Formation of potent hybrid promoters of the mutant llm gene by IS256 transposition in methicillin-resistant Staphylococcus aureus

From high-level methicillin-resistant Staphylococcus aureus SRM551, the low-level heterogeneously resistant mutant, SRM563, was isolated by transposon mutagenesis. The transposon insertion occurred in the 3' region of the llm gene in the mutant (H. Maki, T, Yamaguchi, and K, Murakami, J, Bacteriol, 176:4993-5000, 1994), Resistant revertants were generated from the mutant strain SRM563 on the plate containing methicillin at a concentration of 12.5 mu g/ml or more. In some revertants, the insertion sequence IS256 was observed to be transposed into one of five sites localized 88 to 212 bp upstream of the mutant llm at a frequency of 2.8 x 10(-7) in the bacterial population, The IS256 transposition created a new hybrid promoter in which the -35 region at the end of IS256 was properly arranged in relation to the -10-like sequence upstream of llm. The new promoters greatly enhanced the transcription of the mutant llm, as judged by blotting analysis of llm mRNA, with concomitant elevation of the methicillin resistance. Involvement of the Insertion sequence in the hetero-resistance characteristics of methicillin-resistant S. aureus was suggested.