From cradle to grave: The influence of mRNA metabolism on plant stress responses

Abstract

The sessile nature of plants necessitates complex molecular responses to external stimuli, including environmental change. As the intermediary between DNA and protein, mRNA is a crucial cog in plant gene expression, and consequently a key regulatory target. During stress, control of mRNA metabolism (and downstream protein production) takes on particular importance due to the energy constraints imposed on the cell. However, the fate of stress-induced mRNAs, particularly with regard to RNA degradation, remains underexplored. This thesis aimed to address key questions concerning mRNA metabolism within two contexts: activation of transcriptional changes under drought, and the resetting of the transcriptome during recovery from high light stress. Previously, elevated drought tolerance has been observed in Arabidopsis mutants with inhibited activity of the major RNA decay factor XRN. Here, through analysis of RNA sequencing and RNA polymerase II chromatin immunoprecipitation (ChIP), evidence of transcriptional read-through was identified as a consequence of defective XRN-mediated termination. Furthermore, this read-through promoted expression of downstream genes, including stress-responsive loci, providing a potentially novel mechanism by which altered RNA degradation may alter transcriptional dynamics. In comparison to stress-induced changes in gene expression, the recovery phase following cessation of stress has been comparatively under-explored. Previous findings identified rapid downregulation of gene during recovery from high light; however, the mechanism by which this occurs has not been identified. To investigate the role of RNA decay during recovery, transcriptional inhibition experiments were carried out to measure mRNA half-lives. Surprisingly, no change in RNA stability was observed during the shift to recovery. Instead, it appeared that RNA stability was dynamically regulated during stress, including a decrease in stability as gene expression peaked. A shutoff of transcription during recovery then precipitated the rapid decrease in expression observed. Having identified that changes in RNA stability were present during high light stress, the decay pathway responsible was then explored. Previous research had suggested 5' decay was, unexpectedly, not involved in this process; therefore, particular focus was paid here to deadenylation and the 3' decay pathway. However, little evidence could be found substantiating their involvement, nor that of nonsense-mediated decay or post-transcriptional gene silencing. An alternative method was taken to explore the role of translation, which has previously been linked to changes in RNA stability . Characterisation of the high light stress and recovery translatatome was performed, with particular focus paid to genes induced by high light. However, no changes in polysome loading or unloading could be identified that were correlated with changes in RNA stability or gene expression, suggesting that translational changes were not a major contributor to these processes under high light stress. The overall findings of this thesis, particularly with respect to both nuclear and cytosolic RNA decay, emphasise the important role that RNA metabolism has in regulating gene expression in response to environmental stress. The ability to modulate RNA stability to rapidly respond to or recover from such stresses are both a crucial determinant of a plant's survival and productivity, and may also hold implications for further development of crop species that exhibit efficient use of mRNA metabolism in the regulation of gene expression in challenging environments. Show more