Focus On: Genome Recoding

By reprogramming cells to work from an altered genetic code researchers hope to create organisms with resistance to viral DNA - and exciting new biochemistry

The Biologist 66(1) p30-31

The genetic code that cells use to make proteins from DNA varies very little throughout the entirety of life on Earth. Around 60 different three-letter nucleotide sequences, or codons, are used by cells to represent the 20 amino acids from which all proteins are made. This means a single amino acid can be represented by several different codons – for example, the sequences GCG, GCC and GCA all represent alanine.

Genome recoding exploits this in-built redundancy in the genetic code, reprogramming some of these ‘spare’ codons to create organisms with alternative genetic codes.

How does it work?

Replacing all instances of one codon in a genome with another, synonymous codon means the cell can still function normally. For example, replacing all instances of ‘GCG’ in an organism’s DNA sequence with ‘GCC’ would not affect protein production as they both are translated into alanine when proteins are made. 

If the transfer RNA (tRNA) that translates the deleted codon is also removed, the cell will no longer be able to translate any DNA sequence containing the eliminated codon. So the cell functions as normal, but cannot read other DNA from other (i.e. non-recoded) cells or viruses, because it cannot read the sections containing the deleted codon (i.e. GCG).

Why do that?

Eliminating redundant codons as described above effectively means any viral DNA that enters the cell following recoding will not be translated properly as long as it contains the eliminated codons. Remarkably, recoding in this way could in theory make organisms resistant to all viruses, even ones not yet studied.

‘Redundant’ codons can also be repurposed to code for non-natural amino acids, enabling researchers to create proteins with functions not found in nature. This approach requires the relevant tRNA to be engineered so it binds to a non-natural amino acid, but opens up many possibilities in terms of what DNA can then code for.

“Whole new dimensions of biochemistry emerge when you are not limited to the universal and ordinary 20 amino acids,” Harvard geneticist George Church wrote about genome recoding as part of Nature’s ‘Technology to watch in 2018’[1].

A broader purpose of genome recoding is what has been described as ‘isolation from nature’. In other words, recoding can make a cell chemically incompatible with other cells and environments. If an alternative genetic code is created in a cell or organism, or an organism is recoded to be dependent on a synthetic amino acid, it would mean their genes will not work in other natural (unrecoded) organisms, or the organism would die outside certain environments.

This could be used to prevent genes from genetically modified organisms (GMOs) spreading to other organisms (effectively blocking horizontal gene transfer between bacterial species, for example) or as a biocontainment measure (so the organism dies if outside a highly specific nutrient culture) – providing new ‘firewalls’ to prevent unpredictable consequences from the release of GMOs in the environment.

Recoding in Nature Natural recoding of the genome, where an organism reassigns a codon to a different ‘meaning’ in the cells, has been observed in around 20 species – often for stop codons

What next?

Following the Church group’s first successful recoding of an E. coli genome[2] in 2011, several approaches[3,4] have been employed to successfully incorporate codon changes at thousands of positions in bacteria. Some involve recoding genomes in segments before rebuilding the genome in parts. The Synthetic Yeast Genome Project is a large effort to build five recoded yeast chromosomes from scratch, with segments being worked on by multiple organisations that will eventually be combined into one organism.

More ambitious genome recoding projects will require tens of thousands of genetic changes throughout the genome, rather than hundreds, and work on the de novo synthesis of entire genomes5 is likely to aid future genome recoding work.

Although the actual number of successfully recoded organisms remains quite small, the theoretical benefits and applications of genome recoding (virus resistance, genetic firewalls and novel protein synthesis) make it a very exciting area indeed.

Focus genome lrgeIn genome recoding, all instances of a particular codon are replaced with a synonymous codon (1). In the first example of successful genome-wide recoding, George Church’s Harvard laboratory changed all 321 TAG codons in E. coli to TAA. (Both these codons are translated as ‘stop’ by the cell, terminating translation.) In this diagram, GCG is replaced with GCC. The cell continues to function normally, as the replacement codon and deleted codon are synonymous. Next, the gene for the tRNA which translates the eliminated codon from its mRNA form is also deleted (2). The cell can no longer properly translate DNA sequences with the GCG codon. 

Now, when a virus infects the cell, the cell cannot translate proteins from the viral DNA (or, more accurately, from its mRNA) because of the missing tRNA. The viral DNA is effectively incompatible with the cell’s translation toolkit. 

Further reading
Kuo, J. et al. Synthetic genome recoding: New genetic codes for new features. Current Genetics 64(1) (2017).

References
1) ‘Technology to watch in 2018’, Nature.com, 24 January 2018.
2) Isaacs, F. J. et al. Precise manipulation of chromosomes in vivo enables genome-wide codon replacement. Science 333, 348–353 (2011). doi:10.1126/science.1205822
3) Lau, Y. H. et al. Large-scale recoding of a bacterial genome by iterative recombineering of synthetic DNA. Nucleic Acids Res. 45(11), 6971–6980 (2017).
4) Ostrov, N. et al. Design, synthesis, and testing toward a 57-codon genome. Science 353, 819–822 (2016).
5) Hutchison, C. A. 3rd et al. Design and synthesis of a minimal bacterial genome. Science 351, aad6253 (2016).

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