College of Plant Protection, Shenyang Agricultural University, Shenyang, China.
Division of Applied Life Science (BK21 Program), Plant Molecular Biology and Biotechnology Research Center (PMBBRC), Gyeongsang National University, Jinju, Korea.
Ann Bot. 2019 Nov 27;124(6):947-960. doi: 10.1093/aob/mcy242.
INDETERMINATE DOMAIN 10 (IDD10) is a key transcription factor gene that activates the expression of a large number of NH4+-responsive genes including AMMONIUM TRANSPORTER 1;2 (AMT1;2). Primary root growth of rice (Oryza sativa) idd10 mutants is hypersensitive to NH4+. The involvement of CALCINEURIN B-LIKE INTERACTING PROTEIN KINASE (CIPK) genes in the action of IDD10 on NH4+-mediated root growth was investigated.
Quantitative reverse transcription-PCR was used to analyse NH4+- and IDD10-dependent expression of CIPK genes. IDD10-regulated CIPK target genes were identified using electrophoretic mobility shift assays, chromatin immunoprecipitation and transient transcription assays. Root growth rate, ammonium content and 15N uptake of cipk mutants were measured to determine their sensitivity to NH4+ and to compare these phenotypes with those of idd10. The genetic relationship between CIPK9 OX and idd10 was investigated by crosses between the CIPK9 and IDD10 lines.
AMT1;2 was overexpressed in idd10 to determine whether NH4+-hypersensitive root growth of idd10 resulted from limitations in NH4+ uptake or from low cellular levels of NH4+. High NH4+ levels in idd10/AMT1;2 OX did not rescue the root growth defect. Next, the involvement of CIPK genes in NH4+-dependent root growth and interactions between IDD10 and CIPK genes was investigated. Molecular analysis revealed that IDD10 directly activated transcription of CIPK9 and CIPK14. Expression of CIPK8, 9, 14/15 and 23 was sensitive to exogenous NH4+. Further studies revealed that cipk9 and idd10 had almost identical NH4+-sensitive root phenotypes, including low efficiency of 15NH4+ uptake. Analysis of plants containing both idd10 and CIPK9 OX showed that CIPK9 OX could rescue the NH4+-dependent root growth defects of idd10.
CIPK9 was involved in NH4+-dependent root growth and appeared to act downstream of IDD10. This information will be useful in future explorations of NH4+ signalling in plants.
不确定域 10（IDD10）是一个关键的转录因子基因，它激活大量 NH4+-响应基因的表达，包括铵转运蛋白 1；2（AMT1；2）。水稻（Oryza sativa）idd10 突变体的主根生长对 NH4+敏感。研究了钙调神经磷酸酶 B 样相互作用蛋白激酶（CIPK）基因在 IDD10 对 NH4+-介导的根生长作用中的参与。
使用定量逆转录-PCR 分析 CIPK 基因的 NH4+-和 IDD10 依赖性表达。使用电泳迁移率变动分析、染色质免疫沉淀和瞬时转录分析鉴定 IDD10 调节的 CIPK 靶基因。测量 cipk 突变体的根生长速度、铵含量和 15N 摄取，以确定它们对 NH4+的敏感性，并将这些表型与 idd10 的表型进行比较。通过 CIPK9 线和 IDD10 线之间的杂交，研究了 CIPK9 OX 与 idd10 之间的遗传关系。
在 idd10 中过表达 AMT1；2，以确定 idd10 中 NH4+-敏感的根生长是否是由于 NH4+摄取的限制或由于细胞内 NH4+水平低所致。在 idd10/AMT1；2 OX 中高 NH4+水平并没有挽救根生长缺陷。接下来，研究了 CIPK 基因在 NH4+-依赖性根生长中的参与以及 IDD10 和 CIPK 基因之间的相互作用。分子分析表明，IDD10 直接激活 CIPK9 和 CIPK14 的转录。CIPK8、9、14/15 和 23 的表达对外源 NH4+敏感。进一步的研究表明，cipk9 和 idd10 具有几乎相同的 NH4+-敏感的根表型，包括 15NH4+摄取效率低。分析含有 idd10 和 CIPK9 OX 的植物表明，CIPK9 OX 可以挽救 idd10 中 NH4+-依赖性根生长缺陷。
CIPK9 参与 NH4+-依赖性根生长，似乎在 IDD10 下游发挥作用。这些信息将有助于未来对植物中 NH4+信号的探索。
