Cell growth and homeostasis are disrupted in arabidopsis rns2-2 mutants missing the main vacuolar RNase activity.

Background And Aims: Enzymes belonging to the RNase T2 family are essential for normal rRNA turnover in eukaryotes. In Arabidopsis thaliana, this function is performed by RNS2. The null mutant rns2-2 has increased rRNA half-life and constitutive autophagy. The aim of this work was to determine the molecular changes that take place in the rns2-2 mutant that may lead to altered cellular homeostasis, manifested by the observed cellular phenotype.

Methods: To determine the effect of defective rRNA turnover on cellular homeostasis, comparative transcriptome and metabolome analyses of 10-day-old wild-type and rns2-2 seedlings were used to identify molecular processes affected in the mutant. Bioinformatics analyses suggested additional phenotypes that were confirmed through direct plant size measurements and microscopy.

Key Results: Few genes were differentially expressed in the rns2-2 mutant, indicating that control of autophagy in this genotype is mainly achieved at the post-transcriptional level. Among differentially expressed genes, transcripts related to carbon flux processes, particularly the pentose phosphate pathway (PPP), were identified. Metabolite analyses confirmed changes in the levels of PPP intermediates. Genes related to cell wall loosening were also differentially expressed in the mutant, and a decrease in monosaccharide components of cell wall hemicellulose were found. As a potential effect of weaker cell walls, rns2-2 plants are larger than wild-type controls, due to larger cells and increased water content. Elevated levels of reactive oxygen species (ROS) were also measured in rns2-2, and the constitutive autophagy phenotype was blocked by preventing ROS production via NADPH oxidase.

Conclusions: Lack of rRNA recycling in rns2-2 cells triggers a change in carbon flux, which is redirected through the PPP to produce ribose-5-phosphate for de novo nucleoside synthesis. rRNA or ribosome turnover is thus essential for cellular homeostasis, probably through maintenance of nucleoside levels as part of the salvage pathway.