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<p>The insect midgut is crucial for digestion and nutrient absorption, but its embryonic development is poorly understood. Here, we show that the RNA-binding protein Squid is essential for embryonic midgut epithelium development in the model insect, <i>Bombyx mori</i>. CRISPR/Cas9-mediated knockout of <i>Squid</i> causes embryonic lethality, with mutants exhibiting disorganized midgut epithelium, lipid droplet accumulation, and impaired nutrient absorption. Integrated RNA sequencing (RNA-seq) and RNA-immunoprecipitation sequencing (RIP-seq) analyses reveal that Squid directly regulates the alternative splicing of <i>Axin</i>, a key Wnt/β-catenin pathway component. In <i>Squid</i>-depleted embryos, <i>Axin</i> splicing shifts from the long (<i>Axin-L</i>) to a short isoform (<i>Axin-S</i>), as confirmed by RT-qPCR. Consequently, β-catenin protein levels are significantly reduced in the midgut epithelium. Overexpression in <i>Bm</i>N cells confirms that Axin-L, but not Axin-S, elevates β-catenin. Overall, this study uncovers a critical post-transcriptional pathway wherein Squid ensures proper midgut development by regulating <i>Axin</i> alternative splicing to fine-tune β-catenin levels.</p>

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The RNA-binding protein Squid regulates embryonic midgut development via Axin alternative splicing in Bombyx mori

  • 期刊论文

  • 出版模式：

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  • 发表日期：

    2026年2月10日

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  • Chunmei Tong,
  • Wanyu Mo,
  • Minling Cai,
  • Yuling Peng,
  • Jilei Huang,
  • Kang Li,
  • Huimin Deng

摘要

The insect midgut is crucial for digestion and nutrient absorption, but its embryonic development is poorly understood. Here, we show that the RNA-binding protein Squid is essential for embryonic midgut epithelium development in the model insect, Bombyx mori. CRISPR/Cas9-mediated knockout of Squid causes embryonic lethality, with mutants exhibiting disorganized midgut epithelium, lipid droplet accumulation, and impaired nutrient absorption. Integrated RNA sequencing (RNA-seq) and RNA-immunoprecipitation sequencing (RIP-seq) analyses reveal that Squid directly regulates the alternative splicing of Axin, a key Wnt/β-catenin pathway component. In Squid-depleted embryos, Axin splicing shifts from the long (Axin-L) to a short isoform (Axin-S), as confirmed by RT-qPCR. Consequently, β-catenin protein levels are significantly reduced in the midgut epithelium. Overexpression in BmN cells confirms that Axin-L, but not Axin-S, elevates β-catenin. Overall, this study uncovers a critical post-transcriptional pathway wherein Squid ensures proper midgut development by regulating Axin alternative splicing to fine-tune β-catenin levels.