Paradigm shift for cry gene expression in Bacillus thuringiensis

Area of Science:

• Microbiology
• Molecular Biology
• Biochemistry

Background:

• Bacillus thuringiensis (Bt) produces insecticidal Cry and Vip toxins, crucial for biopesticides.
• Toxin gene expression is typically linked to sporulation via sigma factors E or K.
• The VipR regulator's role in stationary phase expression of Vip3A is known.

Purpose of the Study:

• To investigate the broader role of the VipR regulator in Bt toxin gene expression.
• To identify new VipR-regulated genes and understand their contribution to insecticidal activity.
• To explore sporulation-independent pathways for toxin production in Bt.

Main Methods:

• Identification of VipR-binding sites using genomic analysis.
• Validation of VipR binding and transcriptional activation through transcription assays.
• Gene expression analysis in wild-type and mutant Bt strains (e.g., ∆spo0A).
• Introduction of VipR into strains lacking the regulator.

Main Results:

• VipR directly regulates cry2Aa, cry2Ab, and cry1Ia genes in Bt strain HD1.
• A VipR-binding site was found upstream of an ami-cry1Ac operon in strain HD73, leading to VipR-dependent Cry1Ac production.
• Sporulation-independent expression of Cry1Ac and Cry2Ab was confirmed in relevant mutants.
• Putative VipR-binding sites were identified upstream of other lepidopteran-active cry genes (cry1E, cry1F, cry9D, cry9E).

Conclusions:

• The VipR regulator controls a wider range of insecticidal genes than previously known, including cry1A and cry2A families.
• Sporulation-independent production of Cry toxins is a significant regulatory mechanism in Bt.
• These findings provide a basis for enhancing Bt biopesticide efficacy through genetic engineering.