Inactivation of the monofunctional peptidoglycan glycosyltransferase SgtB allows Staphylococcus aureus to survive in the absence of lipoteichoic acid

The cell wall of Staphylococcus aureus is composed of peptidoglycan and the anionic polymers lipoteichoic acid (LTA) and wall teichoic acid. LTA is required for growth and normal cell morphology in S. aureus. Strains lacking LTA are usually only viable when grown under osmotically stabilizing conditions or after the acquisition of compensatory mutations. LTA negative suppressor strains with inactivating mutations in gdpP, resulting in an increase in intracellular c-di-AMP levels, have been described previously. Here, we sought to identify factors other than c-di-AMP that allow S. aureus to survive without LTA. LTA-negative strains able to grow in un-supplemented medium were obtained and found to contain mutations in sgtB, mazE, clpX or vraT. The growth improvement through mutations in mazE and sgtB was confirmed by complementation analysis. We also show that an S. aureus sgtB transposon mutant, inactivated for the monofunctional peptidoglycan glycosyltransferase SgtB, displays a 4-fold increase in the MIC towards Oxacillin, suggesting that alteration in the peptidoglycan structure could help bacteria compensate for the lack of LTA. Muropeptide analysis of peptidoglycan isolated from a WT and sgtB mutant strains did not reveal any sizable alternations in the peptidoglycan structure. In contrast, the peptidoglycan isolated from an LTA-negative ltaS mutant strain showed a significant reduction in the fraction of highly crosslinked peptidoglycan, which was partially rescued in the sgtB ltaS double mutant suppressor strain. Taken together, these data point towards an important function of LTA in cell wall integrity through its requirement for proper peptidoglycan assembly.

金黄色葡萄球菌（Staphylococcus aureus）的细胞壁由肽聚糖（peptidoglycan）以及阴离子聚合物脂磷壁酸（lipoteichoic acid, LTA）和壁磷壁酸组成。LTA是金黄色葡萄球菌生长及维持正常细胞形态所必需的物质。缺失LTA的菌株通常仅能在渗透压稳定的培养条件下存活，或在获得补偿突变后方可存活。此前已有研究报道，携带gdpP失活突变的LTA阴性抑制突变株可使胞内环二腺苷酸（c-di-AMP）水平升高。本研究旨在鉴定除c-di-AMP之外，能够使金黄色葡萄球菌在缺失LTA的情况下存活的其他因子。我们成功获得了可在未补充培养基中生长的LTA阴性菌株，经检测发现这些菌株的sgtB、mazE、clpX或vraT基因均携带突变。通过互补实验证实，mazE与sgtB突变可改善菌株生长情况。此外，本研究还发现，针对单功能肽聚糖糖基转移酶SgtB的金黄色葡萄球菌sgtB转座子突变株，其对苯唑西林（Oxacillin）的最低抑菌浓度（minimum inhibitory concentration, MIC）提升了4倍，这表明肽聚糖结构的改变可能帮助细菌弥补LTA缺失带来的缺陷。对野生型（wild type, WT）与sgtB突变株分离得到的肽聚糖进行肽谱分析，未发现肽聚糖结构存在显著变化。与之相反，从LTA阴性的ltaS突变株中分离得到的肽聚糖，其高度交联肽聚糖的占比显著降低，而sgtB ltaS双突变抑制株的该表型可得到部分恢复。综上，本研究数据表明，LTA通过调控肽聚糖的正常组装，在维持细胞壁完整性方面发挥重要作用。