mecR1, although truncated in CHE482, was still transcribed and had the same expression pattern as mecA, as both became derepressed over time and had the highest transcript levels Panobinostat in vitro after 30 min of induction. In the mutant ΔCHE482, transcripts of both mecA and mecR1′ were unaffected by SA1665 deletion, indicating that SA1665 had no influence on their expression at

either OD 0.25 (Figure 5D) or OD 1.0 (data not shown). SA1665 deletion also had no effect on mecA transcription or induction in strains ZH37, ZH44 and ZH73 (data not shown). Western blot analysis Mutants of CHE482 and of ZH44 and ZH73, which had the largest differences in oxacillin GW4869 mouse resistance levels, were analysed by Western blot analysis to determine if SA1665 affected production of PBP2a from mecA. As shown in Figure 5E, all pairs of wild type and mutant strains had similar amounts of PBP2a present both before and after induction with cefoxitin, indicating selleck chemical that SA1665 deletion did not alter amounts of PBP2a produced. Therefore it seems that SA1665 exerts no direct control over mecA or PBP2a expression. Discussion Methicillin resistance in MRSA is primarily dependent

on the presence of the mecA gene, however, resistance levels are generally governed by strain-specific factors including mecA regulatory elements and other chromosomal fem/aux factors which either enhance or repress the expression of resistance. For instance, the very low-level methicillin resistance Glycogen branching enzyme of the Zurich drug clone CHE482, was shown to be controlled by its genetic background [12] suggesting that it either contained or lacked certain fem/aux factors involved in controlling resistance expression. Many of the currently known fem/aux factors are directly or indirectly involved in cell wall synthesis and turnover,

or envelope biogenesis, however there still remain factors of unknown function. Most of the currently known fem/aux factors reduce methicillin resistance levels when inactivated. A few genes, such as lytH, dlt, norG, sarV and cidA increase resistance levels upon inactivation or mutation. All of these genes, except norG, which is an efflux pump regulator, play a role in either autolysis or are important for cell physiology and growth [25–30]. Other genes increase β-lactam resistance upon overexpression, such as hmrA coding for a putative amidohydrolase, hmrB coding for a putative acyl carrier protein [31], or the NorG-controlled abcA multidrug efflux pump [28]. SA1665, a predicted DNA-binding transcriptional regulator, was found to bind to a DNA fragment containing the mecA promoter region. However, although this protein shifted the mecA operator/5′ coding sequence, it did not appear to directly control mecA or mecR1 transcription or PBP2a production. Therefore its binding to the mecA region may have no specific regulatory function.