Modulation ofccrABExpression and SCCmecExcision by an Inverted Repeat Element and SarS in Methicillin-Resistant Staphylococcus aureus

ABSTRACTMethicillin-resistantStaphylococcus aureus(MRSA) is a notorious human pathogen that can cause a broad spectrum of infections. MRSA strains are resistant to almost the entire family of β-lactam antibiotics due to the acquisition of staphylococcal cassette chromosomemec(SCCmec). The chromosome cassette recombinases A and B, encoded byccrABgenes located on SCCmec, play a key role in the excision of SCCmec. Studies have shown thatccrABgenes are expressed in only a minority of cells, suggesting the involvement of a subtle regulatory mechanism inccrABexpression which has not been uncovered. Here, we found that an inverted repeat (IR) element, existing extensively and conservatively within theccrABpromoter of different SCCmectypes, played a repressive role inccrABexpression and SCCmecexcision in MRSA strain N315. Replacement of the IR sequence led to a significant increase inccrABexpression and curing of SCCmecfrom strain N315 cells. In addition, we identified the transcriptional regulator SarS using DNA-affinity chromatography and further demonstrated that SarS can bind to the IR sequence and upregulateccrABexpression and SCCmecexcision. These findings reveal a molecular mechanism regulatingccrABexpression and SCCmecexcision and may provide mechanic insights into the lateral transfer of SCCmecand spread of antibiotic resistance inS. aureus.

Transcription factor p53 exhibits increased binding to the α2-macroglobulin gene promoter and decreased glycosylation in fetal and adult rat liver during the acute-phase response

The binding affinity of p53 for the MG promoter was assessed by DNA-affinity chromatography with the extended ?2-macroglobulin (MG) gene promoter (-852/+12) and immunoblot analysis. During the increased MG gene transcription observed in the fetus and the acute-phase (AP) response in both the fetus and the adult, p53 exhibited increased binding to the MG promoter. This increase was accompanied by decreased O-linked N-acetyl glucosamine glycosylation of p53. We suggest that the enzymatic removal of sugar moieties in vivo serves to activate the MG gene promoter binding potential of p53 and its participation in upregulated MG gene transcription.

Chlamydial GroEL Autoregulates Its Own Expression through Direct Interactions with the HrcA Repressor Protein

ABSTRACT In the pathogenic bacterium Chlamydia trachomatis, a transcriptional repressor, HrcA, regulates the major heat shock operons, dnaK and groE. Cellular stress causes a transient increase in transcription of these heat shock operons through relief of HrcA-mediated repression, but the pathway leading to derepression is unclear. Elevated temperature alone is not sufficient, and it is hypothesized that additional chlamydial factors play a role. We used DNA affinity chromatography to purify proteins that interact with HrcA in vivo and identified a higher-order complex consisting of HrcA, GroEL, and GroES. This endogenous HrcA complex migrated differently than recombinant HrcA, but the complex could be disrupted, releasing native HrcA that resembled recombinant HrcA. In in vitro assays, GroEL increased the ability of HrcA to bind to the CIRCE operator and to repress transcription. Other chlamydial heat shock proteins, including the two additional GroEL paralogs present in all chlamydial species, did not modulate HrcA activity.