体细胞胚胎发生（SE）体系是研究组织和器官脱分化和再分化的良好模型，也是植物转基因技术的重要依托，对于其机制的研究具有重要的理论和应用价值。microRNAs（miRNAs）是一类长度通常为19-25个核苷酸（nt）的内源非编码RNA，在植物生长发育、激素信号转导、逆境胁迫响应等方面起着重要的作用。

近日，棉花团队在《Journal of Experimental Botany》发表题为“The GhmiR157a/GhSPL10 regulatory module controls initial cellular dedifferentiation and callus proliferation in cotton by modulating ethylene-mediated flavonoid biosynthesis”的研究论文。论文的通讯作者为杨细燕副教授。该研究基于棉花团队早期对高效体胚发生材料YZ1的小RNA和降解组测序中鉴定到的小RNA及其靶标基因GhmiR157a/GhSPL10 (Yang et al., 2013, Small RNA and degradome sequencing reveal complex miRNA regulation during cotton somatic embryogenesis, Journal of Experimental Botany, 64: 1521–1536,)，进一步探讨GhmiR157a/GhSPL10 在棉花体细胞胚胎发生中的功能。结果发现，超表达GhSPL10（GhmiR157a的靶基因）增加了生长素与乙烯含量及其相关信号通路基因的表达，激活了类黄酮生物合成途径，并促进棉花细胞初始脱分化和愈伤组织增殖。抑制GhSPL10过表达（35S:rSPL10-7）株系中类黄酮合成基因F3H的表达阻断了愈伤组织起始，而外源施几种黄酮醇可促进愈伤组织增殖，并促进细胞周期相关的基因表达。AVG处理35S:rSPL10-7株系抑制了乙烯合成，从而显著抑制愈伤组织的起始；而通过ACC处理、过表达EIN2、或通过RNAi抑制乙烯负调节因子CTR1，均可促进类黄酮相关基因表达和黄酮醇积累。以上结果表明，上调乙烯信号及激活GhSPL10过表达株系中类黄酮生物合成与细胞初始脱分化和愈伤组织增殖有关。该研究表明，GhmiR157a/GhSPL10模块通过激素和类黄酮途径调节棉花的体细胞胚胎发生。

全文链接：https://doi.org/10.1093/jxb/erx475

Authors: Lichen Wang¹, Nian Liu¹, Tianyi Wang¹, Jianying Li¹, Tianwang Wen¹, Xiyan Yang¹*, Keith Lindsey², Xianlong Zhang¹

Abstract

MicroRNAs (miRNAs) modulate many biological processes through inactivation of specific mRNA targets such as those encoding transcription factors (TFs). A delicate spatial/temporal balance between specific miRNA and target is central to achieving the appropriate biological outcomes. Somatic embryogenesis (SE) in cotton (Gossypium hirsutum), which goes through initial cellular dedifferentiation, callus proliferation and somatic embryo development, is of great importance for both fundamental research and for biotechnological applications. In this study, we characterize the function of the GhmiR157a/GhSPL10 miRNA-TF module during SE in cotton. We show that overexpression of GhSPL10, a target of GhmiR157a, increases free auxin and ethylene content and expression of associated signalling pathways, the activation of the flavonoid biosynthesis pathway, and promotes initial cellular dedifferentiation and callus proliferation. Inhibition of expression of the flavonoid synthesis gene F3H in GhSPL10 overexpression line (35S:rSPL10-7) blocked callus initiation, while exogenous application of several types of flavonols promoted callus proliferation, associated with cell cycle related gene expression. Inhibition of ethylene synthesis by AVG treatment in 35S:rSPL10-7 line severely inhibited callus initiation, while activation of ethylene signaling through ACC treatment, EIN2 overexpression, or inhibition of the ethylene negative regulator CTR1 by RNAi all promoted flavonoid-related gene expression and flavonol accumulation. These results show that an upregulation of ethylene signaling and the activation of flavonoid biosynthesis in GhSPL10 overexpression lines were associated with initial cellular dedifferentiation and callus proliferation. Our results demonstrate the importance of a GhmiR157a/GhSPL10 gene module in regulating SE via hormonal and flavonoid pathways.