BmY1314调控家蚕后部丝腺大小发育的分子机制研究

Study on the regulatory mechanism of organ size during development has great significance for revealing the mechanism of tumors and guiding the improvement of plant performance and animal production. The silk gland of the lepidopteran model insect, the silkworm Bombyx mori, is the organ that produces silk and has been used to explore regulation of organ size and gene expression for a long time. However, the regulatory mechanism of this organ has still not been fully elucidated. We have previously found that BmY1314 (the major downstream effector of the Hippo pathway) is closely involved in the size regulation of B. mori posterior silk glands. In this project, the BmY1314-overexpressing and BmY1314-interfering transgenic silkworms will be used as experimental materials. By using methods of electron microscope, RNA-Seq, ChIP-Seq, DNA pull-down and EMSA etc., this project will further investigate the phenotypic characteristics of posterior silk glands of both the transgenic silkworms, transcription factors induced by BmY1314 and differentially expressed in the posterior silk gland of transgenic silkworms, and interactions between BmY1314, transcription factors and silk fibroin genes, to clarify the role and molecular mechanism of BmY1314 in regulating the size of B. mori posterior silk glands. This study is expected to reveal a novel mechanism for the size regulation of B. mori posterior silk glands, which will not only provides new view for the study of silk gland development and regulation, but also has the guiding significance for the modification of silk glands and development of silk-based materials.

器官大小调控研究对揭示肿瘤发生机理、指导动植物生产性能改造等具有重要意义。丝腺是鳞翅目模式昆虫家蚕的产丝器官，很早就被用作器官大小发育及基因表达调控的研究材料，但其调控机制至今仍未完全阐明。本项目将在前期确证BmY1314（Hippo通路下游核心成员）与家蚕后部丝腺大小发育密切相关的基础上，以转基因过表达和转基因干涉BmY1314的家蚕为材料，采用电镜观察、RNA-Seq、ChIP-Seq、DNA pull-down和EMSA等方法，通过分析转基因家蚕及其后部丝腺的表型特征、BmY1314诱导并在转基因家蚕后部丝腺中显著差异表达的转录因子，以及BmY1314-转录因子-丝素基因的相互作用关系等，探明BmY1314调控家蚕后部丝腺大小发育的作用及分子机制。本研究将揭示一种家蚕丝腺大小发育调控的新机制，不仅为丝腺发育调控研究提供新的视点，同时对丝腺定向改造和蚕丝素材研发等也具有指导意义。

家蚕以产丝著称，是蚕丝业的生物学基础。丝腺是家蚕合成分泌蚕丝蛋白的专一器官，对蚕丝产量与品质起着决定性作用。丝腺发育调控机制是蚕学领域的重大科学问题，但具体机制和关键调控因子远未探明。本项目以Hippo通路核心成员BmY1314为靶标，解析其调控家蚕后部丝腺发育的具体功能和作用机理。取得的主要结果有：1）通过制作和分析转基因过表达和敲除BmY1314的家蚕丝腺突变表型，证实了BmY1314是调控家蚕丝腺发育的关键因子；2）通过丝腺RNA-seq等分析，鉴定了丝腺中受BmY1314调控的转录因子等下游靶标；3）通过丝腺scRNA-seq等分析，揭示了丝腺细胞的异质性和胞内基因转录表达动态，鉴定出Hippo、20E等调控丝腺发育的重要信号转导通路和调控因子；4）通过互作等实验，证实了BmSd是BmY1314的主要协同因子，BmY1314与BmSd既能以直接结合丝素基因启动子的方式调控丝蛋白合成，也能通过结合丝腺转录因子的方式间接调控丝蛋白合成。本项目揭示了BmY1314调控家蚕丝腺发育的分子机制，为丝腺发育调控研究提供了新视点；鉴定了调控丝腺发育的靶标基因，获得了多个丝腺突变体素材，可贡献于蚕丝分子改良和丝蛋白开发利用。项目成果已发表于Nature Communications、Scientific Reports、Transgenic Research、Biochemical and Biophysical Research Communications等国际学术期刊，申请蚕丝改良技术和素材专利3项。