Ribosomal Protein Biosynthesis Research Articles

Eukaryotic ribosomal proteins: Interactions with their own pre-mRNAs and their involvement in splicing regulation

The present review summarizes data concerning regulation of eukaryotic ribosomal protein genes expression at the splicing step, including own results. In particular, roles of the ribosomal proteins in regulation of splicing of their coding pre-mRNAs are considered. Special attention is devoted to discussion of the molecular mechanisms that underlie the process and to the analysis of interactions of ribosomal proteins with their own pre-mRNAs and mRNAs. Besides, critical consequences arising by disturbances in the mechanisms of regulation of ribosomal proteins biosynthesis in the cell are noted in the review. Special role of autoregulation in the maintenance of the normal level of ribosomal protein biosynthesis is underlined in the conclusion.

Transcriptome analysis in a rat model of L-DOPA-induced dyskinesia.

We have examined the pattern of striatal messenger RNA expression of over 8000 genes in a rat model of levodopa (L-DOPA)-induced dyskinesia and Parkinson disease (PD). 6-Hydroxydopamine (6-OHDA)-lesioned rats were treated with L-DOPA or physiological saline for 22 days and repeatedly tested for antiakinetic response to L-DOPA and the development of abnormal involuntary movements (AIMs). In a comparison of rats that developed a dyskinetic motor response to rats that did not, we found striking differences in gene expression patterns. In rats that developed dyskinesia, GABA neurons had an increased transcriptional activity, and genes involved in Ca2+ homeostasis, in Ca2+ -dependent signaling, and in structural and synaptic plasticity were upregulated. The gene expression patterns implied that the dyskinetic striatum had increased transcriptional, as well as synaptic activity, and decreased capacity for energy production. Some basic maintenance chores such as ribosome protein biosynthesis were downregulated, possibly a response to expended ATP levels.

Occurrence, functions and biosynthesis of polyamides in microorganisms and biotechnological production.

Microorganisms are able to synthesize three different polyamides by enzymatic processes independently from ribosomal protein biosynthesis: poly(gamma-D-glutamic acid), poly(epsilon-L-lysine) and multi-L-arginyl-poly(L-aspartic acid) which is also referred to as cyanophycin. These polyamides, which occur mainly in Bacillus spp. (and only a few other eubacteria and the nematocysts of Cnidaria, in Streptomyces albulus or in cyanobacteria, respectively), have recently attracted considerable interest of the chemical industry and may be suitable for various applications. This review summarizes our current knowledge on the occurrence, biosynthesis, physiological functions, and biodegradation as well as on the properties and putative applications of these polyamides. Emphasis is placed on the enzymology of the polymerization and on the genes encoding the polymerizing enzymes, which have only recently become available for cyanophycin synthetases. Prospects for novel production processes. in particular for cyanophycin, are also presented.

In vitro production of a TRH-homologous peptide and His-Pro diketopiperazine by human semen.

Fresh human semen, diluted 1:1 v/v with 0.15 M NaCl- 0.05 M phosphate, pH 7.5, undergoes a 3-fold increase in total thyrotropin-releasing hormone (TRH) immunoreactivity on incubation at 4 C for 8 to 16 hours. To identify the mechanism for this increase, pooled human semen was incubated at 4, 37, and 60 C, and the change in composition of the immunoreactive TRH peptides was quantitated by high pressure liquid chromatography and radioimmunoassay of TRH. His-Pro diketopiperazine, a biologically active metabolite of TRH consisting of a cyclic dipeptide of histidine and proline, also was measured by specific RIA. The concentration of TRH (pGlu-His-Pro-NH2) dropped precipitously within the first hour after dilution and incubation at all temperatures studied. A hydrophobic TRH-homologous peptide with the amino acid composition (Glu,X,Y,Pro), where X and Y are neutral, nonaromatic amino acids, increased 8-fold during 16 hours of incubation at 4 C. This TRH-homologous peptide is not derived from TRH because it lacks the histidine is not derived from TRH because it lacks the histidine residue. A 3- to 23-fold increase in His-Pro diketopiperazine levels occurred after 4 hours at 37 C. This was not due primarily to enzymatic removal of the pyroglutamyl residue from TRH by pyroglutamate aminopeptidase, since about 1 hour after ejaculation the initial His-Pro diketopiperazine levels were 9.7 +/- 5.1 micrograms/ml, or approximately 1000-fold greater than the corresponding levels of seminal TRH. Because cycloheximide, which blocks ribosomal protein biosynthesis, did not inhibit in vitro production of the TRH-homologous peptide and the His-Pro cyclic dipeptide, these peptides, like TRH, most likely arise from post-translational cleavage and processing from pre-existing macromolecular precursor proteins.

