Chen Yaping, Chen Wei, Li Xueliu, Jiang Huawu, Wu Pingzhi, Xia Kuaifei, Yang Yali, Wu Guojiang
Key Laboratory of Plant Resources Conservation and Sustainable Utilization, South China Botanical Garden, Chinese Academy of Sciences, Guangzhou 510650, China.
Plant Cell Physiol. 2014 Jan;55(1):183-93. doi: 10.1093/pcp/pct171. Epub 2013 Nov 26.
Cytokinins play important roles in legume-rhizobia symbiosis. Here we report isolation of six genes encoding isopentenyl transferase (IPT) from Lotus japonicus, which catalyze the rate-limiting step of cytokinin biosynthesis. The LjIPT3 gene was found to be up-regulated in infected roots and mature nodules. Histochemical analysis demonstrated expression of Pro(LjIPT3):GUS (β-glucuronidase) in vegetative and reproductive organs, and was especially high in the vascular bundles of roots. When inoculated with Mesorhizobium loti MAFF303099, LjIPT3 was undetectable in the nodule primordia and developing nodules, and later it was expressed only in the vascular bundles of mature nodules. In addition, knockdown of LjIPT3 (LjIPT3i) by RNA interference reduced levels of endogenous cytokinins, affected plant development and accelerated Chl degradation during dark-induced leaf senescence. Compared with the wild type, LjIPT3i plants produced fewer infection threads and nodules. In addition, expression of downstream nodulation-related transcription factor genes LjNSP1, LjNSP2 and LjNIN decreased dramatically in LjIPT3i plants. These results suggest that LjIPT3 regulates the CRE1-dependent cytokinin pathway, affecting nodule initiation and thereby influencing the number of infection threads and nodules. Detection of nitrogenase activity and observation of nodule structure showed that endogenous cytokinins are required for full development of the infected cells in mature nodules by preventing early senescence. Therefore, our results indicate that the LjIPT3 gene product is required for nodule initiation and development, and does not appear to be involved in early infection events.
细胞分裂素在豆科植物与根瘤菌的共生关系中发挥着重要作用。在此，我们报告从百脉根中分离出六个编码异戊烯基转移酶（IPT）的基因，这些基因催化细胞分裂素生物合成的限速步骤。研究发现LjIPT3基因在受侵染的根和成熟根瘤中上调表达。组织化学分析表明，Pro（LjIPT3）：GUS（β-葡萄糖醛酸酶）在营养器官和生殖器官中均有表达，在根的维管束中表达量尤其高。用中慢生根瘤菌MAFF303099接种时，在根瘤原基和发育中的根瘤中未检测到LjIPT3，随后它仅在成熟根瘤的维管束中表达。此外，通过RNA干扰敲低LjIPT3（LjIPT3i）会降低内源细胞分裂素水平，影响植物发育，并加速黑暗诱导的叶片衰老过程中的叶绿素降解。与野生型相比，LjIPT3i植株产生的侵染线和根瘤较少。此外，LjIPT3i植株中与结瘤相关的下游转录因子基因LjNSP1、LjNSP2和LjNIN的表达显著降低。这些结果表明，LjIPT3调节依赖于CRE1的细胞分裂素途径，影响根瘤起始，从而影响侵染线和根瘤的数量。固氮酶活性检测和根瘤结构观察表明，内源细胞分裂素通过防止早期衰老，对成熟根瘤中被侵染细胞的充分发育是必需的。因此，我们的结果表明，LjIPT3基因产物是根瘤起始和发育所必需的，似乎不参与早期感染事件。
