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<p>N<sup>6</sup>-methyladenosine (m<sup>6</sup>A), the most prevalent epitranscriptomic modification in plant mRNAs, functions as a critical regulatory layer governing environmental response. Refrigeration reduces transcripts of cell wall enzymes to delay softening, however whether m<sup>6</sup>A participated in this progress remains unclear. Here, we identified a cold-inducible m<sup>6</sup>A reader PpYTHDFE1 as a regulator that stabilizes peach (<i>Prunus persica</i> L.) fruit firmness. Epitranscriptomic analysis revealed that firmness maintenance correlates with reduced levels of m<sup>6</sup>A-modified transcripts such as expansin <i>PpEXP3</i>, whose overexpressing accelerates fruit softening. Both in vitro RNA-EMSA and in vivo RIP-qPCR assays demonstrate that PpYTHDFE1 recognizes m<sup>6</sup>A-modified <i>PpEXP3</i> transcripts. Transcription inhibition assays further confirm that PpYTHDFE1 accelerates the degradation of <i>PpEXP3</i> through RNA–protein interaction rather than transcriptional modulation. Mechanistically, PpYTHDFE1 participates in liquid–liquid phase separation as evidenced by fluorescence recovery after photobleaching (FRAP) assays, facilitating <i>PpEXP3</i> degradation<i>.</i> Furthermore, overexpression of <i>PpYTHDFE1</i> decreases <i>PpEXP3</i> mRNA levels, while silencing <i>PpYTHDFE1</i> leads to <i>PpEXP3</i> transcript accumulation. A significantly negative correlation was observed between <i>PpYTHDFE1</i> transcripts and firmness during fruit cold storage. Fruit firmness is increased by 44% in peach and 45% in tomato upon <i>PpYTHDFE1</i> overexpressing. Our findings provide molecular insights into RNA modification-based strategies to reduce postharvest losses and develop cold-resilient horticultural crops.</p>

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Cold storage delays peach fruit softening via m6A reader PpYTHDFE1 liquid-liquid phase separation-mediated degradation of cell wall-loosening transcript PpEXP3

  • 期刊论文

  • 出版模式：

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  • 发表日期：

    2026年5月9日

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  • Hanqing Wang,
  • Hanxiao Bian,
  • Xubin Wu,
  • Kunsong Chen,
  • Donald Grierson,
  • Bo Zhang

摘要

N6-methyladenosine (m6A), the most prevalent epitranscriptomic modification in plant mRNAs, functions as a critical regulatory layer governing environmental response. Refrigeration reduces transcripts of cell wall enzymes to delay softening, however whether m6A participated in this progress remains unclear. Here, we identified a cold-inducible m6A reader PpYTHDFE1 as a regulator that stabilizes peach (Prunus persica L.) fruit firmness. Epitranscriptomic analysis revealed that firmness maintenance correlates with reduced levels of m6A-modified transcripts such as expansin PpEXP3, whose overexpressing accelerates fruit softening. Both in vitro RNA-EMSA and in vivo RIP-qPCR assays demonstrate that PpYTHDFE1 recognizes m6A-modified PpEXP3 transcripts. Transcription inhibition assays further confirm that PpYTHDFE1 accelerates the degradation of PpEXP3 through RNA–protein interaction rather than transcriptional modulation. Mechanistically, PpYTHDFE1 participates in liquid–liquid phase separation as evidenced by fluorescence recovery after photobleaching (FRAP) assays, facilitating PpEXP3 degradation. Furthermore, overexpression of PpYTHDFE1 decreases PpEXP3 mRNA levels, while silencing PpYTHDFE1 leads to PpEXP3 transcript accumulation. A significantly negative correlation was observed between PpYTHDFE1 transcripts and firmness during fruit cold storage. Fruit firmness is increased by 44% in peach and 45% in tomato upon PpYTHDFE1 overexpressing. Our findings provide molecular insights into RNA modification-based strategies to reduce postharvest losses and develop cold-resilient horticultural crops.