Stimulation par l'Œstradiol de la synthèse des ribosomes dans les cellules adénohypophysaires

A single dose of 10 micrograms oestradiol injected to a male rat stimulates in the anterior pituitary the synthesis of ribosomal RNA and of the associated proteins. This stimulation is shown using in vitro double-labeling of RNA with adenine or guanine and of proteins with valine. The analysis of polysomes reveals the incorporation of the neo-synthesized molecules into the 40 S and 60 S subunits. Therefore, the stimulation of ribosomal RNA and protein biosynthesis by oestradiol is a coordinated process. No change in the whole polysome distribution is observed in these conditions though such a modification may occur in a specific cell population without being detected by using sucrose gradient analysis.

Effects of low dose actinomycin D treatment in vivo on the biosynthesis of ribosomal proteins in rat liver.

The long-term effects (up to 12 h) of low dose in vivo actinomycin D treatment, which selectively inhibits rRNA synthesis, on the activity of rat liver for the synthesis of ribosomal proteins relative to that for the synthesis of total protein were investigated. The effects of actinomycin D treatment in vivo and in vitro on the template activity of poly(A)-containing mRNA of rat liver for ribosomal proteins were examined by using a wheat germ cell-free system. The following results were obtained. 1. The activity of rat liver for synthesizing total protein observed in vivo and in vitro was inhibited by actinomycin D treatment even at a small dose. 2. A double-labeling technique using [3H] and [14C]leucine in vivo showed that the rate of synthesis of the ribosomal protein fraction relative to that of total protein in actinomycin-treated rat liver (6 + 6 h) was 1.45 times higher than that in the control rat. 3. By using a wheat germ cell-free system, it was shown that the template activity of poly(A)-containing mRNA for the synthesis of total protein was increased slightly by actinomycin D treatment in vivo. Furthermore, the template activity for the ribosomal protein fraction relative to that for total protein was increased. This increase was observed in most of the ribosomal proteins separated on two-dimensional acrylamide gel electrophoresis, although the extents of increase were different among individual ribosomal proteins examined. On the other hand, the selective increase of the template activity for the ribosomal protein fraction was not observed when poly(A)-containing mRNA was incubated with actinomycin D in vitro, although the template activity for total protein was increased slightly.

Ribosomal protein biosynthesis during starvation and refeeding in Tetrahymena pyriformis

The relative abundance of translationally active mRNAs for ribosomal proteins has been measured using Tetrahymena pyriformis subjected to refeeding after a 20-h starvation period. The level of ribosomal proteins synthesized, using total RNA from starved cells and a wheat germ extract for translation, corresponded to less than 3% of the total [ 35S]methionine incorporated into protein. In contrast, total RNA isolated from cells after 1 to 3 h of refeeding yielded a two- to eightfold increase in the amount of the individual ribosomal proteins synthesized, with the maximum level occurring at about 2 h following refeeding. In addition to the increase in the proportion of ribosomal proteins synthesized in vitro, there was also a near linear increase with time in the translational activity of RNA isolated from refed cells. Approximately 25 proteins synthesized in vitro comigrated with authentic ribosomal proteins, as determined by two-dimensional gel electrophoresis. It appears likely that the control of ribosomal protein synthesis in Tetrahymena subjected to starvation and refeeding occurs at the level of transcription.

