Metabolic reprogramming and altered cell envelope characteristics in a pentose phosphate pathway mutant increases MRSA resistance to β-lactam antibiotics.
Metabolic reprogramming and altered cell envelope characteristics in a pentose phosphate pathway mutant increases MRSA resistance to β-lactam antibiotics.
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DOI：
10.1371/journal.ppat.1011536
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发表时间：
2023-07
期刊：
影响因子：
6.7
通讯作者：
O'Gara, James
中科院分区：
文献类型：
--
作者：
Zeden, Merve;Gallagher, Laura;Bueno, Emilio;Nolan, Aaron;Ahn, Jongsam D.;Shinde, Dhananjay;Razvi, Fareha C.;Sladek, Margaret P.;Burke, Orla A.;O'Neill, Eoghan;Fey, Paul A.;Cava, Felipe;Thomas, Vinai A.;O'Gara, James
中枢代谢途径控制着毒力和抗生素耐药性，是抗菌药物的潜在靶点。在金黄色葡萄球菌中，戊糖磷酸途径（PPP）的作用在很大程度上仍未被探索。编码PPP氧化阶段唯一非必需酶的6-磷酸葡萄糖醇内酯酶基因pgl的突变显著增加了MRSA对β-内酰胺类抗生素的耐药性，特别是在具有生理相关葡萄糖浓度的化学定义培养基中，并减少了oxacillin （OX）诱导的裂解。耐甲氧西林青霉素结合蛋白2a的表达和肽聚糖结构未受影响。碳追踪和代谢组学显示pgl突变体中广泛的代谢重编程，包括糖酵解通量增加，TCA循环和几种细胞包膜前体，这与β-内酰胺抗性增加一致。形态学上，pgl突变体细胞在OX中生长时比野生型细胞小，细胞壁厚，表面皱褶。pgl突变体降低了对刚果红、磺胺甲恶唑和氧化应激的抗性，增加了对塔戈西、磷霉素和万古霉素的抗性。脂质胆酸（LTAs）水平在pgl中显著降低，这可能限制了细胞的裂解，而pgl细胞的表面电荷明显更加阳性。pgl的vraG突变逆转了增加的OX抗性表型，并部分恢复了野生型的表面电荷，但没有恢复LTA水平。VraFG/ graRS复合体中的vraF或graRS的突变也恢复了野生型OX敏感性，VraFG/ graRS复合体调节dltabcd介导的天芥酸的d-丙烯酰化（反过来控制β-内酰胺抗性和表面电荷）。总的来说，这些数据表明，在MRSA pgl突变体中，LTAs和OX诱导裂解水平的降低，以及VraFG/ grars依赖性的细胞表面正电荷的增加，伴随着OX抗性的显著增加。对青霉素型（β-内酰胺）抗生素的高耐药性极大地限制了MRSA感染患者的治疗选择，需要使用更新的药物，已经描述了降低敏感性。在这里，我们首次报道中心代谢戊糖磷酸途径控制MRSA对青霉素型抗生素的耐药性。我们全面证明了PPP基因pgl的突变扰乱了MRSA的代谢，导致细胞包膜前体的通量增加，从而增加了抗生素耐药性。此外，抗性的增加与细胞包膜中脂壁酸水平的降低、β-内酰胺应激下细胞裂解率的降低有关，并依赖于控制壁酸d-丙烯酰化和细胞表面电荷的VraRG/GraRS多酶膜复合物。我们的数据为MRSA β-内酰胺耐药机制提供了新的见解，这将支持扩大由这种和其他抗菌素耐药病原体引起的感染的治疗选择。
Central metabolic pathways control virulence and antibiotic resistance, and constitute potential targets for antibacterial drugs. In Staphylococcus aureus the role of the pentose phosphate pathway (PPP) remains largely unexplored. Mutation of the 6-phosphogluconolactonase gene pgl, which encodes the only non-essential enzyme in the oxidative phase of the PPP, significantly increased MRSA resistance to β-lactam antibiotics, particularly in chemically defined media with physiologically-relevant concentrations of glucose, and reduced oxacillin (OX)-induced lysis. Expression of the methicillin-resistance penicillin binding protein 2a and peptidoglycan architecture were unaffected. Carbon tracing and metabolomics revealed extensive metabolic reprogramming in the pgl mutant including increased flux to glycolysis, the TCA cycle, and several cell envelope precursors, which was consistent with increased β-lactam resistance. Morphologically, pgl mutant cells were smaller than wild-type with a thicker cell wall and ruffled surface when grown in OX. The pgl mutation reduced