Obando Manuela Alvarado, Dörr Tobias
Department of Microbiology, Cornell University, Ithaca, NY.
Weill Institute for Cell and Molecular Biology, Cornell University, Ithaca, NY.
bioRxiv. 2023 Jul 12:2023.07.12.548665. doi: 10.1101/2023.07.12.548665.
Peptidoglycan (PG) is the main component of the bacterial cell wall; it maintains cell shape while protecting the cell from internal osmotic pressure and external environmental challenges. PG synthesis is essential for bacterial growth and survival, and a series of PG modifications are required to allow expansion of the sacculus. Endopeptidases (EPs), for example, cleave the crosslinks between adjacent PG strands to allow the incorporation of newly synthesized PG. EPs are collectively essential for bacterial growth and must likely be carefully regulated to prevent sacculus degradation and cell death. However, EP regulation mechanisms are poorly understood. Here, we used TnSeq to uncover novel EP regulation factors in . This screen revealed that the carboxypeptidase DacA1 (PBP5) alleviates EP toxicity. is essential for viability on LB medium, and this essentiality was suppressed by EP overexpression, revealing that EP toxicity both mitigates, and is mitigated by, a defect in . A subsequent suppressor screen to restore viability of Δ in LB medium was answered by hypomorphic mutants in the PG synthesis pathway, as well as mutations that promote PG degradation. Our data thus reveal a key role of DacA1 in maintaining the balance between PG synthesis and degradation.
肽聚糖（PG）是细菌细胞壁的主要成分；它维持细胞形状，同时保护细胞免受内部渗透压和外部环境挑战。PG合成对于细菌生长和存活至关重要，并且需要一系列PG修饰来允许细胞壁的扩展。例如，内肽酶（EPs）切割相邻PG链之间的交联以允许新合成的PG掺入。EPs对于细菌生长总体上是必不可少的，并且很可能必须受到严格调控以防止细胞壁降解和细胞死亡。然而，EPs的调控机制尚不清楚。在这里，我们使用TnSeq来揭示[具体物种]中的新型EP调控因子。该筛选表明羧肽酶DacA1（PBP5）减轻了EP毒性。[具体物种]对于在LB培养基上的生存能力至关重要，并且这种必需性被EP过表达所抑制，这表明EP毒性既减轻了[具体物种]中的缺陷，又被该缺陷所减轻。随后在LB培养基中恢复Δ[具体物种]生存能力的抑制子筛选得到了PG合成途径中的次等位基因突变体以及促进PG降解的突变体的响应。因此，我们的数据揭示了DacA1在维持PG合成与降解之间平衡中的关键作用。