Biosynthesis of ribosomal proteins by poly(A)-containing mRNAs from rat liver in a wheat germ cell-free system and sizes of mRNAs coding ribosomal proteins

(1) Poly(A)-containing mRNAs from total polysomal RNA of regenerating rat liver were incubated with [ 3H]leucine in a wheat germ cell-free system. Ribosomal proteins were purified as described previously [1], and with two-dimensional gel electrophoresis. The proteins on the gel except for less basic protein had appreciable radioactivity, whereas the surrounding areas had very low radioactivity. Acetic acid-soluble proteins labeled in this system were subjected to three-dimensional gel electrophoresis [2]. Except for L1 and L2 proteins, each of the ribosomal proteins, including less basic ones, showed a major radioactive peak coinciding with the protein band on SDS gel. Thus, the wheat germ cell-free system completely translates almost all mRNAs for individual ribosomal proteins. Equimolar amounts of almost all ribosomal proteins were synthesized in the presence of the saturating concentration of mRNAs. (2) Free polysomes from regenerating rat liver were fractionated into three sizes. Each class of polysomes was incubated with [ 3H]leucine. Ribosomal proteins with molecular weights of 40 000 to 21 000 were mainly synthesized by Fraction B (5–14 monomeric ribosomes), L1 and L2 [2] with 60 000 and 54 000, by Fraction C (>15 monomeric ribosomes) and B, and ribosomal proteins smaller than 20 000 by Fractions A (<pentamer) and B. (3) mRNAs from rat liver total polysomes were fractionated into seven classes by size and each was translated in the wheat germ extract. Ribosomal proteins with molecular weights of 54 000 to 30 000 were mainly synthesized by mRNAs of 12 to 14.5 S, ribosomal proteins of 35 000 to 22 000 by those of 9.5 to 12 S, ribosomal proteins of 22 000 to 13 000 by those of 7 to 9.5 S, and smaller ribosomal proteins by those smaller than 7 S. These results indicate that individual ribosomal proteins are synthesized by monocistronic mRNAs, the lengths of which are proportional to the molecular weights of the corresponding ribosomal proteins.

On the control of ribosomal protein biosynthesis in E. coli

On the control of ribosomal protein biosynthesis in E. coli

On the control of ribosomal protein biosynthesis in E. coli

On the control of ribosomal protein biosynthesis in E. coli

On the control of ribosomal protein biosynthesis in Escherichia coli: II. Studies during recovery from amino acid starvation

The rate of synthesis of ribosomal proteins relative to that of total protein was measured at various times during recovery from arginine starvation in isogenic rel + and rel − strains of Escherichia coli K 12. Total ribosomal proteins are preferentially synthesized early during recovery. Higher rates of synthesis are obtained in the rel + strain than in the rel − strain. Differential rates of synthesis of individual ribosomal proteins are observed at the various times studied. The rate of synthesis of individual proteins increases with time up to maximum values then the rates come down to values similar to those found in exponentially growing cells. The time of restart of synthesis of each protein has been estimated (1) by the time at which the maximum value is reached, and (2) by measuring the rate of synthesis at early time (3 min). Most ribosomal proteins behave similarly in rel − and rel + strains. Proteins have been listed from highly labelled (early proteins) to poorly labelled (late proteins). The significance of the order of restart is considered.

