Related Experiment Video
Updated: Mar 14, 2026
Measurement of Specific Mycobacterial Mistranslation Rates with Gain-of-function Reporter Systems
Published on: April 26, 2019
The Expression of Antibiotic Resistance Methyltransferase Correlates with mRNA Stability Independently of Ribosome
1Edward A. Doisy Department of Biochemistry and Molecular Biology, Saint Louis University School of Medicine, St. Louis, Missouri, USA.
Abstract:
Members of the Erm methyltransferase family modify 23S rRNA of the bacterial ribosome and render cross-resistance to macrolides and multiple distantly related antibiotics. Previous studies have shown that the expression of erm is activated when a macrolide-bound ribosome stalls the translation of the leader peptide preceding the cotranscribed erm Ribosome stalling is thought to destabilize the inhibitory stem-loop mRNA structure and exposes the erm Shine-Dalgarno (SD) sequence for translational initiation. Paradoxically, mutations that abolish ribosome stalling are routinely found in hyper-resistant clinical isolates; however, the significance of the stalling-dead leader sequence is largely unknown. Here, we show that nonsense mutations in the Staphylococcus aureus ErmB leader peptide (ErmBL) lead to high basal and induced expression of downstream ErmB in the absence or presence of macrolide concomitantly with elevated ribosome methylation and resistance. The overexpression of ErmB is associated with the reduced turnover of the ermBL-ermB transcript, and the macrolide appears to mitigate mRNA cleavage at a site immediately downstream of the ermBL SD sequence. The stabilizing effect of antibiotics on mRNA is not limited to ermBL-ermB; cationic antibiotics representing a ribosome-stalling inducer and a noninducer increase the half-life of specific transcripts. These data unveil a new layer of ermB regulation and imply that ErmBL translation or ribosome stalling serves as a "tuner" to suppress aberrant production of ErmB because methylated ribosome may impose a fitness cost on the bacterium as a result of misregulated translation.
Insights
Bacterial antibiotic resistance gene ermB expression is regulated by ribosome stalling in its leader peptide. This study reveals that mutations preventing stalling increase ermB expression and antibiotic resistance, suggesting a novel regulatory mechanism.
Area of Science:
- Bacterial genetics and molecular biology
- Antibiotic resistance mechanisms
- Ribosome function and regulation
Background:
- Erm methyltransferases confer resistance to macrolides and other antibiotics by modifying bacterial 23S rRNA.
- Erm gene expression is typically induced by macrolide-bound ribosomes stalling during leader peptide translation, which exposes the Shine-Dalgarno sequence.
- The role of mutations that abolish ribosome stalling in clinical isolates remains unclear.
Purpose of the Study:
- To investigate the regulatory significance of ribosome stalling in the leader peptide of the Staphylococcus aureus ErmB methyltransferase.
- To determine the impact of mutations in the ErmB leader peptide on ErmB expression and antibiotic resistance.
- To explore the influence of antibiotics on the stability of ermBL-ermB transcripts.
Main Methods:
- Analysis of nonsense mutations in the Staphylococcus aureus ErmB leader peptide (ErmBL).
- Measurement of ErmB expression levels in the presence and absence of macrolides.
- Assessment of ribosome methylation levels and antibiotic resistance.
- Transcriptional stability assays for ermBL-ermB and other specific transcripts.
- Investigation of antibiotic effects on mRNA cleavage sites.
Main Results:
- Nonsense mutations in ErmBL result in high basal and induced ErmB expression, increased ribosome methylation, and elevated antibiotic resistance, irrespective of macrolide presence.
- Overexpression of ErmB correlates with reduced turnover of the ermBL-ermB transcript.
- Macrolides appear to inhibit mRNA cleavage downstream of the ermBL Shine-Dalgarno sequence, stabilizing the transcript.
- The antibiotic-mediated mRNA stabilization effect extends to other transcripts, including those affected by cationic antibiotics.
Conclusions:
- Ribosome stalling during ErmBL translation acts as a crucial negative regulator, preventing excessive ErmB production.
- Mutations that eliminate stalling lead to uncontrolled ErmB expression and heightened antibiotic resistance.
- Antibiotics can stabilize ermB transcripts, representing a previously unrecognized layer of gene regulation.
- Aberrant ErmB production due to loss of stalling may impose a fitness cost on bacteria through misregulated translation.
Related Concept Videos
02:51mRNA Stability and Gene Expression
02:51mRNA Stability and Gene Expression
Cis-acting Elements involved in mRNA stability
01:53RNA Stability
01:53RNA Stability
01:29Translational Regulation
01:23Types of RNA
Three main types of RNA are involved in protein synthesis: messenger RNA (mRNA), transfer RNA (tRNA), and ribosomal RNA (rRNA). These RNAs perform diverse functions and can be broadly classified as protein-coding or non-coding RNA. Non-coding RNAs play important roles in the regulation of gene expression in response to developmental and environmental changes. Non-coding RNAs in prokaryotes can be manipulated to develop more effective antibacterial drugs for human or animal use.
RNA...