Translational Sensitivity and tRNA Availability – Applications in Synthetic Biology

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Sibylle Wohlgemuth, Thomas Gorochowski, Hans Roubos

DSM/ETH Zurich, Netherlands

Translational processes play a role in regulating genome-wide protein expression levels. While the translational initiation step and codon usage have received much attention, we focus on another aspect, namely tRNA availability. This factor conditionally influences translational speed along a transcript and has been linked to co-translational processes like protein folding. Recent studies have shown that tRNA pools can significantly vary under different growth and stress conditions. Due to tRNAs acting as a shared resource and the ability for synonymous codons to code the same amino acid, modulating tRNA concentrations offers the cell a way to precisely adapt translational dynamics across the genome. With the aim of understanding how such a mechanism is used, we developed a generalized computational workflow that estimates the translational speed of codons along a given transcript and generated speed profiles. We show data for all coding sequences in the E.coli genome. Unlike previous studies that assumed fixed tRNA concentrations, we investigated the sensitivity of these profiles to large shifts in tRNA availability, as experimentally measured during amino acid starvation and for differing growth rates. Visualization of profiles across the entire genome highlighted key gene clusters that were either particularly sensitive or insensitive to tRNA level change, and surprising temporal responses with links to genes that are known to use translational control to alter protein expression level. In addition to analyzing how tRNA availability might be used by E.coli as such, we also consider applications as a control mechanism for synthetic biology. We show how our developed workflow can be applied to: (i) forward-engineering of new protein parts where translational dynamics will respond in a specific way to known changes in tRNA pools; and (ii) help in the design of circuits to artificially modulate tRNA availability to control translational dynamics of endogenous genes across the genome.