Studies on ribosomal protein biosynthesis in an RNA polymerase temperature sensitive E. coli mutant

In E. coli strain XH56 the synthesis of all RNA species is blocked upon shifting the culture to the non-permissive temperature. The decay of specific messenger RNA species coding for individual ribosomal (r) proteins was followed by measuring the rate of r-protein synthesis by pulse labelling at various times after the shift. The half-lives of the average 30S r-protein and 50S r-protein mRNA species are identical (1.75 min) and shorter than those of the average messenger coding for total cell proteins (2.75 min). Most individual r-protein messengers have a half-life in the same range (1.50-2.00). Only a few r-protein messengers have significantly longer half-lives: S1 (2.80 min), S17 (3.29 min), L29 (2.30 min), L31 (2.30 min), L32 (2.33 min) and L16 (2.60 min). The results indicate that the degradation of most individual r-protein mRNA species is not specifically controlled. After a few min at the non-permissive temperature, all protein synthesis is blocked. The restart of r-protein synthesis was followed after shifting the culture back to the permissive temperature. The recovery of cell growth is very slow. During this period preferential r-protein synthesis was observed. Moreover differential rates of bisynthesis of r-proteins was obtained, it may be indicative of specific regulatory process(es).

Synthesis and soluble pools of ribosomal proteins in Rhodopseudomonas palustris

The specific antibodies prepared against two purified ribosomal proteins (19 and 24) and total 66-S ribosomal proteins have been utilized to measure free ribosomal proteins in Rhodopseudomonas palustris. The free ribosomal protein pool (66 S) amounts to 1–2% of the total soluble proteins of R. palustris. In addition, the size of free ribosomal protein pool (66 S) is calculated to be approx. 7% of the total ribosomal protein on the mature 66-S ribosomes from the pulse-labelling data. A study of the pool size indicates that ribosomal proteins are synthesized at a constant rate during exponential growth. However, during abnormal conditions such as antibiotic treatment, individual ribosomal proteins behave in a manner distinct from the average behavior of total 66-S ribosomal proteins. The co-ordination in the biosynthesis of free ribosomal proteins is no longer apparent during chloramphenicol treatment. Measurements on the free ribosomal protein pools following physiological stress treatment indicate that duing the recovery period, rate of ribosome assembly is greater than the rate of ribosomal protein synthesis.

Preferential biosynthesis of ribosomal structural proteins by free and loosely bound polysomes from regenerating rat liver

Homologous cell-free systems were prepared using free, total bound, tightly bound or KCl-sensitive loosely bound (KCl-sensitive) polysomes from regenerating rat liver. [ 14C] Leucine was incubated with one kind of polysomes and [ 3H] leucine with another kind. The reaction mixtures were then combined, and ribosomal structural proteins were purified as described previously [4], using two-dimensional polyacrylamide gel electrophoresis as the final step [5]. The 3H to 14C ratios of the purified fractions were estimated to compare the activities of the two kinds of polysomes for biosynthesis of ribosomal structural proteins. The following results were obtained: 1. (1) The activity of free polysomes for biosynthesis of ribosomal structural proteins was about 3.6 or 2.4 times higher than that of total bound polysomes in two experiments in which 14C and 3H labeling was reversed. The radioactivities incorporated by free polysomes into most of the proteins separated on two-dimensional gel were found to be definitely higher than those in the surrounding areas, suggesting that most of the ribosomal structural proteins were synthesized by free polysomes. The activity of free polysomes for biosynthesis of ribosomal structural proteins was about 7 times higher than that of tightly bound polysomes, which were prepared by washing the microsomal membrane fraction with 0.5 M KCl. The radioactivities incorporated by tightly bound polysomes into the proteins separated on two-dimensional gel were only slightly higher than those in the surrounding areas, indicating that these polysomes had very low synthetic activity. 2. (2) Preferential synthesis of histones by free polysomes was also shown using the same procedures. 3. (3) KCl-sensitive polysomes which were released by washing the microsomal membrane fraction with 0.5 M KCl, were shown to have definitely higher activity than tightly bound polysomes for biosynthesis of ribosomal structural proteins. 4. (4) From these results, it is concluded that most of the ribosomal structural proteins are preferentially synthesized by free and KCl-sensitive polysomes in regenerating rat liver.