main interactors of PfSET10 include ApiAP2 transcription factors, chromatin modulators like PfMORC and PfISWI, mediators of RNA polymerase II, and DNA replication licensing factors. The combined data pinpoint PfSET10 as a histone H3 lysine K18 methyltransferase of the P. falciparum blood stages that regulates nucleic acid metabolic processes as part of a comprehensive chromatin modulation network. ImportanceThe fine-tuned regulation of DNA replication and transcription is particularly crucial for the rapidly multiplying blood stages of malaria parasites and proteins involved in these processes represent important drug targets. This study demonstrates that contrary to previous reports the histone methyltransferase PfSET10 of the malaria parasite Plasmodium falciparum methylates histone 3 at lysine K18, a histone mark to date not well understood. Deficiency of PfSET10 due to genetic knockout affects genes involved in intraerythrocytic development. Furthermore, in the nuclei of blood stage parasites, PfSET10 interacts with various protein complexes crucial for DNA replication, remodeling and repair, as well as for transcriptional regulation and mRNA processing. In summary, this study highlights PfSET10 as a H3K18 methyltransferase with critical functions in chromatin maintenance during the development of P. falciparum in red blood cells.

microbiology↗

bioRxiv2023

Proximity interaction analysis of the Plasmodium falciparum putative ubiquitin ligase PfRNF1 reveals a role in RNA regulation

Some proteins have acquired both ubiquitin ligase activity and RNA-binding properties and are therefore known as RNA-binding Ubiquitin ligases (RBULs). These proteins provide a link between the RNA metabolism and the ubiquitin proteasome system (UPS). The UPS is a crucial protein surveillance system of eukaryotes primarily involved in the selective proteolysis of proteins which are covalently marked with ubiquitin through a series of steps involving ubiquitin E1 activating, E2 conjugating and E3 ligating enzymes. The UPS also regulates other key cellular processes such as cell cycle, proliferation, cell differentiation, transcription and signal transduction. While RBULs have been characterized in other organisms, little is known about their role in Plasmodium falciparum, the causative agent of the deadliest human malaria, malaria tropica. In this study, we characterized a previously identified putative P. falciparum RING finger E3 ligase PfRNF1. We show that the protein is highly expressed in sexual stage parasites and mainly present in immature male gametocytes. Using proximity interaction studies with parasite lines expressing PfRNF1 tagged with the Biotin ligase BirA, we identified an interaction network of PfRNF1 in both the asexual blood stages and gametocytes composed mainly of ribosomal proteins, RNA-binding proteins including translational repressors such DOZI, CITH, PUF1 and members of the CCR4-NOT complex, as well as proteins of the UPS such as RPN11, RPT1 and RPT6. Our interaction network analysis reveals PfRNF1 as a potential RNA-binding E3 ligase which links RNA dependent processes with protein ubiquitination to regulate gene expression. ImportanceRBULs provide a link between RNA-mediated processes with the ubiquitin system. Only a few RBULs have been identified and none has been characterized in the malaria parasite P. falciparum. In this study, we unveiled the interactome of the putative P. falciparum E3 ligase PfRNF1. We show that PfRNF1 interacts with both proteins of the ubiquitin system as well as RNA-binding proteins therefore indicating that it is a putative RBUL which links RNA regulation with the ubiquitin system in P. falciparum.

cell biology↗