Ribosomal protein mutants and their effects on plant growth and development

Abstract

Ribosomes, large enzymatic complexes containing an RNA catalytic core, drive protein synthesis in all living organisms. 80S cytoplasmic eukaryotic ribosomes are comprised of four rRNAs and approximately 80 ribosomal proteins (r-proteins). R-proteins are encoded by gene families with large families (average of twelve members) predominating in mammals and smaller families (two to seven members) in plants. Increased ribosome heterogeneity is possible in plant ribosomes due to multiple transcriptionally active members in each family, whereas, in mammalian r-protein gene families, only one member is typically active. Multiple functional paralogs provide for greater plasticity in response to environmental/developmental cues, as well as, increasing the possibility of individual paralogs procuring or retaining extraribosomal functions. This research investigated the effects of r-protein mutations on plant growth and development. Through RNA interference (RNAi) mediated knockdown (KD) of type I (cytoplasmic: RPS15aA/D and F) and type II (non-cytosolic: RPS15aB and E) RPS15a family members I was able to confirm the delineation between the two types. Subcellular localization of the type I isoforms was nuclear/nucleolar while localization of type II isoforms was non-mitochondrial and probably cytosolic. Illumina sequencing of two r-protein mutant transcriptomes, pfl1 (rps18a) and pfl2 (rps13a), identified a novel set of up and down regulated genes, previously unknown or linked to r-protein mutants. The 20 genes identified were classified into four groups (1) plant defense, (2) transposable elements, (3) nitrogen metabolism and (4) genes with unknown function. Illumina miRNOME analysis revealed no changes in the miRNA profile of pfl1 and pfl2 plants. These data do not support the previously proposed theory that a disruption in ribosome biogenesis (by decreased r-protein synthesis) disrupts miRNA-mediated degradation of a range of auxin response genes. Finally, a novel double r-protein mutant, rps18a:HF/RPL18B, presented a late flowering/thickened bolt phenotype not seen in a rps13a:HF/RPL18B mutant, suggesting that RPS18A has an extraribosomal role in plant growth and development in Arabidopsis.