Methicillin-Resistance in Staphylococcus aureus Is Not Affected by the Overexpression in Trans of the mecA Gene Repressor: A Surprising Observation

Abstract

Methicillin-resistant Staphylococcus aureus (MRSA) is intrinsically cross-resistant to virtually all β-lactam antibiotics. The central determinant for the MRSA phenotype is the mecA gene, whose transcriptional control may be mediated by a repressor (mecI) and a sensor/inducer (mecR1). The mecI-mecR1-mediated induction of mecA takes several hours rendering the strains phenotypically susceptible in spite of the presence of the resistance gene. Therefore, it has been proposed that the full resistance to β-lactams observed in many contemporary clinical MRSA strains requires a non-functional mecI-mecR1 regulatory system. The mecA gene is embedded in a large chromosomal cassette (the SCCmec element) for which several structural types have been described. Some epidemic MRSA clones, typically expressing full β-lactam resistance, carry SCCmec elements that contain an intact mecI-mecR1 locus (e.g. SCCmec types II and III). We have addressed this apparent contradiction by first sequencing the mecI coding region and mecA promoter sequences in a collection of prototype MRSA strains characterized by different SCCmec types. A conserved non-sense mutation within mecI was detected in all SCCmec type III strains tested, presumably responsible for a non-functional truncated MecI protein and, therefore, explaining the full resistance phenotype. In SCCmec type II strains no conserved mutations were found. We next transformed a collection of prototype MRSA epidemic strains with a recombinant plasmid overexpressing a wild-type copy of mecI. Surprisingly, for the great majority of the strains no significant alterations in the phenotypic expression of β-lactam resistance could be detected. These findings were confirmed and further explored, challenging the currently accepted mechanism of mecA transcriptional control. Our observations suggest the existence of yet unidentified additional determinants involved in the transcriptional control of mecA gene and point to a revision of the mecA regulatory mechanism in contemporary MRSA strains.

Redefining the Role of the β-Lactamase Locus in Methicillin-Resistant Staphylococcus aureus: β-Lactamase Regulators Disrupt the MecI-Mediated Strong Repression on mecA and Optimize the Phenotypic Expression of Resistance in Strains with Constitutive mecA Expression

“…In previous studies, upon the overexpression in trans of MecI in a representative collection of MRSA strains, we have unexpectedly observed that in most cases there was no significant effect on the phenotypic expression of ␤-lactam resistance (20). Follow-up studies have demonstrated that, for those strains with functional mecR1-mecI regulatory loci, the observed phenomenon was due to the presence of a previously unrecognized third regulatory gene, mecR2, coding for a potent antirepressor (19).…”

“…MecR2 interacts with MecI, disturbing its binding to the mecA promoter and fostering its inactivation by proteolytic cleavage (19). The function of MecR2 as an antirepressor accounts for the unexpected lack of effect on the phenotypic expression of ␤-lactam resistance upon overexpression in trans of a wild-type copy of MecI observed in most MRSA strains (20). Nevertheless, in a few strains negative for the bla locus, there was a massive decline of the resistance phenotype upon the overexpression of MecI in trans.…”

Proteolysis of mecA Repressor Is Essential for Expression of Methicillin Resistance by Staphylococcus aureus

“…In previous studies, we observed that, contrary to what was expected, the overexpression in trans of MecI in a representative collection of prototype MRSA strains did not cause significant effects on the expression of ␤-lactam resistance in most cases (7). Later, we demonstrated that the "MecI overexpression protection effect" was due to the presence of a third regulator in the mecA locus, MecR2, acting as a potent antirepressor and fostering the proteolysis of MecI (1).…”

Structure-Function Studies of the Staphylococcal Methicillin Resistance Antirepressor MecR2

“…Second, the structure of MecI and BlaI in complexes with target DNA revealed that the repressor cleavage site is found within an ␣-helix and is not surface-accessible (29 -32), as would be required for proper proteolytic processing. Third, highly resistant MRSA strains did not show significant variation in the phenotypic expression of resistance when wild-type MecI was overexpressed in trans (33). Lastly, the presence of functional MecR1-MecI did not correlate with the level of BLA resistance in a representative collection of epidemic MRSA strains (33).…”

“…Third, highly resistant MRSA strains did not show significant variation in the phenotypic expression of resistance when wild-type MecI was overexpressed in trans (33). Lastly, the presence of functional MecR1-MecI did not correlate with the level of BLA resistance in a representative collection of epidemic MRSA strains (33). These and other findings led several authors to postulate the existence of a further regulatory element, MecR2/BlaR2, although no candidate molecules were suggested (4, 10, 17, 18, 31, 32, 34 -38).…”