Mutations in an upstream regulatory sequence that increase expression of the bacteriophage T4 lysozyme gene

A P22 hybrid phage bearing the bacteriophage T4 lysozyme gene (e), as well as T4 sequences upstream from the lysozyme gene, was constructed. Amber mutations were introduced into gene e in the hybrid phage, and the resulting mutant phages were tested for the ability to form plaques on amber suppressor strains. Revertant phages that were able to form plaques on amber suppressors that did not suppress the parent amber mutant phages were isolated following UV mutagenesis. Secondary site pseudorevertants were identified among the revertants by a genetic test. Four of the suppressing secondary site mutations were mapped and sequenced. They were found to consist of small sequence alterations immediately upstream from gene e, all of which would tend to destabilize potential base-pairing interactions in the transcript. The mutations were shown to increase lysozyme expression when introduced into an otherwise wild-type hybrid phage, but were found to have little effect on transcription of the lysozyme gene.