Comparative transcriptomic analysis among MRSA strains reveals a brominated carbazole disrupts translation and biosynthesis of cell walls, amino acids, and nucleotides to potentiate oxacillin.

Abstract

Methicillin-resistant Staphylococcus aureus (MRSA) is a priority pathogen that has acquired resistance to multiple antibiotic classes. One approach to minimizing this threat to human health is to identify and examine novel antibiotic adjuvants that potentiate these existing antibiotics in vitro. We have identified a chemical scaffold that potentiates multiple cell wall active antibiotics in MRSA and demonstrated that it disrupts the transcriptome of MRSA 43300 in a pleiotropic fashion. Treatment of cultures with this brominated carbazole (compound 8) in combination with oxacillin resulted in downregulation of genes involved in carbohydrate utilization, antibiotic resistance, and virulence. An interaction network constructed in this strain supported our previously proposed molecular target, the serine-threonine kinase, Stk1. Given the genetic diversity among both hospital- and community-associated strains of MRSA, we hypothesized that identifying the conserved transcriptome disruptions among multiple strains would provide biologically and clinically relevant information on compound 8’s mode of action as an antibiotic adjuvant. We also hypothesized that a standard meta-analysis of differentially expressed gene (DEG) lists that were common to all analyzed strains would provide predictive power for future studies and allow transposon mutants of interest such as Tn::stk1 to be interrogated for this master regulatory protein’s requirement in compound 8’s mode of action. Using weighted gene correlation network analysis, we examined four strains of MRSA that showed potentiation of oxacillin with compound 8 to reveal these conserved transcriptome disruption signatures. We uncovered widespread, conserved disruptions in bacterial translation, as well as cell wall, amino acid, and nucleotide synthesis. Finally, we validated a list of conserved DEGs in a fifth strain and showed that mutants for either stk1 or its cognate phosphatase stp1 still had compound 8 modulating these genes in a similar fashion. The exception was that vraD (an efflux pump) was no longer detectable when either stk1 or stp1 were disrupted. We concluded that compound 8 influences these conserved DEGs in all tested strains in an stk1- and stp1-independent fashion.

Affiliated Institutions

• High Point University US

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