Gene regulatory network analysis of somatic embryogenesis identifies morphogenic genes that increase maize transformation frequency

Somatic embryogenesis allows a somatic plant cell to develop into an embryo, and potentially into a fertile plant. Transcription factors such as BABY BOOM can induce somatic embryogenesis when ectopically expressed and are widely used for aiding regeneration in tissue culture for transformation and gene editing of crops. Nevertheless, regeneration remains a bottleneck and alternative morphogenic genes are highly desired. Here, we co-expressed BABY BOOM and WUSCHEL2 in zygotic maize (Zea mays L.) embryos and studied gene regulatory networks in induced somatic embryos at the single-cell level. By inferring cell-type-specific regulons, we prioritized candidate regulators and confirmed functionality of four transcription factors, bHLH48, EREB152, GRF4, and HB77, for enhanced maize transformation frequency, leading to fertile, transgenic plants. Interestingly, the basic helix-loop-helix and homeodomain-leucine zipper families had previously not been associated with induced somatic embryogenesis. Our work will contribute to more efficient transformation, much needed to deliver on the promise of gene editing for agriculture. The provided files inlude: mini-ex__input__priors.zip: list of priors used to perform MINI-EX runs mini-ex__input__seurat.zip: scRNA-seq related inputs extracted from the Seurat object for the MINI-EX runs mini-ex__output.zip: outputs provided by MINI-EX for the eight priors体细胞胚胎发生（somatic embryogenesis）可使植物体细胞发育为胚胎，进而有望形成可育植株。诸如BABY BOOM的转录因子（transcription factor）在异位表达时可诱导体细胞胚胎发生，目前被广泛应用于作物转化与基因编辑的组织培养过程中，以辅助植株再生。然而，植株再生仍是作物遗传转化的一大瓶颈，因此亟需开发替代性的形态发生相关基因。本研究在玉米（Zea mays L.）合子胚中共表达BABY BOOM与WUSCHEL2，并在单细胞层面解析了诱导型体细胞胚胎中的基因调控网络。通过推断细胞类型特异性调控子，本研究筛选得到候选调控因子，并验证了bHLH48、EREB152、GRF4及HB77这四个转录因子可提升玉米转化频率，最终获得可育的转基因植株。值得注意的是，碱性螺旋-环-螺旋（basic helix-loop-helix, bHLH）与同源域-亮氨酸拉链（homeodomain-leucine zipper, HD-Zip）蛋白家族此前从未被报道与诱导型体细胞胚胎发生相关。本研究可为更高效的作物遗传转化提供技术支撑，而高效转化正是实现农业基因编辑应用前景的核心需求。 本次提供的文件包括： mini-ex__input__priors.zip：用于运行MINI-EX分析的先验因子列表 mini-ex__input__seurat.zip：从用于MINI-EX分析的Seurat对象中提取的单细胞RNA测序（single-cell RNA sequencing, scRNA-seq）相关输入数据 mini-ex__output.zip：MINI-EX针对8种先验因子生成的分析结果