Synonymous mutations in the phosphoglycerate kinase 1 gene induce an altered response to protein misfolding in .

Background: Proteostasis refers to the processes that regulate the biogenesis, folding, trafficking, and degradation of proteins. Any alteration in these processes can lead to cell malfunction. Protein synthesis, a key proteostatic process, is highly-regulated at multiple levels to ensure adequate adaptation to environmental and physiological challenges such as different stressors, proteotoxic conditions and aging, among other factors.

Oxidative stress strongly restricts the effect of codon choice on the efficiency of protein synthesis in .

Introduction: The response of enterobacteria to oxidative stress is usually considered to be regulated by transcription factors such as OxyR and SoxR. Nevertheless, several reports have shown that under oxidative stress the levels, modification and aminoacylation of tRNAs may be altered suggesting a role of codon bias in regulation of gene expression under this condition.

Methods: In order to characterize the effects of oxidative stress on translation elongation we constructed a library of 61 plasmids, each coding for the green fluorescent protein (GFP) translationally fused to a different set of four identical codons.

Regulation of Leaderless mRNA Translation in Bacteria.

In bacteria, the translation of genetic information can begin through at least three different mechanisms: canonical or Shine-Dalgarno-led initiation, readthrough or 70S scanning initiation, or leaderless initiation. Here, we discuss the main features and regulation of the last, which is characterized mainly by the ability of 70S ribosomal particles to bind to AUG located at or near the 5' end of mRNAs to initiate translation. These leaderless mRNAs (lmRNAs) are rare in enterobacteria, such as , but are common in other bacteria, such as and where they may represent more than 20% and even up to 60% of the genes.

Genomic analysis of the diversity, antimicrobial resistance and virulence potential of clinical Campylobacter jejuni and Campylobacter coli strains from Chile.

Campylobacter jejuni and Campylobacter coli are the leading cause of human gastroenteritis in the industrialized world and an emerging threat in developing countries. The incidence of campylobacteriosis in South America is greatly underestimated, mostly due to the lack of adequate diagnostic methods. Accordingly, there is limited genomic and epidemiological data from this region.

Modification of Transfer RNA Levels Affects Cyclin Aggregation and the Correct Duplication of Yeast Cells.

Codon usage bias (the preferential use of certain synonymous codons (optimal) over others is found at the organism level (intergenomic) within specific genomes (intragenomic) and even in certain genes. Whether it is the result of genetic drift due to GC/AT content and/or natural selection is a topic of intense debate. Preferential codons are mostly found in genes encoding highly-expressed proteins, while lowly-expressed proteins usually contain a high proportion of rare (lowly-represented) codons.

Modulation of Translation by the Specific Inactivation of tRNA Under Oxidative Stress.

Bacterial oxidative stress responses are generally controlled by transcription factors that modulate the synthesis of RNAs with the aid of some sRNAs that control the stability, and in some cases the translation, of specific mRNAs. Here, we report that oxidative stress additionally leads to inactivation of tRNA in , inducing a series of physiological changes. The observed inactivation of tRNA correlated with altered efficiency of translation of Gly codons, suggesting a possible mechanism of translational control of gene expression under oxidative stress.

Codon usage revisited: Lack of correlation between codon usage and the number of tRNA genes in enterobacteria.

It is widely believed that if a high number of genes are found for any tRNA in a rapidly replicating bacteria, then the cytoplasmic levels of that tRNA will be high and an open reading frame containing a higher frequency of the complementary codon will be translated faster. This idea is based on correlations between the number of tRNA genes, tRNA concentration and the frequency of codon usage observed in a limited number of strains as well as from the fact that artificially changing the number of tRNA genes alters translation efficiency and consequently the amount of properly folded protein synthesized. tRNA gene number may greatly vary in a genome due to duplications, deletions and lateral transfer which in turn would alter the levels and functionality of many proteins.

Non-canonical roles of tRNAs and tRNA mimics in bacterial cell biology.

Transfer RNAs (tRNAs) are the macromolecules that transfer activated amino acids from aminoacyl-tRNA synthetases to the ribosome, where they are used for the mRNA guided synthesis of proteins. Transfer RNAs are ancient molecules, perhaps even predating the existence of the translation machinery. Albeit old, these molecules are tremendously conserved, a characteristic that is well illustrated by the fact that some bacterial tRNAs are efficient and specific substrates of eukaryotic aminoacyl-tRNA synthetases and ribosomes.

The non-canonical hydroxylase structure of YfcM reveals a metal ion-coordination motif required for EF-P hydroxylation.

EF-P is a bacterial tRNA-mimic protein, which accelerates the ribosome-catalyzed polymerization of poly-prolines. In Escherichia coli, EF-P is post-translationally modified on a conserved lysine residue. The post-translational modification is performed in a two-step reaction involving the addition of a β-lysine moiety and the subsequent hydroxylation, catalyzed by PoxA and YfcM, respectively.

Translation initiation rate determines the impact of ribosome stalling on bacterial protein synthesis.

