Phosphorylation in the Plasmodium falciparum Proteome: A Meta-Analysis of Publicly Available Data Sets

Malaria is a deadly disease caused by Apicomplexan parasites of the Plasmodium genus. Several species of the Plasmodium genus are known to be infectious to humans, of which P. falciparum is the most virulent. Post-translational modifications (PTMs) of proteins coordinate cell signaling and hence regulate many biological processes in P. falciparum homeostasis and host infection, of which the most highly studied is phosphorylation. Phosphosites on proteins can be identified by tandem mass spectrometry (MS) performed on enriched samples (phosphoproteomics), followed by downstream computational analyses. We have performed a large-scale meta-analysis of 11 publicly available phosphoproteomics data sets to build a comprehensive atlas of phosphosites in the P. falciparum proteome, using robust pipelines aimed at strict control of false identifications. We identified a total of 26,609 phosphorylated sites on P. falciparum proteins, split across three categories of data reliability (gold/silver/bronze). We identified significant sequence motifs, likely indicative of different groups of kinases responsible for different groups of phosphosites. Conservation analysis identified clusters of phosphoproteins that are highly conserved and others that are evolving faster within the Plasmodium genus, and implicated in different pathways. We were also able to identify over 180,000 phosphosites within Plasmodium species beyond falciparum, based on orthologue mapping. We also explored the structural context of phosphosites, identifying a strong enrichment for phosphosites on fast-evolving (low conservation) intrinsically disordered regions (IDRs) of proteins. In other species, IDRs have been shown to have an important role in modulating protein-protein interactions, particularly in signaling, and thus warranting further study for their roles in host-pathogen interactions. All data have been made available via UniProtKB, PRIDE, and PeptideAtlas, with visualization interfaces for exploring phosphosites in the context of other data on Plasmodium proteins.