Genomics and Proteomics of Mycobacteriophage Patience, an Accidental Tourist in the Mycobacterium Neighborhood

Pope, Welkin H.; Jacobs-Sera, Deborah; Russell, Daniel A.; Rubin, Daniel H. F.; Kajee, Afsana; Msibi, Zama N. P.; Larsen, Michelle H.; Jacobs, William R.; Lawrence, Jeffrey G.; Hendrix, Roger W.; Hatfull, Graham F.

Although codon usage shows that Patience spent a significant portion of its evolutionary past in hosts with moderate GC contents, it is not clear what its current host range is. If Patience currently exploits more GC-rich hosts, then it would experience not only mutational biases, which would increase its GC content, but selective pressure on its most highly expressed genes to use tRNA pools poised to translate GC-rich genes. This selective pressure would be reflected in a preferred usage of GC-rich codons within highly expressed genes. Patience does experience codon selection, which is shown by the robust positive correlation between codon selection (adapative codon enrichment [ACE z ] [25] ) (see Materials and Methods) and level of gene expression, using total numbers of peptides as identified by mass spectrometry under stringent conditions as a surrogate for expression (Fig. 6B) . Codon selection is measured using ␦ values, or the ratio of codon frequencies in genes experiencing codon selection normalized to their frequencies in genes lacking codon selection. Preferred codons are those with ␦ values greater than 1, indicating more frequent use in genes experiencing codon selection, frequently those expressed to greater levels. Patience favors the use of GCrich codons in genes experiencing codon selection (Table 2) , with 24 of 33 preferred codons bearing GC base pairs in their third positions, whereas 21 of 26 nonpreferred codons end with AT base pairs. Moreover, the patterns of codon preference in Patience (which codons are preferred and which are not) are strongly correlated with that of M. smegmatis (Fig. 6C) , suggesting that this host imposes codon selection congruent with that currently being experienced by Patience. In contrast, Patience codon selection is not congruent with that imposed by hosts with more moderate GC content, such as Escherichia coli (Fig. 6C, inset) , although such surveys can never be conclusive.