PRJNA212003
Trypanosoma brucei rhodesiense siRNA sequencing
Parasitic protozoa of the flagellate order Kinetoplastida represent one of the deepest branches of the eukaryotic lineage and are therefore considered descendants of “ancient” eukaryotes. Among this group of organisms the mechanism of RNA interference (RNAi) has been investigated in Trypanosoma brucei and to a lesser degree in Leishmania (Viannia) spp. The pathway is triggered by long double-stranded (ds) RNA and in T. brucei requires a set of five core genes, including a single Argonaute protein, TbAGO1. The five genes are conserved in Leishmania (Viannia) spp, but are absent in other major kinetoplastid species, such as T. cruzi and L. major. In T. brucei small interfering RNAs (siRNAs) are methylated at the 3’ end, whereas Leishmania (Viannia) spp siRNAs are not. Here we report that T. brucei HEN1, an ortholog of the metazoan HEN1 2’-O-methyltransferases, is required for methylation of siRNAs. Loss of TbHEN1 causes a reduction in the length of siRNAs. The shorter siRNAs in hen1-/- parasites are single-stranded, associated with TbAGO1 and a subset carries a non-templated uridine at the 3’ end. These findings support a model wherein TbHEN1 methylates siRNA 3’ ends after they are loaded into TbAGO1 and this methylation protects siRNAs from uridylation and 3' trimming.