PSRP1 Is Not a Ribosomal Protein, but a Ribosome-binding Factor That Is Recycled by the Ribosome-recycling Factor (RRF) and Elongation Factor G (EF-G)

Abstract

Plastid-specific ribosomal proteins (PSRPs) have been proposed to play roles in the light-dependent regulation of chloroplast translation. Here we demonstrate that PSRP1 is not a bona fide ribosomal protein, but rather a functional homologue of the Escherichia coli cold-shock protein pY. Three-dimensional Cryo-electron microscopic (Cryo-EM) reconstructions reveal that, like pY, PSRP1 binds within the intersubunit space of the 70S ribosome, at a site overlapping the positions of mRNA and A-and P-site tRNAs. PSRP1 induces conformational changes within ribosomal components that comprise several intersubunit bridges, including bridge B2a, thereby stabilizes the ribosome against dissociation. We find that the presence of PSRP1/pY lowers the binding of tRNA to the ribosome. Furthermore, similarly to tRNAs, PSRP1/pY is recycled from the ribosome by the concerted action of the ribosome-recycling factor (RRF) and elongation factor G (EF-G). These results suggest a novel function for EF-G and RRF in the post-stress return of PSRP1/pY-inactivated ribosomes to the actively translating pool.Chloroplasts are intracellular organelles present in higher plants and algae; they contain the entire machinery with which the process of photosynthesis is conducted. According to the endosymbiotic theory of chloroplast evolution (1-3), this organelle originated through engulfment of a photosynthetic unicellular prokaryote by a eukaryotic host cell, and the subsequent integration of the two genomes (that of the engulfed prokaryote, and the eukaryotic nucleus) through a process of gene transfers from the chloroplast to the nuclear genome. Thus, although the chloroplast carries its own transcriptional and translational machineries, the development and maintenance of the chloroplast are dependent on the coordinated expression of chloroplast-and nuclear-encoded gene products.The light-dependent process of photosynthesis is the primary function of the chloroplast. Because the components that are crucial for the biogenesis of the photosynthetic apparatus are encoded by both the chloroplast and nuclear genomes, the plant cell has evolved several mechanisms to achieve concerted regulation of gene expression in the two cellular compartments, in response to changes in illumination (4 -6). Regulation of gene expression is primarily post-transcriptional, and is achieved through altered mRNA processing and stability, and the control of the translational apparatus itself in response to environmental signals like light (7-9). It has been demonstrated that the redox state of the chloroplast achieved in response to photosynthetic electron transport can regulate protein synthesis within the chloroplast at the stages of initiation and elongation (10 -12).A detailed analysis of the chloroplast translational machinery, the chloroplast ribosome together with its trans-acting translational factors, will provide important clues as to how such gene regulation is achieved. Proteomic characterization of the chloroplast ribosomes (chlororibosomes) from spinach has r...

Disassembly of theStaphylococcus aureushibernating 100S ribosome by an evolutionarily conserved GTPase

“…These findings suggest that the dissociation of 100S ribosome involves an active mechanism to dislodge the dimerizing factors from the ribosome. Although factors such as initiation factor 3 (IF3) and ribosome recycling factor (RRF)/elongation factor G (EF-G) have been implicated in counteracting the formation of hibernating 100S and PSRP1-mediated 70S ribosomes in vitro (30,33), the proposed activity of these factors on hibernating ribosomes does not corroborate with their canonical roles in ribosome recycling (34,35) (Discussion, and see Fig. S7).…”

“…E. coli also possesses an HPF paralog (referred to as YfiA or pY or RaiA) that silences the 70S ribosome (27)(28)(29) and antagonizes the formation of the 100S complex (21,23). YfiA is absent from Firmicutes and the majority of bacteria but is functionally homologous to PSRP1 in plant chloroplasts (30,31).…”

“…E. coli also possesses an HPF paralog (referred to as YfiA or pY or RaiA) that silences the 70S ribosome (27-29) and antagonizes the formation of the 100S complex (21, 23). YfiA is absent from Firmicutes and the majority of bacteria but is functionally homologous to PSRP1 in plant chloroplasts (30,31).The two 70S monomers in the 100S complex are not irreversibly interlocked; instead, the disassembly of the E. coli 100S ribosome can occur within 1 min after transferring the aged culture into fresh medium (16,32). The 100S ribosomes connected by the long-form HPFs in Firmicutes are more stable by one order of magnitude than the ones generated by the short-form HPF and RMF (2, 14).…”

Expression of Plastid Genes: Organelle-Specific Elaborations on a Prokaryotic Scaffold

“…PSRP1 has proven to be neither a ribosomal protein nor plastid specific (Sharma et al, 2010). In fact, PSRP1 and its bacterial orthologs inhibit translation by blocking tRNA binding sites.…”

The mechanisms of extracellular electron transfer

“…RaiA interferes with protein synthesis by competing for the ribosomal binding sites of tRNAs [195,196]. Hence, RaiA will only be active during times of stress or starvation when levels of tRNA are low [85].…”