Ribosome stalling during translation can be caused by a number of characterized mechanisms. However, the impact of elongation stalls on protein levels is variable, and the reasons for this are often unclear. To investigate this relationship, we examined the bacterial translation elongation factor P (EF-P), which plays a critical role in rescuing ribosomes stalled at specific amino acid sequences including polyproline motifs.

EF-P dependent pauses integrate proximal and distal signals during translation.

Elongation factor P (EF-P) is required for the efficient synthesis of proteins with stretches of consecutive prolines and other motifs that would otherwise lead to ribosome pausing. However, previous reports also demonstrated that levels of most diprolyl-containing proteins are not altered by the deletion of efp. To define the particular sequences that trigger ribosome stalling at diprolyl (PPX) motifs, we used ribosome profiling to monitor global ribosome occupancy in Escherichia coli strains lacking EF-P.

Molecular evolution of protein-RNA mimicry as a mechanism for translational control.

Elongation factor P (EF-P) is a conserved ribosome-binding protein that structurally mimics tRNA to enable the synthesis of peptides containing motifs that otherwise would induce translational stalling, including polyproline. In many bacteria, EF-P function requires post-translational modification with (R)-β-lysine by the lysyl-tRNA synthetase paralog PoxA. To investigate how recognition of EF-P by PoxA evolved from tRNA recognition by aminoacyl-tRNA synthetases, we compared the roles of EF-P/PoxA polar contacts with analogous interactions in a closely related tRNA/synthetase complex.

ICE Afe 1, an actively excising genetic element from the biomining bacterium Acidithiobacillus ferrooxidans.

Integrative conjugative elements (ICEs) are self-transferred mobile genetic elements that contribute to horizontal gene transfer. An ICE (ICEAfe1) was identified in the genome of Acidithiobacillus ferrooxidans ATCC 23270. Excision of the element and expression of relevant genes under normal and DNA-damaging growth conditions was analyzed.

A pseudo-tRNA modulates antibiotic resistance in Bacillus cereus.

Bacterial genomic islands are often flanked by tRNA genes, which act as sites for the integration of foreign DNA into the host chromosome. For example, Bacillus cereus ATCC14579 contains a pathogenicity island flanked by a predicted pseudo-tRNA, tRNA(Other), which does not function in translation. Deletion of tRNA(Other) led to significant changes in cell wall morphology and antibiotic resistance and was accompanied by changes in the expression of numerous genes involved in oxidative stress responses, several of which contain significant complementarities to sequences surrounding tRNA(Other).

β-Lysine discrimination by lysyl-tRNA synthetase.

Elongation factor P is modified with (R)-β-lysine by the lysyl-tRNA synthetase (LysRS) paralog PoxA. PoxA specificity is orthogonal to LysRS, despite their high similarity. To investigate α- and β-lysine recognition by LysRS and PoxA, amino acid replacements were made in the LysRS active site guided by the PoxA structure.

Cellular levels of heme affect the activity of dimeric glutamyl-tRNA reductase.

Glutamyl-tRNA reductase (GluTR) is the first enzyme committed to tetrapyrrole biosynthesis by the C(5)-pathway. This enzyme transforms glutamyl-tRNA into glutamate-1-semi-aldehyde, which is then transformed into 5-amino levulinic acid by the glutamate-1-semi-aldehyde 2,1-aminomutase. Binding of heme to GluTR seems to be relevant to regulate the enzyme function.

Redox status affects the catalytic activity of glutamyl-tRNA synthetase.

Glutamyl-tRNA synthetases (GluRS) provide Glu-tRNA for different processes including protein synthesis, glutamine transamidation and tetrapyrrole biosynthesis. Many organisms contain multiple GluRSs, but whether these duplications solely broaden tRNA specificity or also play additional roles in tetrapyrrole biosynthesis is not known. Previous studies have shown that GluRS1, one of two GluRSs from the extremophile Acidithiobacillus ferrooxidans, is inactivated when intracellular heme is elevated suggesting a specific role for GluRS1 in the regulation of tetrapyrrole biosynthesis.

Regulation of a glutamyl-tRNA synthetase by the heme status.

Glutamyl-tRNA (Glu-tRNA), formed by Glu-tRNA synthetase (GluRS), is a substrate for protein biosynthesis and tetrapyrrole formation by the C(5) pathway. In this route Glu-tRNA is transformed to delta-aminolevulinic acid, the universal precursor of tetrapyrroles (e.g.

A tRNA(Glu) that uncouples protein and tetrapyrrole biosynthesis.

Glu-tRNA is either bound to elongation factor Tu to enter protein synthesis or is reduced by glutamyl-tRNA reductase (GluTR) in the first step of tetrapyrrole biosynthesis in most bacteria, archaea and in chloroplasts. Acidithiobacillus ferrooxidans, a bacterium that synthesizes a vast amount of heme, contains three genes encoding tRNA(Glu). All tRNA(Glu) species are substrates in vitro of GluRS1 from A.