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GIARDIA DUODENALIS MRNA BOUND PROTEOME REVEALED THE EMERGENCE OF EUKARYOTIC SPECIFIC POST-TRANSCRIPTIONAL REGULATORY NETWORKS (ID 1094)
Abstract
Introduction
RNA binding proteins (RBPs) are major post-transcriptional regulators (PTR). In higher eukaryotes RBPs control transcription, RNA transport, splicing and degradation, translation and translational repression and play key roles in cell fating, pluripotency and differentiation. Surprisingly, the eukaryotic RBPome is fundamentally unchanged from yeast to humans. This suggests many novel RBPs emerged in basal eukaryotes. Yet this is largely unstudied. Here, we characterised the RBPome of G. duodenalis, which belongs to one of the earliest branching eukaryotic lineages, and used this to understand the evolution of a conserved and essential layer of eukaryotic regulation.
Methods
We undertook in silico curation, RNA interactome capture (RIC) and high-resolution mass spectrometry to characterise Giardia’s RBPome. We then explored the function of key RBPs including the earliest known Pumilio homologs (PUF, PUM) and two significant helicases, DDX3X and EIF4A in Giardia, CRISPRi mediated knockdowns, RBP-crosslinking immunoprecipitation (CLiP), quantitative proteomics and in vitro protein expression.
Results
Despite Giardia’s basal evolutionary origins and minimalistic regulatory systems, its RBPome has most novel functions found in higher eukaryotes including the capacity to form membraneless organelles, translationally repressing complex mRNA networks and use this to control major changes in its life-cycle and stress responses.
Conclusions
Giardia provides the earliest eukaryotic record of many of the RBP families acquired by higher eukaryotes and highlights the central role these have played in eukaryotic evolution.