CmBt and CmBr synergistically regulate cucurbitacin B biosynthesis in melon

Area of Science:

• Plant Molecular Biology
• Plant Biochemistry
• Agricultural Science

Background:

• Cucurbitacin B (CuB) causes bitterness in melon (Cucumis melo L.), impacting fruit quality and consumer acceptance.
• The precise tissue-specific regulatory mechanisms of CuB biosynthesis by transcription factors are not fully understood.

Purpose of the Study:

• To elucidate the roles of the basic helix-loop-helix (bHLH) transcription factors CmBt and CmBr in CuB biosynthesis across different melon tissues.
• To investigate the interaction and synergistic effects of CmBt and CmBr on CuB accumulation and the activation of the CuB biosynthetic gene CmBi.

Main Methods:

• CRISPR-Cas9 gene editing was used to generate knockout lines for the CmBt gene.
• Analysis of CuB content in various tissues of single and double mutant lines (Cmbt, Cmbr, CmbtCmbr).
• Reverse transcription quantitative PCR (RT-qPCR) and dual-luciferase assays were employed to assess gene activation.

Main Results:

• Loss of CmBt reduced CuB in fruits and roots, while CmBr predominantly regulated CuB in fruits and roots, redefining their tissue-specific roles.
• CmBt and CmBr interact synergistically, leading to a more significant reduction in CuB content in double mutants across all tested tissues.
• CmBt and CmBr cooperatively activate the CuB biosynthetic gene CmBi and completely prevent CPPU-induced fruit bitterness in double mutants.

Conclusions:

• CmBt and CmBr exhibit tissue-specific, partially redundant, and synergistic functions in activating CmBi and promoting CuB biosynthesis.
• The CmBt-CmBr locus represents a key genetic target for developing non-bitter melon cultivars with improved fruit quality and consumer appeal.