Chemotranscriptomic Profiling Defines Drug-Specific Signatures of the Glycopeptide Antibiotics Dalbavancin, Vancomycin and Chlorobiphenyl-Vancomycin in a VanB-Type-Resistant Streptomycete

Abstract

Dalbavancin, vancomycin and chlorobiphenyl-vancomycin share a high degree of structural similarity and the same primary mode of drug action. All inhibit bacterial cell wall biosynthesis through complexation with intermediates in peptidoglycan biosynthesis mediated via interaction with peptidyl-d-alanyl-d-alanine (d-Ala-d-Ala) residues present at the termini of the intermediates. VanB-type glycopeptide resistance in bacteria encodes an inducible reprogramming of bacterial cell wall biosynthesis that generates precursors terminating with d-alanyl-d-lactate (d-Ala-d-Lac). This system in Streptomyces coelicolor confers protection against the natural product vancomycin but not dalbavancin or chlorobiphenyl-vancomycin, which are semi-synthetic derivatives and fail to sufficiently activate the inducible VanB-type sensory response. We used transcriptome profiling by RNAseq to identify the gene expression signatures elucidated in S. coelicolor in response to the three different glycopeptide compounds. An integrated comparison of the results defines both the contribution of the VanB resistance system to the control of changes in gene transcription and the impact at the transcriptional level of the structural diversity present in the glycopeptide antibiotics used. Dalbavancin induces markedly more extensive changes in the expression of genes required for transport processes, RNA methylation, haem biosynthesis and the biosynthesis of the amino acids arginine and glutamine. Chlorobiphenyl-vancomycin exhibits specific effects on tryptophan and calcium-dependent antibiotic biosynthesis and has a stronger repressive effect on translation. Vancomycin predictably has a uniquely strong effect on the genes controlled by the VanB resistance system and also impacts metal ion homeostasis and leucine biosynthesis. Leaderless gene transcription is disfavoured in the core transcriptional up- and down-regulation taking place in response to all the glycopeptide antibiotics, while HrdB-dependent transcripts are favoured in the down-regulated group. This study illustrates the biological impact of peripheral changes to glycopeptide antibiotic structure and could inform the design of future semi-synthetic glycopeptide derivatives.

Keywords: Streptomyces; antibiotic; chemotranscriptomics; dalbavancin; glycopeptide; resistance; vancomycin.

FIGURE 1

The glycopeptide antibiotics used in this study. Structures of vancomycin (Van), chlorobiphenyl-vancomycin (CbpVan) and dalbavancin (Dal), and their corresponding minimum inhibitory concentration (MIC) values against S. coelicolor in liquid cultures (Kwun and Hong, 2014) (A) and pairwise Tanimoto chemical similarity scores (B). In (A), distinctive structural features are highlighted, including the following that are also discussed in the text: N-methylleucine group (green); chlorobiphenyl group (blue); lipid side chain (orange); dimethylaminopropyl amide side chain (red).

FIGURE 2

The transcriptional responses of S. coelicolor cells to exposure to vancomycin (Van), chlorobiphenyl-vancomycin (CbpVan) and dalbavancin (Dal). Principal components analysis (PCA) (A) and hierarchical clustering of Euclidean sample distances (B) indicate closer similarity between the data for Van and CbpVan compared to Dal. Differential expression analysis identifies gene transcripts significantly changing in abundance in response to one or more glycopeptide antibiotic compared to the control (Ctrl) (C). In (B), darker blue grid colours indicate closer similarity between pairs of samples.

FIGURE 3

Differences in activation of the VanB-type resistance system (A), the SigE-controlled cell wall stress response (B) and zinc starvation stress response (C). In each panel, the diagonal line illustrates theoretical perfect correlation between the data on each x- and y-axis (slope = 1). In (A), global correlations between the changes induced by each antibiotic treatment are shown (R² and p-values).

FIGURE 4

Heatmap summarising the transcripts uniquely changing significantly in response to only one of the glycopeptide antibiotics used, relative to the control treatment. Significant biological process categories from gene ontology (GO) functional enrichment analysis of each group of genes are shown to the right (“ns” denotes cases where no significant categories were identified). Complete GO enrichment analysis results are provided in Supplementary Data 2.

FIGURE 5

Heatmap summarising the transcripts changing significantly in response to two or more of the glycopeptide antibiotics used relative to the control treatment. Significant biological process categories from gene ontology (GO) functional enrichment analysis of each group of genes are shown to the right (“ns” denotes cases where no significant categories were identified). Only groups containing more than 10 transcripts are labelled. Complete GO enrichment analysis results are provided in Supplementary Data 2.

FIGURE 6

sRNAs responding significantly to glycopeptide antibiotic stress.