Abstract:

Removing cellular transfer RNAs (tRNAs), making their cognate codons unreadable, creates a genetic firewall that prevents viral replication and horizontal gene transfer. However, numerous viruses and mobile genetic elements encode parts of the translational apparatus, including tRNAs, potentially rendering a genetic-code-based firewall ineffective. In this paper, we show that such horizontally transferred tRNA genes can enable viral replication in Escherichia coli cells despite the genome-wide lack of three codons and the previously essential cognate tRNAs and release factor 1. By repurposing viral tRNAs, we then develop recoded cells bearing an amino-acid-swapped genetic code that reassigns two of the six serine codons to leucine during translation. This amino-acid-swapped genetic code renders cells completely resistant to viral infections by mistranslating viral proteomes and prevents the escape of synthetic genetic information by engineered reliance on serine codons to produce leucine-requiring proteins. Finally, we also repurpose the third free codon to biocontain this virus-resistant host via dependence on an amino acid not found in nature.Competing Interest StatementThe authors declare competing financial interests. Harvard Medical School has filed a provisional patent application related to this work on which A.N., S.V., and G.M.C. are listed as inventors. M.L., K.C., and F.H. are employed by GenScript USA Inc., but the company had no role in designing or executing experiments. G.M.C. is a founder of the following companies in which he has related financial interests: GRO Biosciences, EnEvolv, and 64x Bio. Other potentially relevant financial interests of G.M.C. are listed at http://arep.med.harvard.edu/gmc/tech.html.