The Spx stress regulator confers high-level β-lactam resistance and decreases susceptibility to last-line antibiotics in methicillin resistant Staphylococcus aureus

Infections caused by methicillin resistant Staphylococcus aureus (MRSA) are a leading cause of mortality worldwide. MRSA have acquired resistance to next generation {beta}-lactam antibiotics through the horizontal acquisition of the mecA resistance gene. Development of high resistance is, however, often associated with additional mutations in a set of chromosomal core genes, known as potentiators which through poorly described mechanisms enhance resistance. The yjbH gene was recently identified as a hot spot for adaptive mutations during severe infections. Here, we show that inactivation of yjbH increased {beta}-lactam MICs up to 16-folds and transformed MRSA cells with low level of resistance to being homogenously highly resistant to {beta}-lactams. The yjbH gene encodes an adaptor protein that targets the transcriptional stress regulator Spx for degradation by the ClpXP protease. Using CRISPRi to knock down spx transcription, we unambiguously linked hyper-resistance to accumulation of Spx. Spx was previously proposed to be essential, however, our data indicate that Spx is dispensable for growth at 37{degrees}C but becomes essential in the presence of antibiotics with various targets. On the other hand, high Spx levels bypassed the role of PBP4 in {beta}-lactam resistance and broadly decreased MRSA susceptibility to compounds targeting the cell wall or the cell membrane including vancomycin, daptomycin, and nisin. Strikingly, Spx potentiated resistance independently of its redox sensing switch. Collectively, our study identifies a general stress pathway that, in addition to promoting the development of high-level, broad-spectrum {beta}-lactam resistance, also decreases MRSA susceptibility to critical antibiotics of last resort.