Overexpression\nof l,d‑Transpeptidase\nA Induces Dispensability of Rod Complex in Escherichia\ncoli

Antimicrobial resistance (AMR) is a significant global\nthreat,\nand the presence of resistance-determinant genes is one of the major\ndriving forces behind it. The bacterial rod complex is an essential\nset of proteins that is crucial for cell survival due to its role\nin cell wall biogenesis and shape maintenance. Therefore, these proteins\noffer excellent potential as drug targets; however, compensatory mutations\nin nontarget genes render this complex nonessential. The MreB protein\nof this complex is an actin homologue that rotates along the longitudinal\naxis of the cell to provide rod shape to the bacteria. In this study,\nusing chemical–chemical interaction profiling and FtsZ suppression\nassay, we identified the MreB targeting activity of IITR07865, a previously\ndiscovered small molecule in our lab. Escherichia coli suppressors against IITR07865 revealed mutations in two cell division-associated\ngenes, <i>min C</i> and <i>pal</i>, that have\nnot been previously implicated in rod complex essentiality. IITR07865\nresistant mutants were found to inactivate and render the rod complex\nnonessential, making the rod complex inhibitors ineffective. Further,\nthrough transcriptome analysis, we reveal the primary cause of resistance\nin suppressor strains to be the overexpression of an l, d-transpeptidase A enzyme, which is involved in peptidoglycan\nand Braun’s lipoprotein cross-linking. Our results demonstrate\na novel mechanism of resistance development in rod-shaped Gram-negative\nbacterial pathogen E. coli involved\nin UTIs where mecillinam, a clinically used antibiotic that targets\nrod complex, is a drug of choice.