Stress response inEntamoeba histolyticais associated with robust processing of tRNA to tRNA-halves

Abstract

tRNA-derived fragments have been reported in many different organisms and have diverse cellular roles such as regulating gene expression, inhibiting protein translation, silencing transposable elements and modulating cell proliferation. In particular tRNA halves, a class of tRNA fragments produced by the cleavage of tRNAs in the anti-codon loop, have been widely reported to accumulate under stress and regulate translation in cells. Here we report the presence of tRNA-derived fragments in Entamoeba with tRNA halves being the most abundant. We further established that tRNA halves accumulate in the parasites upon different stress stimuli such as oxidative stress, heat shock, and serum starvation. We also observed differential expression of tRNA halves during developmental changes of trophozoite to cyst conversion with various tRNA halves accumulating during early encystation. In contrast to other systems, the stress response does not appear to be mediated by a few specific tRNA halves as multiple tRNAs appear to be processed during the various stresses. Furthermore, we identified some tRNA-derived fragments are associated with Entamoeba Argonaute proteins, EhAgo2-2, and EhAgo2-3, which have a preference for different tRNA-derived fragment species. Finally, we show that tRNA halves are packaged inside extracellular vesicles secreted by amoeba. The ubiquitous presence of tRNA-derived fragments, their association with the Argonaute proteins, and the accumulation of tRNA halves during multiple different stresses including encystation suggest a nuanced level of gene expression regulation mediated by different tRNA-derived fragments in Entamoeba.

Transfer RNA-Derived Small RNAs in the Pathogenesis of Parasitic Protozoa

“…It has also been shown that amoebic EVs contain tRNA halves and tRFs which associate with E. histolytica AGO proteins Eh AGO2-2 and Eh AGO2-3 [ 48 ]. Prior to EV collection, parasites were cultivated in serum-free medium to prevent contamination with bovine serum EVs.…”

“…Thus, serum starvation is a trigger that prompted increased production of tRNA halves which is common in other organisms where tRNA cleavage into halves is induced during nutritional and oxidative stress [ 17 , 70 , 71 , 72 , 73 ]. Most of the tRNA halves originated from a few tRNAs, particularly tRNA Ala (AGC), tRNA Ala (UGC), tRNA Arg (UCU), and tRNA Asp (GUC) [ 48 ]. The biogenesis of tsRNA can be highly affected by tRNA modifications [ 16 ].…”

“…It has also been reported that queuine can induce a C38 hypermethylation in the tRNA Asp (GUC) anticodon loop to promote the resistance of the parasite against oxidative stress [ 74 ], thus suggesting that a modulating mechanism may exist to regulate the synthesis of tRNA halves in this organism. However, the clear role of tRNA halves in the EVs of E. histolytica remains unknown, although it is hypothesized that they mediate intercellular communication [ 48 ].…”