Interplay among Membrane-Bound Lytic Transglycosylase D1, the CreBC Two-Component Regulatory System, the AmpNG-AmpD _I -NagZ-AmpR Regulatory Circuit, and L1/L2 β-Lactamase Expression in Stenotrophomonas maltophilia

Abstract

e Lytic transglycosylases (LTs) are an important class of enzymes involved in peptidoglycan (PG) cleavage, with the concomitant formation of an intramolecular 1,6-anhydromuramoyl reaction product. There are six annotated LT genes in the Stenotrophomonas maltophilia genome, including genes for five membrane-bound LTs (mltA, mltB1, mltB2, mltD1, and mltD2) and a gene for soluble LT (slt). Six LTs of S. maltophilia KJ were systematically mutated, yielding the ⌬mltA, ⌬mltB1, ⌬mltB2, ⌬mltD1, ⌬mltD2, and ⌬slt mutants. Inactivation of mltD1 conferred a phenotype of elevated uninduced ␤-lactamase activity. The underlying mechanism responsible for this phenotype was elucidated by the construction of several mutants and determination of ␤-lactamase activity. The expression of the genes assayed was assessed by quantitative reverse transcriptase PCR and a promoter transcription fusion assay. The results demonstrate that ⌬mltD1 mutant-mediated L1/L2 ␤-lactamase expression involved the creBC two-component regulatory system (TCS) and the ampNG-ampD I -nagZ-ampR regulatory circuit. The inactivation of mltD1 resulted in mltB1 and mltD2 upexpression in a creBC-and ampNG-dependent manner. The overexpressed MltB1 and MltD2 activity contributed to the expression of the L1/L2 ␤-lactamase genes via the ampNG-ampD I -nagZ-ampR regulatory circuit. These findings reveal, for the first time, a linkage between LTs, the CreBC TCS, the ampNG-ampD I -nagZ-ampR regulatory circuit, and L1/L2 ␤-lactamase expression in S. maltophilia.

Impacts of Penicillin Binding Protein 2 Inactivation on β-Lactamase Expression and Muropeptide Profile in Stenotrophomonas maltophilia

“…The CreBC TCS has been shown to be responsible for strain ΔdacB -mediated β-lactam resistance increases in P. aeruginosa ( 29 ), srtrain ΔdacB -mediated β-lactamase increases in A. hydrophilia ( 12 ), and strain Δ mltD1 -mediated L1/L2 expression increases in S. maltophilia ( 30 ). To elucidate whether the mutant ΔmrdA -mediated basal L1/L2 activity increase is related to the creBC TCS, the ΔcreBC allele was introduced into strain KJΔ mrdA , yielding the mutant KJΔ mrdA Δ BC , and its basal β-lactamase activity was determined.…”

Lytic transglycosylases: concinnity in concision of the bacterial cell wall

“…Individual deletion of MltB, MltF, Slt, and SltB1 mutant strains had no significant effect on either basal or cefoxitin-induced AmpC expression compared to the wild-type strain (Cavallari et al , 2013). Similarly, LT-gene knockouts in Stenotrophomonas maltophilia (including Slt, MltA, MltB1, MltB2, MltD1, and MltD2) explored the effects of loss of LT function in this species (Huang et al , 2015; Wu et al , 2016). Removal of one LT (MltD1) yielded high levels of uninduced β-lactamase activity, an undesirable phenotype.…”

Expression and Functions of CreD, an Inner Membrane Protein in Stenotrophomonas maltophilia

“…aeruginosa . Nevertheless, we recently observed that the creD transcript has a 3.83 ± 0.33-fold increase when creBC is inactivated [ 14 ], which suggests that the regulatory circuit of creD expression in S . maltophilia is distinct and perhaps more complicated than that in E .…”