| Lipoteichoic and wall teichoic acids (TA) are highly anionic cell envelope-associated polymers containing repeating glycerol/ribitol phosphate moieties. Substitution of TA with D-alanine is important for modulation of cell envelope-dependent processes such as activity of autolytic enzymes, binding of divalent cations and susceptibility of Staphylococcus aureus to innate host defenses such as mammalian Group IIA phospholipase A2 (PLA 2). Highly cationic properties of these PLA2 are important for Ca2+-independent binding and cell wall penetration, pre-requisites for Ca2+-dependent degradation of membrane phospholipids and bacterial killing. To further delineate charge properties of the bacterial envelope important in Group IIA PLA2 action against S. aureus, we examined the effects of mutations that prevent specific modifications of cell wall (dltA) and cell membrane ( mprF) polyanions. Differences in PLA2 sensitivity of intact bacteria reflect differences in cell wall, not cell membrane, properties since protoplasts from wild type and mutant strains are equally sensitive to the PLA2. Diminished positive charge in PLA2 reduces PLA 2 binding and antibacterial activity. In contrast, diminished cell wall negative charge by substitution of TA with D-alanine reduces antibacterial activity of bound PLA2, but not initial PLA2 binding. Therefore, the potent antistaphylococcal activity of Group IIA PLA2 depends on cationic properties of the enzyme that promote binding to the cell wall, and polyanionic properties of TA that promote attack of membrane phospholipids by bound PLA2.;D-alanylation of TA is diminished when bacteria are grown in medium containing increased NaCl concentrations, but effects of increased salt concentration on expression of the dlt operon encoding proteins mediating D-alanylation of TA are unknown. We demonstrate that S. aureus transcriptionally represses dlt expression in response to high concentrations of Na+ and moderate concentrations of Mg 2+ and Ca2+, but not sucrose. Changes in dlt mRNA are induced within 15 min and sustained for several generations of growth. Mg2+-induced dlt repression depends on the ArlSR two component system. Northern blot, RT-PCR and SMART(TM)-RACE analyses suggest that the dlt transcript begins 250 bp upstream of the dltA start codon and includes an open reading frame immediately upstream of dltA that is conserved in many Gram-positive bacterial species. |
