The genetic landscape of origins of replication in P. falciparum
A variety of origin mapping approaches have enabled genome-wide identification of origins of replication (ORI) in model organisms but only a limited number of studies have focused on divergent organisms. Here we employed three approaches to map ORIs in Plasmodium falciparum, the deadliest human malaria parasite. We mapped the distribution of ORC binding sites by ChIP-seq and active ORIs using two strategies: Fork-seq and SNS-seq. We show that ORIs lack sequence specificity but are not randomly distributed, and group in initiation zones or clusters. Licensing is biased towards regions of higher GC content and associated with G-quadruplex forming sequences (G4FS).   While strong transcription likely enhances origin firing, active origins are depleted from transcription start sites. Instead, most accumulate in transcriptionally active gene bodies. With the exception of centromeres and telomeres, fork speed is similar across the genome and in different moments of schizogony. Single molecule analysis of nanopore reads containing multiple initiation events, which could have only come from individual replicating nuclei, showed a relationship between the pace at which replication forks travel and the distance to the nearest origin. While some similarities could be drawn with the canonic eukaryote model, the distribution of origins of replication in P. falciparum is likely shaped by unique genomic features such as extreme AT-richness – a product of the evolutionary pressure imposed by the parasitic lifestyle. Our multifaceted approach provides a holistic understanding of the genetic and epigenetic makeup of the origins of replication in malaria parasites.