UBC Theses and Dissertations

UBC Theses Logo

UBC Theses and Dissertations

Investigating plant-nepovirus interactions : non-canonical translation of tomato ringspot virus RNAs and plant antiviral defence responses associated with symptom recovery Paudel, Dinesh Babu

Abstract

Tomato ringspot virus (ToRSV, genus Nepovirus) was used as a model to study two essential steps of the infection cycle: the translation of uncapped viral RNAs and the induction of host antiviral defences. Using reporter transcripts containing the 5’ and 3’ untranslated regions (UTRs) of ToRSV RNAs and a portion of the viral coding region, I show that translation initiates at the first AUG of the viral RNA (nt 77) in spite of its suboptimal context. A predicted downstream stem-loop structure (5’ SL) may enhance initiation at this AUG. I show that both viral UTRs are required for efficient translation. A region of 386 nt in the viral 3’ UTR was necessary and sufficient for translation in conjunction with the viral 5’ UTR. In this region, a predicted stem-loop (SL 3’) may be involved in a long-distance RNA-RNA interaction with the 5’ SL as suggested by the complementarity of the loop sequences. The viral 3’ UTR inhibited the cap-dependent translation of control transcripts, suggesting that it contains a cis-acting translation element that binds one or several translation factors. These results highlight a unique non-canonical translation initiation mechanism that shares similarities with that described for Blackcurrant reversion virus (another nepovirus), although the cis-acting elements required for translation differed between the two viruses. Symptom recovery (an asymptomatic phase of infection that follows an initial symptomatic phase) has been attributed to antiviral RNA silencing. Using Nicotiana benthamiana plants infected with two ToRSV isolates (Rasp1 and GYV), I show that symptom recovery is dependent on environmental conditions and the isolate. The recovery from GYV- or Rasp1-infection shared similar hallmarks of RNA silencing but GYV-infected plants recovered at a wider range of temperatures (21–27°C). ARGONAUTE (AGO) proteins are key components of the RNA silencing pathway. Silencing of AGO1, which prevents the recovery of Rasp1-infected plants, did not prevent recovery from GYV infection. I show differential accumulation of AGO2 depending on the temperature and the isolate. However, although silencing or mutation of AGO2 increased virus accumulation in infected plants, it did not prevent symptom recovery. Taken together, these results highlight the complexity of plant-virus interactions.

Item Citations and Data

Rights

Attribution-NonCommercial-NoDerivatives 4.0 International