MicroRNA390-Directed TAS3 Cleavage Leads to the Production of tasiRNA-ARF3/4 During Somatic Embryogenesis in Dimocarpus longan Lour
MicroRNA390-Directed TAS3 Cleavage Leads to the Production of tasiRNA-ARF3/4 During Somatic Embryogenesis in Dimocarpus longan Lour
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MicroRNA390 引导的 TAS3 裂解导致龙眼体细胞胚胎发生过程中 tasiRNA-ARF3/4 的产生
DOI：
10.3389/fpls.2075.07719
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发表时间：
2015-12-16
影响因子：
5.6
通讯作者：
Lai, Zhongxiong
中科院分区：
文献类型：
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作者：
Lin, Yuling;Lin, Lixia;Lai, Zhongxiong
反式作用短干扰RNA（tasiRNA）来源于TAS 3家族，通过microRNA（miRNA）390引导初级转录物和靶向生长素反应因子（ARF 3/-4）的切割，其参与植物侧根和花的正常发育。然而，它们在胚胎发育中的作用仍不清楚。首次系统鉴定了龙眼体细胞胚发育过程中miR 390-TAS 3-ARF 3/-4途径。我们鉴定了miR 390的初级转录本和启动子。该启动子含有顺式作用元件，响应于刺激，如光，水杨酸，厌氧诱导，真菌诱导子，昼夜节律控制，和热应激。分离了含有两个miR 390结合位点的龙眼TAS 3转录物;验证了位于TAS 3转录物3'端附近的miR 390引导的切割位点。在龙眼小RNA数据中发现了8个具有21个核苷酸相位的TAS 3-tasiRNA，进一步证实了miR 390指导的TAS 3切割导致龙眼中tasiRNA的产生。其中，TAS3_5 'D5+和5' D 6 + tasiRNA的丰度较高，并被证实靶向ARF 3和ARF-4，这表明miR 390引导的TAS 3切割具有21个核苷酸的阶段，导致tasiRNA-ARF的产生在植物中是保守的。Pri-miR 390在脆性胚性愈伤组织（EC）中高表达，在不完全致密的原胚性愈伤组织中表达较少，而miR 390在EC中表达最低，在鱼雷形胚（TE）中表达最高。DITAS 3和D1 ARF 4在EC中的表达量最低，在球形胚期达到最高，与miR 390的表达量成反比，尤其是在球形胚至子叶胚（CEs）期。而DIARF 3从EC到TE阶段变化很小，并且在CE阶段表现出最低表达。D1 ARF 3和miR 390的表达之间普遍缺乏相关性。此外，2，4-D以浓度依赖性方式上调pri-miR 390、DITAS 3、D1 ARF 3和-4。它们在四季蜜龙眼的根、果肉和种子中也有表达，表明它们在龙眼根和果实发育中的作用。该研究为miR 390-tasiRNA-ARF在植物体细胞胚发育中的可能作用提供了新的见解。
Trans-acting short-interfering RNAs (tasiRNAs) originate from TAS3 families through microRNA (miRNA) 390 guided cleavage of primary transcripts and target auxin response factors (ARF3/-4), which are involved in the normal development of lateral roots and flowers in plants. However, their roles in embryo development are still unclear. Here, the pathway miR390-TAS3-ARF3/-4 was identified systematically for the first time during somatic embryo development in Dimocarpus longan. We identified the miR390 primary transcript and promoter. The promoter contained cis-acting elements responsive to stimuli such as light, salicylic acid, anaerobic induction, fungal elicitor, circadian control, and heat stress. The longan TAS3 transcript, containing two miR390-binding sites, was isolated; the miR390- guided cleavage site located near the 3' end of the TAS3 transcript was verified. Eight TAS3-tasiRNAs with the 21-nucleotides phase were found among longan small RNA data, further confirming that miR390-directed TAS3 cleavage leads to the production of tasiRNA in longan. Among them, TAS3_5'D5+ and 5'D6+ tasiRNAs were highly abundant, and verified to target ARF3 and -4, implying that miR390-guided TAS3 cleavage with 21-nucleotides phase leading to the production of tasiRNA-ARF is conserved in plants. Pri-miR390 was highly expressed in friable-embryogenic callus (EC), and less expressed in incomplete compact pro-embryogenic cultures, while miR390 showed its lowest expression in EC and highest expression in torpedo-shaped embryos (TEs). DITAS3 and D1ARF4 both exhibited their lowest expressions in EC, and reached their peaks in the globular embryos stage, which were mainly inversely proportional to the expression of miR390, especially at the globular embryos to cotyledonary embryos (CEs) stages. While DIARF3 showed little variation from the EC to TEs stages, and exhibited its lowest expression in the CEs stage. There was a general lack of correlation between the expressions of D1ARF3 and miR390. In addition, pri-miR390, DITAS3, D1ARF3 and -4 were up-regulated by 2,4-D in a concentration-dependent manner. They were also preferentially expressed in roots, pulp, and seeds of 'Sijimi' longan, implying their extended roles in the development of longan roots and fruit. This study provided insights into a possible role of miR390-tasiRNAs-ARF in plant somatic embryo development.