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Key message <p>Tissue-specific <i>RDR6</i> compensation rescues Arabidopsis defects while maintaining seed polyunsaturated fatty acid accumulation, balancing co-suppression relief and key trait retention for modular engineering.</p> Abstract <p>The transgene-induced co-suppression of fatty acid desaturase 2 (FAD2) can be effectively released in <i>rdr6</i> mutant, enabling a significant increase in polyunsaturated fatty acid (PUFA) content in seeds. However, the global suppression of RNA-dependent RNA polymerase 6 (RDR6) compromises plant growth and disease resistance. To address this limitation, we developed a spatiotemporal compensation strategy by restoring <i>RDR6</i> expression in non-seed tissues using tissue-specific promoters while maintaining its low expression during seed maturation. To implement this goal, we identified P<sub>BnTC06</sub>, a <i>Brassica napus</i> promoter, through transcriptomic data mining and functional characterization. GUS staining revealed that the P<sub>BnTC06</sub> promoter drives strong gene expression in vegetative tissues (e.g., leaves, stems, and flowers) but exhibits negligible activity in mid- to late-stage-developing seeds. We introduced P<sub>BnTC06</sub>::<i>RDR6</i> into Pha::<i>AtFAD2/rdr6-11</i>, the previously established high- PUFA Arabidopsis line. This intervention rescued the <i>rdr6</i> mutant phenotype (characterized by gracile, downward-curling leaves) to wild-type morphology and restored <i>RDR6</i> expression across non-seed tissues, while maintaining minimal expression in middle and late developing seeds. Crucially, <i>FAD2</i> transcript levels remained at a high level during late seed development, resulting in sustained high PUFA accumulation in mature seeds. This strategy establishes a practical strategy to circumvent transgene co-suppression and proposes a modular framework for precision breeding of complex traits in crops.</p>

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A method for maintaining the release of co-suppression and maximally restoring the RDR6 expression

  • 期刊论文

  • 发表日期：

    2025年5月9日

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  • Na Li,
  • Hailan Wang,
  • Jiayin Wen,
  • Xiangling Liu,
  • Meng Zhang

摘要

Key message

Tissue-specific RDR6 compensation rescues Arabidopsis defects while maintaining seed polyunsaturated fatty acid accumulation, balancing co-suppression relief and key trait retention for modular engineering.

Abstract

The transgene-induced co-suppression of fatty acid desaturase 2 (FAD2) can be effectively released in rdr6 mutant, enabling a significant increase in polyunsaturated fatty acid (PUFA) content in seeds. However, the global suppression of RNA-dependent RNA polymerase 6 (RDR6) compromises plant growth and disease resistance. To address this limitation, we developed a spatiotemporal compensation strategy by restoring RDR6 expression in non-seed tissues using tissue-specific promoters while maintaining its low expression during seed maturation. To implement this goal, we identified PBnTC06, a Brassica napus promoter, through transcriptomic data mining and functional characterization. GUS staining revealed that the PBnTC06 promoter drives strong gene expression in vegetative tissues (e.g., leaves, stems, and flowers) but exhibits negligible activity in mid- to late-stage-developing seeds. We introduced PBnTC06::RDR6 into Pha::AtFAD2/rdr6-11, the previously established high- PUFA Arabidopsis line. This intervention rescued the rdr6 mutant phenotype (characterized by gracile, downward-curling leaves) to wild-type morphology and restored RDR6 expression across non-seed tissues, while maintaining minimal expression in middle and late developing seeds. Crucially, FAD2 transcript levels remained at a high level during late seed development, resulting in sustained high PUFA accumulation in mature seeds. This strategy establishes a practical strategy to circumvent transgene co-suppression and proposes a modular framework for precision breeding of complex traits in crops.