Proteomic Changes of Klebsiella pneumoniae in Response to Colistin Treatment and crrB Mutation-Mediated Colistin Resistance

Abstract

Polymyxins are increasingly used as the critical last-resort therapeutic options for multidrug-resistant Gram-negative bacteria. Unfortunately, polymyxin resistance has increased gradually over the past few years. Although studies on polymyxin mechanisms are expanding, systemwide analyses of the underlying mechanism for polymyxin resistance and stress response are still lacking. To understand how Klebsiella pneumoniae adapts to colistin (polymyxin E) pressure, we carried out proteomic analysis of a K. pneumoniae strain cultured with different concentrations of colistin. Our results showed that the proteomic responses to colistin treatment in K. pneumoniae involve several pathways, including (i) gluconeogenesis and the tricarboxylic acid (TCA) cycle, (ii) arginine biosynthesis, (iii) porphyrin and chlorophyll metabolism, and (iv) enterobactin biosynthesis. Interestingly, decreased abundances of class A β-lactamases, including TEM, SHV-11, and SHV-4, were observed in cells treated with colistin. Moreover, we present comprehensive proteome atlases of paired polymyxin-susceptible and -resistant K. pneumoniae strains. The polymyxin-resistant strain Ci, a mutant of K. pneumoniae ATCC BAA 2146, showed a missense mutation in crrB. This crrB mutant, which displayed lipid A modification with 4-amino-4-deoxy-l-arabinose (l-Ara4N) and palmitoylation, showed striking increases in the expression of CrrAB, PmrAB, PhoPQ, ArnBCADT, and PagP. We hypothesize that crrB mutations induce elevated expression of the arnBCADTEF operon and pagP via PmrAB and PhoPQ. Moreover, the multidrug efflux pump KexD, which was induced by crrB mutation, also contributed to colistin resistance. Overall, our results demonstrated proteomic responses to colistin treatment and the mechanism of CrrB-mediated colistin resistance, which may offer valuable information on the management of polymyxin resistance.

The History of Colistin Resistance Mechanisms in Bacteria: Progress and Challenges

“…The authors identified a large amount of differentially expressed proteins that may contribute to mcr-1 -mediated antimicrobial resistance through regulation of glycerophospholipid metabolism, LPS biosynthesis and phosphoethanolamine substrate accumulation [ 94 ]. Additionally, to adapt to colistin stress, some proteins involved in the TCA cycle and pentose phosphate pathway were highly expressed in K. pneumoniae strains [ 97 ]. These recent advances in proteomics highlight significant metabolic pathways and suggest that the processes involved in the acquisition of the colistin response are related to energy, carbohydrate, and lipid metabolism [ 96 , 97 ].…”

“…Additionally, to adapt to colistin stress, some proteins involved in the TCA cycle and pentose phosphate pathway were highly expressed in K. pneumoniae strains [ 97 ]. These recent advances in proteomics highlight significant metabolic pathways and suggest that the processes involved in the acquisition of the colistin response are related to energy, carbohydrate, and lipid metabolism [ 96 , 97 ]. Thus, the inhibition of the metabolic process may potentially offer a way to overcome colistin resistance [ 97 ].…”

CrrB Positively Regulates High-Level Polymyxin Resistance and Virulence in Klebsiella pneumoniae

“…This suggests that crrB also induces the arnBCADTEF operon ( Cheng et al, 2016 , 2018 ; Jayol et al, 2017 ; Jeannot et al, 2017 ). Additional studies have taken proteomic and transcriptomic approaches to better understand PR and the role of crrB and have supported the observation that CrrA/B activates the arnBCADTEF operon ( Sun et al, 2020 ; Ramos et al, 2016 ). They also suggest that CrrA/B may alter the expression of several carbon metabolism pathways, including gluconeogenesis and the tricarboxylic acid (TCA) cycle.…”

A Molecular Perspective on Colistin and Klebsiella pneumoniae: Mode of Action, Resistance Genetics, and Phenotypic Susceptibility

“…The CrrC protein can activate the expression of both arnBCADTEF operon and pmrC gene via the PmrA/PmrB TCS (through an unknown mechanism) [ 40 , 41 ]. CrrB can also induce the expression of arnBCADTEF operon through phoP/phoQ but it is currently unknown if CrrC plays a role in this activation process [ 42 ] ( Figure 3 ).…”

“…The wild-type CrrB seems to repress kexD , but upon genetic alterations such as those, it is able to induce kexD through CrrA. However, crrC is simultaneously transcribed with kexD hampering a direct association between colistin resistance and kexD overexpression [ 42 , 79 ].…”