Gundavarapu Bhargavi, Nallamotu Krishna Chaitanya, Murapaka Vishnu Vachana, Venkataraman Balaji, Saisree Lutikurti, Reddy Manjula
CSIR-Centre for Cellular and Molecular Biology, Hyderabad, India.
Academy of Scientific and Innovative Research (AcSIR), Ghaziabad, India.
PLoS Genet. 2025 May 22;21(5):e1011712. doi: 10.1371/journal.pgen.1011712. eCollection 2025 May.
Gram-negative bacterial cell envelope consists of a surface-exposed lipid bilayer (outer membrane or OM) that serves as a permeability barrier to maintain the cellular integrity. Beneath the OM is the periplasmic space that harbours peptidoglycan (PG), a highly cross-linked mesh-like glycan polymer closely encasing the inner membrane (IM). During growth of a bacterium balanced synthesis of the envelope components is required to maintain the cellular integrity, of which little is known. In this study, we identify sanA, an ORF of unknown function encoding a predicted IM-anchored protein as a factor contributing to balanced synthesis of PG in E. coli. Absence of SanA increased the rate of nascent PG strand incorporation, and restored growth and viability to several mutants defective in either cell division or cell elongation. Detailed mutant analysis of sanA showed that it is defective in the envelope barrier properties. Interestingly, overexpression of the periplasmic endopeptidases that cleave the cross-links of the PG mesh was able to alleviate the phenotypes of sanA mutant implying the envelope defects are due to alterations in the PG sacculus. Additionally, a SanA variant (SSDsbA-SanA) targeted to the periplasm, complemented the SanA- phenotypes suggesting it functions in the periplasmic phase of the PG synthesis. Further, we find that SanA functions independently of its paralog, ElyC, known to regulate the synthesis of enterobacterial common antigen (ECA), a surface polysaccharide found in the cell envelopes of most enteric bacteria. Overall, our results suggest a role for SanA in the maintenance of optimal PG synthesis, providing evidence for the existence of an additional layer of regulation in Gram-negative cell envelope biogenesis.
革兰氏阴性菌的细胞包膜由一个表面暴露的脂质双层（外膜或OM）组成，该双层作为一种渗透屏障以维持细胞的完整性。在外膜下方是周质空间，其中含有肽聚糖（PG），这是一种高度交联的网状聚糖聚合物，紧密包裹着内膜（IM）。在细菌生长过程中，需要包膜成分的平衡合成来维持细胞的完整性，但对此了解甚少。在本研究中，我们鉴定出sanA，一个功能未知的开放阅读框，其编码一种预测的内膜锚定蛋白，是有助于大肠杆菌中PG平衡合成的一个因子。SanA的缺失增加了新生PG链掺入的速率，并恢复了几个在细胞分裂或细胞伸长方面有缺陷的突变体的生长和活力。对sanA的详细突变分析表明它在包膜屏障特性方面存在缺陷。有趣的是，切割PG网交联的周质内肽酶的过表达能够减轻sanA突变体的表型，这意味着包膜缺陷是由于PG囊泡的改变所致。此外，一个靶向周质的SanA变体（SSDsbA-SanA）补充了SanA的表型，表明它在PG合成的周质阶段发挥作用。进一步地，我们发现SanA独立于其旁系同源物ElyC发挥作用，ElyC已知可调节肠杆菌共同抗原（ECA）的合成，ECA是大多数肠道细菌细胞包膜中发现的一种表面多糖。总体而言，我们的结果表明SanA在维持最佳PG合成中发挥作用，为革兰氏阴性菌细胞包膜生物合成中存在额外一层调控提供了证据。
