Phosphorylation of the Saccharomyces cerevisiae equivalent of ribosomal protein S6 has no detectable effect on growth.

Abstract

The phosphorylation of mammalian ribosomal protein S6 is affected by a variety of agents, including growth factors and tumor promoters, as well as by expressed oncogenes. Its potential role in the regulation of protein synthesis has been the object of much study. We have developed strains of Saccharomyces cerevisiae in which the phosphorylatable serines of the equivalent ribosomal protein (S10) were converted to alanines by site-directed mutagenesis. The S10 of such cells is not phosphorylated. Comparison of these cells with the parental cells, whose genomes differ by only six nucleotides, revealed no differences in the lag phase or logarithmic phase of a growth cycle, in growth on different carbon sources, in sporulation, or in sensitivity to heat shock. We conclude that in S. cerevisiae the phosphorylation of ribosomal protein S10 may play no role in regulating the synthesis of proteins. This conclusion leads one to ask whether certain protein phosphorylations are simply the adventitious, if easily observable, result of the imperfect specificity of one or another protein kinase.Ribosomal protein S6 is the major phosphoprotein of eucaryotic ribosomes. It can be phosphorylated at multiple sites (two in Saccharomyces cerevisiae and at least five in vertebrates), and the degree of phosphorylation is somehow related to cell proliferation. A variety of stimuli and agents have been observed to affect the phosphorylation of S6, including insulin (35,40,44), growth factors (40), serum treatment (40), viral transformation (7), phorbol esters (3), and fertilization (2, 26). Phosphorylation of the S6 protein is carried out by a number of kinases, one of which is apparently specific for S6, and is activated by at least two regulatory mechanisms (4).In S. cerevisiae the homologous ribosomal protein, referred to as S10, is phosphorylated after transfer of a stationary culture to fresh nutrient medium (16). Phosphorylation also occurs at an early stage of germination (37). As in other eucaryotes, the protein is dephosphorylated during heat shock (16).At present, much is known about the phenomenology of S6 phosphorylation, but little about its function. Generally, as phosphorylation of protein S6 increases, the level of protein synthesis increases (14, 40), and as dephosphorylation of S6 occurs, the level of protein synthesis decreases (13, 34). However, phosphorylation of S6 has been observed to be stimulated under conditions in which protein synthesis increased, decreased, or remained unchanged (14,17,41). In serum-stimulated 3T3 cells, ribosomes containing highly phosphorylated S6 are preferentially incorporated into new polysomes, suggesting a role in mRNA recruitment (8,40). Such a role is supported by the localization of S6 in the mRNA-binding site of 80S ribosomes (39). In vitro attempts to demonstrate an effect of S6 phosphorylation have been successful in some cases (6) and unsuccessful in others (20,25).Because of the potential importance of the phosphorylation of S6 to the regulation of cell growth and because of th...

Regulation of Ribosome Biogenesis by the Rapamycin-sensitive TOR-signaling Pathway inSaccharomyces cerevisiae

“…These differences are consistent with the fact that in yeast r-protein mRNAs lack 5ЈTOP sequences and phosphorylation of ribosomal protein S6 is not essential for normal cell growth (Johnson and Warner, 1987), features that are required for the observed translational regulation in mammalian cells. On the other hand, it has not been reported whether rapamycin affects r-protein gene transcription in mammalian cells.…”

“…Moreover, although yeast ribosomal protein S6 is normally phosphorylated in a manner that parallels the growth rate of the cell, no obvious growth defect results when the phosphorylation sites are mutated (Johnson and Warner, 1987). Thus the question arises as to what role TOR signaling plays in ribosome synthesis in yeast.…”

Fission yeast TORC1 regulates phosphorylation of ribosomal S6 proteins in response to nutrients and its activity is inhibited by rapamycin

“…We also indicated that phosphorylation of both the serine residues is dispensable for cell viability in fission yeast. Similarly, in budding yeast, phosphorylation of the conserved serine residues is not necessary for cell survival and protein synthesis (Johnson and Warner, 1987). In the case of mouse, phosphorylation of S6 is also dispensable for cell viability, whereas phosphorylation of the ribosomal protein is involved in cell size and glucose homeostasis (Ruvinsky et al, 2005).…”

Ribosomal Protein rpS2 Is Hypomethylated in PRMT3-deficient Mice

“…To some extent, this enzyme/substrate pair of PRMT3/rpS2 is evocative of the S6 kinase/rpS6 pairing. Both enzyme substrate pairs have been conserved over an extended evolutionary period and are found from yeast to man (4,5,21,22). The phosphorylation of rpS6 occurs in response to numerous growth stimulatory agents and temporally correlates with the initiation of protein synthesis (23).…”