A phosphorylation-deficient ribosomal protein eS6 is largely functional inArabidopsis thaliana, rescuing mutant defects from global translation and gene expression to photosynthesis and growth

Abstract

ABSTRACTThe eukaryote-specific ribosomal protein of the small subunit eS6 is phosphorylated through the Target of rapamycin (TOR) kinase pathway. Although this phosphorylation event responds dynamically to environmental conditions and has been studied for over 50 years, its biochemical and physiological significance remains controversial and poorly understood. Here we report data fromArabidopsis thaliana, which indicate that plants expressing only a largely phospho-deficient isoform of eS6 grow essentially normally under laboratory conditions. The eS6A (RPS6A) paralog of eS6 functionally rescued double mutations in bothrps6aandrps6bgenes when expressed at approximately twice the wild-type dosage. A mutant isoform of eS6A lacking the major six phosphorylatable serine and threonine residues in its carboxyl-terminal tail also rescued the lethality, rosette growth, and polyribosome loading of the double mutant. It also complemented many mutant phenotypes ofrps6that were newly characterized here, including photosynthetic efficiency, and the vast majority of gene expression defects that were measured by transcriptomics and proteomics. However, compared to plants rescued with a phospho-enabled version of eS6A, the phospho-deficient seedlings retained a mild pointed-leaf phenotype, root growth was reduced, and certain cell cycle related mRNAs and ribosome biogenesis proteins were misexpressed. The residual defects of the phospho-deficient seedlings could be understood as an incomplete rescue of therps6mutant defects, with little or no evidence for gain-of-function defects. As expected, the phospho-deficient eS6A also rescued therps6aandrps6bsingle mutants; however, phosphorylation of the eS6B paralog remained lower than predicted, further underscoring that plants can tolerate phospho-deficiency of eS6 well. Our data also yield new insights into how plants cope with mutations in essential, duplicated ribosomal protein isoforms.