The anti-Shine-Dalgarno sequence drives translational pausing and codon choice in bacteria.

Department of Cellular and Molecular Pharmacology, Howard Hughes Medical Institute, University of California, San Francisco, California 94158, USA.
Nature (Impact Factor: 42.35). 03/2012; 484(7395):538-41.
Source: PubMed

ABSTRACT Protein synthesis by ribosomes takes place on a linear substrate but at non-uniform speeds. Transient pausing of ribosomes can affect a variety of co-translational processes, including protein targeting and folding. These pauses are influenced by the sequence of the messenger RNA. Thus, redundancy in the genetic code allows the same protein to be translated at different rates. However, our knowledge of both the position and the mechanism of translational pausing in vivo is highly limited. Here we present a genome-wide analysis of translational pausing in bacteria by ribosome profiling--deep sequencing of ribosome-protected mRNA fragments. This approach enables the high-resolution measurement of ribosome density profiles along most transcripts at unperturbed, endogenous expression levels. Unexpectedly, we found that codons decoded by rare transfer RNAs do not lead to slow translation under nutrient-rich conditions. Instead, Shine-Dalgarno-(SD)-like features within coding sequences cause pervasive translational pausing. Using an orthogonal ribosome possessing an altered anti-SD sequence, we show that pausing is due to hybridization between the mRNA and 16S ribosomal RNA of the translating ribosome. In protein-coding sequences, internal SD sequences are disfavoured, which leads to biased usage, avoiding codons and codon pairs that resemble canonical SD sites. Our results indicate that internal SD-like sequences are a major determinant of translation rates and a global driving force for the coding of bacterial genomes.

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    ABSTRACT: The codon composition of the coding sequence's (ORF) 5' end first few dozen codons is known to be distinct to that of the rest of the ORF. Various explanations for the unusual codon distribution in this region have been proposed in recent years, and include, among others, novel regulatory mechanisms of translation initiation and elongation. However, due to the fact that many overlapping regulatory signals are suggested to be associated with this relatively short region, its research is challenging. Here, we review the currently known signals that appear in this region, the theories related to the way they regulate translation and affect the organismal fitness, and the debates they provoke. © The Author(s) 2014. Published by Oxford University Press on behalf of Nucleic Acids Research.
    Nucleic Acids Research 12/2014; · 8.81 Impact Factor
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    ABSTRACT: Ribosome profiling data report on the distribution of translating ribosomes, at steady-state, with codon-level resolution. We present a robust method to extract codon translation rates and protein synthesis rates from these data, and identify causal features associated with elongation and translation efficiency in physiological conditions in yeast. We show that neither elongation rate nor translational efficiency is improved by experimental manipulation of the abundance or body sequence of the rare AGG tRNA. Deletion of three of the four copies of the heavily used ACA tRNA shows a modest efficiency decrease that could be explained by other rate-reducing signals at gene start. This suggests that correlation between codon bias and efficiency arises as selection for codons to utilize translation machinery efficiently in highly translated genes. We also show a correlation between efficiency and RNA structure calculated both computationally and from recent structure probing data, as well as the Kozak initiation motif, which may comprise a mechanism to regulate initiation.
    Molecular Systems Biology 12/2014; 10(12). · 14.10 Impact Factor
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    ABSTRACT: Background Codon decoding time is a fundamental property of mRNA translation believed to affect the abundance, function, and properties of proteins. Recently, a novel experimental technology--ribosome profiling--was developed to measure the density, and thus the speed, of ribosomes at codon resolution. Specifically, this method is based on next-generation sequencing, which theoretically can provide footprint counts that correspond to the probability of observing a ribosome in this position for each nucleotide in each transcript. Results In this study, we report for the first time various novel properties of the distribution of codon footprint counts in five organisms, based on large-scale analysis of ribosomal profiling data. We show that codons have distinctive footprint count distributions. These tend to be preserved along the inner part of the ORF, but differ at the 5' and 3' ends of the ORF, suggesting that the translation-elongation stage actually includes three biophysical sub-steps. In addition, we study various basic properties of the codon footprint count distributions and show that some of them correlate with the abundance of the tRNA molecule types recognizing them. Conclusions Our approach emphasizes the advantages of analyzing ribosome profiling and similar types of data via a comparative genomic codon-distribution-centric view. Thus, our methods can be used in future studies related to translation and even transcription elongation.
    BMC Genomics 10/2014; 15(Suppl 6):S13. · 4.04 Impact Factor


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