丛枝菌根是土壤中的菌根真菌与植物根系间建立形成的互惠共生体。研究表明植物中存在着受菌根共生诱导、负责共生界面养分（如Pi、NH4+）吸收的转运体。但至今尚未有直接证据证明菌根共生界面上存在运输NO3-的转运体。前期我们从水稻菌根中鉴定到一个被强烈诱导表达的NPF家族（该家族大部分成员都可以转运NO3-）新成员OsNPF4.x。该基因主要在含有丛枝的细胞中表达，对NO3-有响应，ABA和SA处理显著下调其表达。鉴于NO3-可以作为信号分子调控根系发育，我们提出了该假说：OsNPF4.x能够通过NO3-或ABA/SA介导的信号途径调控水稻丛枝菌根共生。本项目将利用超表达和突变体材料研究该基因对水稻菌根途径NO3-吸收和菌根共生的影响。项目还将利用启动子功能分析和酵母单杂交系统筛选启动子中响应菌根共生信号的元件和与之作用的转录因子，以揭示OsNPF4.x响应和可能调控水稻丛枝菌根共生的作用机制。

Arbuscular mycorrhizal (AM) symbiosis between plant roots and arbuscular mycorrhizal fungi is one of the important evolutionary mechanisms of terrestrial plants to increase fitness to nutrient stresses. Several types of mycorrhiza-induced transporters responsible for translocating nutrients, such as Pi and NH4+, across the intraradical symbiotic interface have been isolated and well studied. However, no genes associated with transporting NO3- across the symbiotic interface have been evidenced to be present in plants. In our previous work, we have identified a mycorrhiza-induced gene, OsNPF4.x, from the rice NPF gene family, within which members are always characterized to have the ability of transporting NO3-. Expression analysis using GUS reporter revealed that the expression of OsNPF4.x was mainly confined to distinct cells containing arbuscules in rice mycorrhizal roots. OsNPF4.x was also observed to show response to NO3-, and its transcripts were significantly downregulated in response to ABA and SA applications. Given that NO3- has been repeatedly documented to be a signaling molecule that is able to modulate plant root development, it is tempting to make a hypothesis that OsNPF4.x could modulate the establishment of AM symbiosis, probably through mediating NO3- and/or ABA/SA (or other phytohormones) signaling pathways. In this study, we will functionally characterize this mycorrhiza-induced NPF gene, by enhancing or silencing its expression in transgenic plants, to testify its potential roles in transporting NO3- across the symbiotic interface and regulating mycorrhizal symbiosis and the related signaling pathways. The transgenic plants carrying mutated promoters (a series of truncations and deletions) fusing GUS reporter gene will also be employed to explore the putative cis-elements conferring AM-induced expression in OsNPF4.x promoter. In addition, EMSA (Electrophoretic Mobility Shift Assay) and Yeast One-Hybridization system will be performed to screen and testify the potential transcription factor(s) recognizing the AM-responsive cis-element(s). Accomplishing these studies, it will allow us to elucidate the molecular mechanisms underlying the response regulation and physiological roles of OsNPF4.x during the establishment of AM symbiosis in rice.