Attenuator LRR - a regulatory tool for modulating gene expression in Gram-positive bacteria

Area of Science:

• Synthetic biology
• Molecular biology
• Bacterial genetics

Background:

• Riboswitches are crucial RNA devices for gene regulation in prokaryotic and eukaryotic cells.
• Existing riboswitches have limited performance and context-dependence, restricting their use in biotechnology.
• The need for robust and versatile RNA-based regulatory tools is critical for advancing synthetic biology.

Purpose of the Study:

• To evaluate the regulatory performance of a previously discovered ribosome-dependent ribo-regulator, LRR.
• To assess LRR's applicability across different bacterial species and genetic contexts.
• To demonstrate LRR's utility in controlling metabolic pathways for synthetic biology applications.

Main Methods:

• Characterization of LRR's gene expression regulation in the presence and absence of retapamulin.
• Testing LRR's functionality with various promoters in multiple Bacillus species and Escherichia coli.
• Application of LRR to modulate melanin metabolism in Bacillus thuringiensis BMB171.

Main Results:

• LRR demonstrated a 60-fold change in gene expression with retapamulin and low leaky expression (1-2%) without antibiotics.
• LRR exhibited robust performance across diverse Bacillus species (B. thuringiensis, B. cereus, B. amyloliquefaciens, B. subtilis) and in Escherichia coli.
• LRR successfully controlled melanin metabolism in Bacillus thuringiensis BMB171, showcasing its metabolic engineering potential.

Conclusions:

• LRR is a high-performance ribo-regulator with broad applicability in bacterial gene expression control.
• LRR facilitates the construction of genetic circuits and tuning of metabolic pathways.
• LRR holds significant potential for diverse applications within synthetic biology and biotechnology.