A Forward-Genetic Screen and Dynamic Analysis of Lambda Phage Host-Dependencies Reveals an Extensive Interaction Network and a New Anti-Viral Strategy
Nathaniel D. Maynard, Elsa W. Birch, Jayodita C. Sanghvi, Lu Chen, Miriam V. Gutschow, Markus W. Covert
PLoS Genet 6(7): e1001017. doi:10.1371/journal.pgen.1001017
Latently infecting viruses are an important class of virus that plays a key role in viral evolution and human health. Here we report a genome-scale forward-genetics screen for host-dependencies of the latently-infecting bacteriophage lambda. This screen identified 57 Escherichia coli (E. coli) genes—over half of which have not been previously associated with infection—that when knocked out inhibited lambda phage’s ability to replicate. Our results demonstrate a highly integrated network between lambda and its host, in striking contrast to the results from a similar screen using the lytic-only infecting T7 virus. We then measured the growth of E. coli under normal and infected conditions, using wild-type and knockout strains deficient in one of the identified host genes, and found that genes from the same pathway often exhibited similar growth dynamics. This observation, combined with further computational and experimental analysis, led us to identify a previously unannotated gene, yneJ, as a novel regulator of lamB gene expression. A surprising result of this work was the identification of two highly conserved pathways involved in tRNA thiolation—one pathway is required for efficient lambda replication, while the other has anti-viral properties inhibiting lambda replication. Based on our data, it appears that 2-thiouridine modification of tRNAGlu, tRNAGln, and tRNALys is particularly important for the efficient production of infectious lambda phage particles.
In this study, we took advantage of a new genetic resource for E. coli mutants to screen for previously undiscovered lambda phage host-dependencies. We then assessed the dynamics of infection in these different E. coli mutants and applied a mathematical model of infection in an attempt to further classify the role of these novel interactions. This model-driven approach to biological discovery led us to identify the previously uncharacterized gene yneJ as a regulator of lamB gene expression. In addition, we identified two highly conserved pathways involved in post-transcriptional modification of tRNA—one pathway was required for efficient lambda replication, while the other has anti-viral properties inhibiting lambda replication. This finding is important as it illustrates a new potential anti-viral strategy that could be applied broadly to other viruses.