resistance to Congo Red, sulfamethoxazole and oxidative stress, and increased resistance to targocil, fosfomycin and vancomycin. Levels of lipoteichoic acids (LTAs) were significantly reduced in pgl, which may limit cell lysis, while the surface charge of pgl cells was significantly more positive. A vraG mutation in pgl reversed the increased OX resistance phenotype, and partially restored wild-type surface charge, but not LTA levels. Mutations in vraF or graRS from the VraFG/GraRS complex that regulates DltABCD-mediated d-alanylation of teichoic acids (which in turn controls β-lactam resistance and surface charge), also restored wild-type OX susceptibility. Collectively these data show that reduced levels of LTAs and OX-induced lysis combined with a VraFG/GraRS-dependent increase in cell surface positive charge are accompanied by significantly increased OX resistance in an MRSA pgl mutant. High-level resistance to penicillin-type (β-lactam) antibiotics significantly limits the therapeutic options for patients with MRSA infections necessitating the use of newer agents, for which reduced susceptibility has already been described. Here we report for the first time that the central metabolism pentose phosphate pathway controls MRSA resistance to penicillin-type antibiotics. We comprehensively demonstrated that mutation of the PPP gene pgl perturbed metabolism in MRSA leading to increased flux to cell envelope precursors to drive increased antibiotic resistance. Moreover, increased resistance was associated with reduced levels of the lipoteichoic acids in the cell envelope, reduced rates of cell lysis under β-lactam stress and was dependent on the VraRG/GraRS multienzyme membrane complex that controls d-alanylation of teichoic acids and cell surface charge. Our data provide new insights on MRSA mechanisms of β-lactam resistance, which will support efforts to expand the treatment options for infections caused by this and other antimicrobial resistant pathogens.
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琥珀酰辅酶A的积累使甲氧西林抗葡萄球菌金黄色葡萄球菌（MRSA）琥珀酰基琥珀酰基琥珀酰基琥珀酰基琥珀酰基琥珀酸酯菌，与β-内酰胺抗生素的易感性增加有关。
影响因子：
6.4
作者：
Campbell C;Fingleton C;Zeden MS;Bueno E;Gallagher LA;Shinde D;Ahn J;Olson HM;Fillmore TL;Adkins JN;Razvi F;Bayles KW;Fey PD;Thomas VC;Cava F;Clair GC;O'Gara JP
通讯作者：
O'Gara JP
DOI：
10.1007/978-1-4939-3676-2_2
发表时间：
2016-01-01
期刊：
BACTERIAL CELL WALL HOMEOSTASIS: METHODS AND PROTOCOLS
影响因子：
--
作者：
Alvarez, Laura;Hernandez, Sara B.;Cava, Felipe
通讯作者：
Cava, Felipe
影响因子：
6.7
作者：
Cho J;Costa SK;Wierzbicki RM;Rigby WFC;Cheung AL
通讯作者：
Cheung AL
DOI：
10.1073/pnas.1209126109
发表时间：
2012-11-13
影响因子：
11.1
作者：
Brown, Stephanie;Xia, Guoqing;Walker, Suzanne
通讯作者：
Walker, Suzanne
影响因子：
3.4
作者：
BARBER, M
通讯作者：
BARBER, M