The Trypanosoma brucei RNA-binding protein DRBD18 ensures correct mRNA trans splicing and polyadenylation patterns

Abstract

The parasite Trypanosoma brucei grows as bloodstream forms in mammals, and as procyclic forms in tsetse flies. Transcription is polycistronic, all mRNAs are trans spliced, and polyadenylation sites are defined by downstream splicing signals. Expression regulation therefore depends heavily on post-transcriptional mechanisms. The RNA-binding protein DRBD18 was previously implicated in the export of some mRNAs from the nucleus in procyclic forms. It copurifies the outer ring of the nuclear pore, mRNA export factors and exon-junction-complex proteins. We show that for >200 mRNAs, DRBD18 depletion caused preferential accumulation of versions with shortened 3'-untranslated regions, arising from use of polyadenylation sites that were either undetectable or rarely seen in non-depleted cells. The shortened mRNAs were often, but not always, more abundant in depleted cells than the corresponding longer versions in normal cells. Their appearance was linked to the appearance of trans spliced, polyadenylated RNAs containing only downstream 3'-untranslated-region-derived sequences. Experiments with one mRNA suggested that nuclear retention alone, through depletion of MEX67, did not affect mRNA length, suggesting a specific effect of DRBD18 on processing. Since DRBD18-bound mRNAs were enriched in polypyrimidine tract motifs, and it is found in both the nucleus and the cytoplasm, we suggest that DRBD18 acts in the nucleus by binding to polypyrimidine tracts in 3'-UTRs. DRBD18 binding might both prevent polypyrimidine tract recognition by splicing factors, and promote export of the bound RNAs to the cytosol.