Eukaryote-Conserved Methylarginine Is Absent in Diplomonads and Functionally Compensated inGiardia

Abstract

Methylation is a common post-translational modification of arginine and lysine in eukaryotic proteins. Methylproteomes are best characterised for higher eukaryotes, where they are functionally expanded and evolved complex regulation. However, this is not the case for protist species evolved from the earliest eukaryotic lineages. Here, we integrated bioinformatic, proteomic and drug-screening datasets to comprehensively explore the methylproteome of Giardia duodenalis - a deeply-branching parasitic protist. We demonstrate Giardia and related diplomonads lack arginine-methyltransferases and have remodelled conserved RGG/RG motifs targeted by these enzymes. We also provide experimental evidence for methylarginine absence in proteomes of Giardia, but readily detect methyllysine. We bioinformatically infer 11 lysine-methyltransferases in Giardia, including highly-diverged SET-domain proteins with reduced domain architectures, and novel annotations demonstrating conserved methyllysine regulation of eukaryote Elongation Factor 1 alpha (eEF1a). Using mass spectrometry, we identify more than 200 methyllysine sites in Giardia, including in species-specific gene families involved in cytoskeletal regulation, enriched in coiled-coil features. Finally, we use known methylation-inhibitors to show that methylation plays key roles in replication and cyst formation in this parasite. This study highlights reduced methylation enzymes, sites and functions early in eukaryote evolution, including absent methylarginine networks in the Diplomonadida. These results challenge the view that arginine methylation is eukaryote-conserved, and demonstrates functional compensation of methylarginine was possible preceding expansion and diversification of these key networks in higher eukaryotes.

MOB: Pivotal Conserved Proteins in Cytokinesis, Cell Architecture and Tissue Homeostasis

“…This research also demonstrated that TbMOB1A interacts with the T. brucei NDR kinase TbPK50, a protein with high homology to the T. gondii NDR kinases. More recently, MOB proteins were also detected and are conserved in other kinetoplastids, i.e., Trypanosoma rangeli, Trypanosoma cruzi, and Leishmania major [160,161] and in the fornicatan Giardia [162][163][164]. Conserved Mob1 genes have also been identified in Trichomonas vaginalis and Entamoeba histolytica, protozoans belonging to the phyla Parabasalia and Amoebozoa, respectively [165,166].…”

ROS inhibits RORα degradation by decreasing its arginine methylation in liver cancer

“…Protein arginine and lysine residues are commonly methylated in eukaryotic cells. 27 Arginine methylation is an influential PTM that occurs on nonhistone and histone proteins, affecting their interactions, such as protein‐protein and protein–nucleic acid interactions. 28 In mammals, nine enzymes, PRMTs 1‐9, promote arginine methylation.…”

Non-histone protein methylation inTrypanosoma cruziepimastigotes

“…Our analysis of amino acids patterns surrounding methylation sites reveal different residues in K-Me and R-Me. The vicinity of lysine methylated sites are enriched for glutamic acids (E/Glu), something also seen in P. falciparum [21] and Giardia duodenalis [22] and arginine methylated sites are enriched upstream for arginine (R/Arg) and downstream for methionine (M/Met). Well-defined patterns of amino acids, such “RGG”, are not always surrounding targets of PRMTs [57] and, likewise, are not always defined for PKMT [58].…”

“…In Plasmodium falciparum , arginine [21] and lysine [22] methylated proteins are involved in diverse biological pathways, for example, RNA metabolism, protein synthesis, transport, proteolysis, protein folding and chromatin organization. In the deep-branching Giardia duodenalis [23], interestingly, methyl-arginines (and arginine-methyltransferases) are absent, but their biological functions appear to be compensated by methyl-lysines, which are regulated between its different life-cycle stages. For the TriTryps, global studies about protein methylation are only available for Leishmania and T. brucei .…”