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A dual-mechanism antimicrobial peptide with antimutagenic activity targets the replisome and induces cell envelope stress

ABSTRACT: To combat the growing threat of multidrug-resistant bacteria, we need to develop novel antibiotics with unique modes of action. This study investigates the antibacterial properties of BTP-001 towards Escherichia coli. BTP-001 is a β-clamp targeting antimicrobial peptide containing an AlkB homologu 2 PCNA-interacting motif (APIM) linked to a cell penetrating peptide composed of 11 arginine residues (R11). Our data indicates that R11 mediates energy-dependent transport of BTP-001 across the cell membrane, possibly via iron transport systems such as the TonB-system. The full-length BTP-001 peptide rapidly affects the bacterial membrane, inducing increased expression and activation of the Cpx, E and Rcs cell envelope stress responses, which trigger the production of reaction oxygen species (ROS). This contributes to a rapid bactericidal effect as evidenced by increased short-term survival by addition of a ROS scavenger. Furthermore, we show that BTP-001 reduces the development of resistance to ciprofloxacin likely by binding to the β-clamp via APIM, thereby inhibiting translesion synthesis. In addition, our multi-omics data suggests that the -clamp is part of ribosomal complexes, and that regulation of translation is targeted by BTP-001. In conclusion, BTP-001 exhibits a multifaceted mode of action which strengthens its potential as a novel therapeutic drug against antibiotic resistant bacteria

ORGANISM(S): Escherichia coli

PROVIDER: GSE276254 | GEO | 2025/08/25

REPOSITORIES: GEO

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Project description:The bundle sheath cells (BSCs) layer – a presumed control point for radial transport of water and solutes between the vasculature and the leaf mesophyll cells (MCs) – is still largely understudied. Using isolated protoplasts, we found that 45% of the 90 genes differentially expressed in BSCs vs. MCs are membrane related and 20% are transport related, suggesting unique functionality of membrane transport in the BSCs, supported also by functional assays (electrophysiology and fluorescence imaging). A tight control of long distance transport is required for the optimization of the hydro-mineral homeostasis of organs in plants under fluctuating ambient conditions. The mechanism of ion selectivity and absorption from the xylem to the leaf is still poorly understood. The bundle sheath (BS), tightly enwrapping the leaf vasculature, has been suggested as a part of this mechanism, acting as a selective barrier which regulates the radial transport of water and solutes from the xylem to leaf cells. This suggestion relies on the anatomy, as well as on the recent physiological transport assays of the BS and its cells (BSCs). We hypothesized that the unique transport functionality of the BSCs is manifested in its transcriptome. To test this, we compared the transcriptomes of individually hand-picked protoplasts of GFP-labeled BSCs and non-labeled mesophyll cells (MCs) of Arabidopsis thaliana leaves. Indeed, in conformation of our hypothesis, 45% of the 90 genes differentially expressed in BSCs vs. MCs are membrane related and 20% are transport related, with the proton-pump AHA2 as a prominent example. Moreover, fluorescence imaging using the potentiometric dye (di-8 ANEPPS) revealed a more negative membrane potential of the BSCs protoplasts compared to those of MCs, and electrophysiological assays (patch-clamp) showed that the major, AKT2-like, membrane K+ conductances of BSCs and MCs had different voltage dependency ranges. Combined, these differences are compatible with an expected possibility of simultaneous but oppositely directed transmembrane K+ fluxes in BSCs and MCs in otherwise similar conditions.

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A dual-mechanism antimicrobial peptide with antimutagenic activity targets the replisome and induces cell envelope stress

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