Data_Sheet_1_High-Resolution Translatome Analysis Reveals Cortical Cell Programs During Early Soybean Nodulation.xlsx

Nodule organogenesis in legumes is regulated temporally and spatially through gene networks. Genome-wide transcriptome, proteomic, and metabolomic analyses have been used previously to define the functional role of various plant genes in the nodulation process. However, while significant progress has been made, most of these studies have suffered from tissue dilution since only a few cells/root regions respond to rhizobial infection, with much of the root non-responsive. To partially overcome this issue, we adopted translating ribosome affinity purification (TRAP) to specifically monitor the response of the root cortex to rhizobial inoculation using a cortex-specific promoter. While previous studies have largely focused on the plant response within the root epidermis (e.g., root hairs) or within developing nodules, much less is known about the early responses within the root cortex, such as in relation to the development of the nodule primordium or growth of the infection thread. We focused on identifying genes specifically regulated during early nodule organogenesis using roots inoculated with Bradyrhizobium japonicum. A number of novel nodulation gene candidates were discovered, as well as soybean orthologs of nodulation genes previously reported in other legumes. The differential cortex expression of several genes was confirmed using a promoter-GUS analysis, and RNAi was used to investigate gene function. Notably, a number of differentially regulated genes involved in phytohormone signaling, including auxin, cytokinin, and gibberellic acid (GA), were also discovered, providing deep insight into phytohormone signaling during early nodule development.豆科植物根瘤器官发生（nodule organogenesis）通过基因网络在时间与空间维度受到精准调控。此前，研究人员已通过全基因组转录组、蛋白质组及代谢组分析，阐明了各类植物基因在结瘤进程中的功能作用。尽管该领域已取得显著进展，但多数此类研究均面临组织稀释的局限：仅少量细胞或根组织区域会响应根瘤菌侵染，大部分根系并无响应。为部分解决这一问题，我们采用翻译核糖体亲和纯化（translating ribosome affinity purification, TRAP）技术，借助皮层特异性启动子，特异性监测根皮层对根瘤菌接种的响应。既往研究多聚焦于根表皮（如根毛）或发育中的根瘤内的植物响应，而针对根皮层内的早期响应——例如与根瘤原基发育或侵染线生长相关的响应——的认知仍较为有限。本研究以大豆慢生根瘤菌（Bradyrhizobium japonicum）接种的根系为实验材料，旨在鉴定早期根瘤器官发生过程中特异性调控的基因。研究不仅发现了多个全新的结瘤基因候选靶点，还鉴定出其他豆科植物中已报道结瘤基因的大豆同源基因。我们通过启动子-GUS分析验证了部分基因的皮层差异表达特性，并借助RNA干扰（RNAi）技术探究了相关基因的功能。值得注意的是，本研究还筛选到一批参与生长素、细胞分裂素及赤霉素（gibberellic acid, GA）信号通路的差异调控基因，为深入解析早期根瘤发育过程中的植物激素信号调控机制提供了重要的理论依据。