Role of Low-Molecular-Mass Penicillin-Binding Proteins, NagZ and AmpR in AmpC β-lactamase Regulation of Yersinia enterocolitica.

Chang Liu,Ran Duan,Jing Zhang,Zhaoke Guo,Xin Wang,Yuhuang Chen,Zhongzhi Zhao,Junrong Liang,Shihe Shao,Huaiqi Jing,Chuchu Li,Huijing Hao

doi:10.3389/fcimb.2017.00425

Abstract

Yersinia enterocolitica encodes a chromosomal AmpC β-lactamase under the regulation of the classical ampR-ampC system. To obtain a further understanding to the role of low-molecular-mass penicillin-binding proteins (LMM PBPs) including PBP4, PBP5, PBP6, and PBP7, as well as NagZ and AmpR in ampC regulation of Y. enterocolitica, series of single/multiple mutant strains were systematically constructed and the ampC expression levels were determined by luxCDABE reporter system, reverse transcription-PCR (RT-PCR) and β-lactamase activity test. Sequential deletion of PBP5 and other LMM PBPs result in a continuously growing of ampC expression level, the β-lactamse activity of quadruple deletion strain YEΔ4Δ5Δ6Δ7 (pbp4, pbp5, pbp6, and pbp7 inactivated) is approached to the YEΔD123 (ampD1, ampD2, and ampD3 inactivated). Deletion of nagZ gene caused two completely different results in YEΔD123 and YEΔ4Δ5Δ6Δ7, NagZ is indispensable for YEΔ4Δ5Δ6Δ7 ampC derepression phenotype but dispensable for YEΔD123. AmpR is essential for ampC hyperproduction in these two types of strains, inactivation of AmpR notable reduced the ampC expression level in both YEΔD123 and YEΔ4Δ5Δ6Δ7.

Highlights

Yersinia enterocolitica, a member of Enterobacteriaceae, is a zoonotic pathogen widely distributed in nature (Wang et al, 2011; Liang et al, 2012)
In the group of double and triple mutant strains, ampC derepression only appeared in pbp5 background, the ampC promoter activity of YE 4 5, YE 5 6, and YE 5 7 exhibited a marked rise compared with YE 4 6, YE 4 7, or YE 6 7
The quadruple deletion strain YE 4 5 6 7 displayed the highest level of ampC promoter activity. These results suggested that PBP5 plays the most important roles in Y. enterocolitica ampC regulation

Summary

Introduction

A member of Enterobacteriaceae, is a zoonotic pathogen widely distributed in nature (Wang et al, 2011; Liang et al, 2012). The process of ampC (blaB) regulation is tightly linked to the peptidoglycan recycling and controlled by AmpG, AmpD, AmpR, and NagZ (Vollmer et al, 2008; Zeng and Lin, 2013). Peptidoglycan degradation products including GlcNAc-1,6-anhydromuropeptide is transported into the cytoplasm by AmpG and further hydrolyzedcosaminidase) to yielding 1,6-anhydromuropeptides, which is the AmpR activator ligand for ampC derepression (Zamorano et al, 2010; Huang et al, 2012; Yang et al, 2014). AmpC Regulation of Yersinia enterocolitica the stem peptides of GlcNAc-1,6-anhydromuropeptide and 1,6-anhydromuropeptides can be removed by AmpD (N-acetylmuramyl-L-alanine amidase) and eventually recycled into UDP-MurNAc-pentapeptide, which is the AmpR repressor ligand to repress ampC expression level (Juan et al, 2006; Balasubramanian et al, 2015; Liu et al, 2016). Recent studies have found that in P. aeruginosa, PBP4 (DacB), PBP5 (DacC), and PBP7 (PbpG) are involved in ampC regulation, and PBP4 is the major cause of ampC derepressed in clinical strains (Moya et al, 2009; Ropy et al, 2015)

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