Proteomic identification of all plastid‐specific ribosomal proteins in higher plant chloroplast 30S ribosomal subunit

Abstract

Six ribosomal proteins are specific to higher plant chloroplast ribosomes [Subramanian, A.R. (1993) Trends Biochem. Sci.18, 177–180]. Three of them have been fully characterized [Yamaguchi, K., von Knoblauch, K. & Subramanian, A. R. (2000) J. Biol. Chem. 275, 28455–28465; Yamaguchi, K. & Subramanian, A. R. (2000) J. Biol. Chem. 275, 28466–28482]. The remaining three plastid‐specific ribosomal proteins (PSRPs), all on the small subunit, have now been characterized (2D PAGE, HPLC, N‐terminal/internal peptide sequencing, electrospray ionization MS, cloning/ sequencing of precursor cDNAs). PSRP‐3 exists in two forms (α/β, N‐terminus free and blocked by post‐translational modification), whereas PSRP‐2 and PSRP‐4 appear, from MS data, to be unmodified. PSRP‐2 contains two RNA‐binding domains which occur in mRNA processing/stabilizing proteins (e.g. U1A snRNP, poly(A)‐binding proteins), suggesting a possible role for it in the recruiting of stored chloroplast mRNAs for active protein synthesis. PSRP‐3 is the higher plant orthologue of a hypothetical protein (ycf65 gene product), first reported in the chloroplast genome of a red alga. The ycf65 gene is absent from the chloroplast genomes of higher plants. Therefore, we suggest that Psrp‐3/ycf65, encoding an evolutionarily conserved chloroplast ribosomal protein, represents an example of organelle‐to‐nucleus gene transfer in chloroplast evolution. PSRP‐4 shows strong homology with Thx, a small basic ribosomal protein of Thermus thermophilus 30S subunit (with a specific structural role in the subunit crystallographic structure), but its orthologues are absent from Escherichia coli and the photosynthetic bacterium Synechocystis. We would therefore suggest that PSRP‐4 is an example of gene capture (via horizontal gene transfer) during chloro‐ribosome emergence. Orthologues of all six PSRPs are identifiable in the complete genome sequence of Arabidopsis thaliana and in the higher plant expressed sequence tag database. All six PSRPs are nucleus‐encoded. The cytosolic precursors of PSRP‐2, PSRP‐3, and PSRP‐4 have average targeting peptides (62, 58, and 54 residues long), and the mature proteins are of 196, 121, and 47 residues length (molar masses, 21.7, 13.8 and 5.2 kDa), respectively. Functions of the PSRPs as active participants in translational regulation, the key feature of chloroplast protein synthesis, are discussed and a model is proposed.

Protein Profiling of Plastoglobules in Chloroplasts and Chromoplasts. A Surprising Site for Differential Accumulation of Metabolic Enzymes

“…It likely partners with RAP41 (At3g63140; Peltier et al, 2006), also identified here in the PGs. C. reinhardtii homologs of RAP41 (At3g63140) and RAP38 (At1g09340) were copurified with 70S chloroplast ribosomes (Yamaguchi and Subramanian, 2003), and curiously a homolog in spinach (Spinacia oleracea) was reported to be an RNA nuclease (Yang et al, 1996;Bollenbach and Stern, 2003).…”

Proteomic study of the Bacillus subtilis ribosome: Finding of zinc-dependent replacement for ribosomal protein L31 paralogues

“…The proteome of chloro-ribosomes of a higher plant (spinach) detected 58 distinct proteins: 52 of 54 Escherichia coli r-protein orthologues (except L25 and L30) and six plastid-specific ribosomal proteins (PSRPs) Subramanian, 2000, 2003;. It is likely that PSRPs are involved in specific functions in the chloroplast, such as light-activated translation initiation (Yamaguchi and Subramanian, 2003). On the other hand, the mitochondrial ribosome (mitoribosome) is less similar to that of bacteria than the chloro-ribosome.…”

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)

“…Proteomic characterization of the chloroplast ribosomes (chlororibosomes) from spinach has revealed the presence of six plastid-specific ribosomal proteins (PSRPs), 5 four of which are associated with the 30S subunit and two with the 50S subunit, in addition to the plastid orthologs of bacterial r-proteins (13)(14)(15)(16). All six PSRPs are encoded by genes located in the nucleus and are synthesized in the cytoplasm as precursor polypeptides.…”