m ⁶ A positions polyadenylation in Plasmodium falciparum

N ⁶ -methyladenosine (m ⁶ A) is the most abundant modification in eukaryotic mRNA and can influence gene expression, but the molecular mechanisms by which it does this at an individual gene level are poorly understood. The eukaryote with the highest known percentage of adenosine in its RNA is P. falciparum , causative agent of the deadliest form of malaria. P. falciparum has a high genomic AT content (>80% A+T) and a particular A-bias in mRNA - coding strands have an average adenosine content of 44%. Plasmodium parasites employ extensive transcriptional and post-transcriptional control over gene expression to differentiate within and between sexual and asexual stages in response to changing environments and hosts. We used direct Nanopore RNA sequencing and GLORI-seq to interrogate m ⁶ A methylation of the asexual Plasmodium falciparum transcriptome, revealing m ⁶ A depletion in protein coding regions relative to the 3’UTR, where m ⁶ A is deposited in precise patterns approximately 50 nt upstream of the polyadenylation site. We used an inducible protein mislocalisation system to disrupt the methyltransferase, which writes m ⁶ A to mRNA, and observed m ⁶ A depletion associated with transcriptional readthrough and chimeric transcripts. We show that m ⁶ A in transcript 3’UTRs is required for faithful positioning of polyadenylation and transcription termination. This work highlights the importance of m ⁶ A to mRNA processing in P. falciparum , and potentially a wider role in post-transcriptional regulation.