Engineering bacterial translation initiation — Do we have all the tools we need?

• Published in: Biochimica et biophysica acta. General subjects Vol. 1861; no. 11; pp. 3060 - 3069
• Main Authors: Vigar, Justin R.J., Wieden, Hans-Joachim
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 01.11.2017
• Subjects: bacteria; Bacteria - genetics; Bacteria - metabolism; Codon, Initiator - genetics; Codon, Initiator - metabolism; Escherichia coli; gene expression; Gene Expression Regulation, Bacterial; ligands; messenger RNA; Peptide Chain Initiation, Translational - genetics; Protein Biosynthesis - genetics; Protein Engineering - methods; protein synthesis; Rational design; regulatory sequences; Riboregulation; Ribosome engineering; ribosomes; Ribosomes - metabolism; RNA engineering; RNA, Messenger - genetics; RNA, Messenger - metabolism; RNA-binding proteins; Synthetic biology; synthetic genes; translation (genetics); Translation initiation; Synthetic biology; Rational design; Ribosome engineering; RNA engineering; Translation initiation; Riboregulation
• ISSN: 0304-4165; 1872-8006
• DOI: 10.1016/j.bbagen.2017.03.008

Reliable tools that allow precise and predictable control over gene expression are critical for the success of nearly all bioengineering applications. Translation initiation is the most regulated phase during protein biosynthesis, and is therefore a promising target for exerting control over gene expression. At the translational level, the copy number of a protein can be fine-tuned by altering the interaction between the translation initiation region of an mRNA and the ribosome. These interactions can be controlled by modulating the mRNA structure using numerous approaches, including small molecule ligands, RNAs, or RNA-binding proteins. A variety of naturally occurring regulatory elements have been repurposed, facilitating advances in synthetic gene regulation strategies. The pursuit of a comprehensive understanding of mechanisms governing translation initiation provides the framework for future engineering efforts. Here we outline state-of-the-art strategies used to predictably control translation initiation in bacteria. We also discuss current limitations in the field and future goals. Due to its function as the rate-determining step, initiation is the ideal point to exert effective translation regulation. Several engineering tools are currently available to rationally design the initiation characteristics of synthetic mRNAs. However, improvements are required to increase the predictability, effectiveness, and portability of these tools. Predictable and reliable control over translation initiation will allow greater predictability when designing, constructing, and testing genetic circuits. The ability to build more complex circuits predictably will advance synthetic biology and contribute to our fundamental understanding of the underlying principles of these processes. "This article is part of a Special Issue entitled "Biochemistry of Synthetic Biology - Recent Developments" Guest Editor: Dr. Ilka Heinemann and Dr. Patrick O’Donoghue. [Display omitted] •RNA-mediated regulation mechanisms control gene expression post-translationally.•Unraveling translation initiation mechanisms has enabled rational design tools.•Bioengineers have successfully exploited RNA-based regulation strategies.•Limitations exist regarding predictability, portability, and dynamic range.•Non-canonical mechanisms will provide tools to address engineering challenges.