Translation efficiency is maintained at elevated temperature in Escherichia coli

Abstract

Cellular protein levels are dictated by the balance between gene transcription, mRNA translation, and protein degradation, among other factors. Translation requires the interplay of several RNA hybridization processes, which are expected to be temperature-sensitive. We used ribosome profiling to monitor translation in at 30 °C and to investigate how this changes after 10-20 min of heat shock at 42 °C. Translation efficiencies are robustly maintained after thermal heat shock and after mimicking the heat-shock response transcriptional program at 30 °C by overexpressing the heat shock σ factor encoded by the gene. We compared translation efficiency, the ratio of ribosome footprint reads to mRNA reads for each gene, to parameters derived from gene sequences. Genes with stable mRNA structures, non-optimal codon use, and those whose gene product is cotranslationally translocated into the inner membrane are generally less highly translated than other genes. Comparison with other published datasets suggests a role for translational elongation in coupling mRNA structures to translation initiation. Genome-wide calculations of the temperature dependence of mRNA structure predict that relatively few mRNAs show a melting transition between 30 and 42 °C, consistent with the observed lack of changes in translation efficiency. We developed a linear model with six parameters that can predict 38% of the variation in translation efficiency between genes, which may be useful in interpreting transcriptome data.

Proteolytic queues at ClpXP increase antibiotic tolerance

“…The increase in antibiotic tolerance due to queue formation at ClpXP may be specific to overexpression of the LAA-tag, especially when considering that the number of LAA tagged proteins naturally increases during stress. The number of proteins with LAA tags increase during heat shock 52 , and queue formation at the proteases is likely a consequence of the increasing cellular traffic. If the native LAA tag is removed from SsrA while maintaining the ribosome rescue function, the survival of ampicillin treatment decreases in E. coli 21 .…”

From reporters to endogenous genes: the impact of the first five codons on translation efficiency in Escherichia coli

“…Next, we compared the score of each E. coli gene calculated at codon positions 3-5 (SCORE GENOME) with independent metrics used to estimate TE [18][19][20] , protein abundance 19,21 , mRNA abundance 20,22-24 and mRNA secondary structure (Gini index) 20 . As a control, we used an E. coli scramble genome that maintained the proportions of all 61 amino acid codons but randomized the codon choice for each transcript (SCORE SCRAMBLE).…”

“…The translation efficiency of these two groups of genes was compared using three different datasets [18][19][20] . Regardless of the TE dataset utilized, the group of genes with higher scores have higher TE when compared to genes with lower scores ( Figure 4B-D).…”

“…Ribosome profiling data E. coli ribosome profiling data were treated as described previously 7,17,18 . The data were analyzed as described by Ingolia and collaborators 17 except that the program used here was Geneious R11…”

Multimapping confounds ribosome profiling analysis: A case‐study of the Hsp90 molecular chaperone

“…In addition to publicly available computational tools, lab-specific scripts also have been used to deal with multimapping reads on a case by case basis. 5,26,27 In addition to computational approaches, coupling RNA-seq and Ribo-seq data is proposed to reduce multimapping when ambiguous reads are the result of multiple isoforms, 21 such as in human cell lines. For example, Ribomap, 21 uses RNA-seq to estimate mRNA isoform abundance which is then used to guide assignment of the ambiguous Ribo-seq reads.…”