The ClpX chaperone controls theStaphylococcus aureuscell cycle but can be bypassed by β-lactam antibiotics

AbstractThe worldwide spread ofStaphylococcus aureusstrains resistant to almost all β-lactam antibiotics is of major clinical concern. β-lactams interfere with cross-linking of the bacterial cell wall, but the killing mechanism of this important class of antibiotics is not fully understood. Here we show that sub-lethal doses of β-lactams stimulate the growth ofS. aureusmutants lacking the widely conserved chaperone ClpX.S. aureus clpXmutants have a severe growth defect at temperatures below 37°C, and we reasoned that a better understanding of this growth defect could provide novel insights into how β-lactam antibiotics interfere with growth ofS. aureus. We demonstrate that ClpX is important for coordinating theS. aureuscell cycle, and thatS. aureuscells devoid of ClpX fail to divide, or lyze spontaneously, at high frequency unless β-lactams are added to the growth medium. Super-resolution imaging revealed thatclpXcells display aberrant septum synthesis, and initiate daughter cell separation prior to septum completion at 30°C, but not at 37°C. FtsZ localization and dynamics were not affected in the absence of ClpX, suggesting that ClpX affects septum formation and autolytic activation downstream of Z-ring formation. Interestingly, β-lactams restored septum synthesis and prevented premature autolytic splitting ofclpXcells. Strikingly, inhibitors of wall teichoic acid (WTA) biosynthesis that work synergistically with β-lactams to kill MRSA synthesis also rescued growth of theclpXmutant, underscoring a functional link between the PBP activity and WTA biosynthesis. The finding that β -lactams can prevent lysis and restore septum synthesis of a mutant with dysregulated cell division lends support to the idea that PBPs function as coordinators of cell division and that β -lactams do not killS. aureussimply by weakening the cell wall.Author SummaryThe bacteriumStaphylococcus aureusis a major cause of human disease, and the rapid spread ofS. aureusstrains that are resistant to almost all β-lactam antibiotics has made treatment increasingly difficult. β-lactams interfere with cross-linking of the bacterial cell wall but the killing mechanism of this important class of antibiotics is still not fully understood. Here we provide novel insight into this topic by examining a definedS. aureusmutant that has the unusual property of growing markedly better in the presence of β-lactams. Without β-lactams this mutant dies spontaneously at a high frequency due to premature separation of daughter cells during cell division. Cell death of the mutant can, however, be prevented either by exposure to β-lactam antibiotics or by inhibiting synthesis of wall teichoic acid, a major component of the cell wall in Gram-positive bacteria with a conserved role in activation of autolytic splitting of daughter cells. The finding that the detrimental effect of β-lactam antibiotics can be reversed by a mutation that affect the coordination of cell division emphasizes the idea that β-lactams do not killS. aureussimply by weakening the cell wall but rather by interference with the coordination of cell division.