Characterization of PfYTH.2 as an m6A-binding protein in Plasmodium falciparum

Post-transcriptional regulation of gene expression is central to the development and replication of the malaria parasite, Plasmodium falciparum, within its human host. The timely coordination of RNA maturation, homeostasis and protein synthesis relies on the recruitment of specific RNA-binding proteins to their cognate target mRNAs. One possible mediator of such mRNA-protein interactions is the N6-methylation of adenosines (m6A), a prevalent mRNA modification of parasite mRNA transcripts. Here, we used RNA protein pull-downs, RNA modification mass-spectrometry, and quantitative proteomics to identify two YTH-domain proteins (PfYTH.1 and PfYTH.2) as bona fide m6A-binding proteins during parasite blood-stage development. Interaction proteomics revealed that PfYTH.2 associates with the translation machinery, including multiple subunits of the eukaryotic initiation factor 3 (eIF3) and poly(A)-binding proteins. Furthermore, knock-sideways coupled with ribosome profiling showed that PfYTH.2 is essential for parasite survival and is a repressor of mRNA translation. Together, these data reveal an important missing link in the m6A-mediated mechanism controlling mRNA translation in a unicellular eukaryotic pathogen.