Key Laboratory of Horticultural Plant Biology (Ministry of Education), College of Horticulture and Forestry Sciences, Huazhong Agricultural University, Wuhan, 430070, China.
Guangdong Key Laboratory for Innovative Development and Utilization of Forest Plant Germplasm, College of Forestry and Landscape Architecture, South China Agricultural University, Guangzhou, 510642, China.
Plant Cell Rep. 2022 Jun;41(6):1403-1415. doi: 10.1007/s00299-022-02865-y. Epub 2022 Apr 5.
Overexpression of miR171 restored SE competence in the recalcitrant citrus callus, and inhibition of miR171 function weakened SE competence in the strongly embryogenic citrus callus. Somatic embryogenesis (SE) is an important way of in vitro regeneration for plants. For perennial woody crops such as citrus, embryogenic callus is usually induced from unfertilized aborted ovules and widely used in biotechnology aided breeding. However, SE capacity always declines in callus during subculture, which makes regeneration difficult and hinders the application of biotechnology. We previously found that miR171 may be a regulator of SE in citrus, based on the abundant expression of csi-miR171c in the embryogenic callus and during SE of citrus. Here, we report that miR171 promotes SE and is required for SE in citrus. Overexpression of miR171 restored SE competence in the recalcitrant callus of 'Guoqing No.1' Satsuma mandarin (G1), whereas inhibition of miR171 function by Short Tandem Target Mimic (STTM) weakened SE competence in the strongly embryogenic callus of 'Valencia' sweet orange (V). The comparative transcriptomic analysis in miR171 overexpressed callus line (OE) and the wild type callus (WT) indicated that overexpression of miR171 decreased the expression level of its SCARECROW-LIKE (CsSCL) targets, and activated stress response related biological processes and metabolic processes that are required for cell differentiation. However, CsSCLs were up-regulated in the OE callus during SE induction process, which activated the cell division and developmental processes that are required for embryogenesis progress. Our results validate the function of miR171 in regulation of SE and reveal the biological responses provoked by miR171 in citrus that may promote SE.
miR171 的过表达恢复了顽固柑橘愈伤组织的 SE 能力，而 miR171 功能的抑制则削弱了强胚胎发生柑橘愈伤组织的 SE 能力。体细胞胚胎发生 (SE) 是植物体外再生的重要途径。对于柑橘等多年生木本作物，胚胎发生愈伤组织通常是从未受精的败育胚珠中诱导产生的，并广泛应用于生物技术辅助育种。然而，SE 能力在愈伤组织的继代培养中总是会下降，这使得再生变得困难，阻碍了生物技术的应用。我们之前发现，基于 csi-miR171c 在胚胎发生柑橘中的大量表达，miR171 可能是柑橘 SE 的调节剂。在这里，我们报告说 miR171 促进 SE 并在柑橘中是 SE 所必需的。miR171 的过表达恢复了‘国光 1 号’温州蜜柑（G1）顽固愈伤组织的 SE 能力，而通过短串联靶标模拟（STTM）抑制 miR171 功能则削弱了‘瓦伦西亚’甜橙（V）强胚胎发生愈伤组织的 SE 能力。miR171 过表达愈伤组织系（OE）和野生型愈伤组织（WT）的比较转录组分析表明，miR171 的过表达降低了其 SCARECROW-LIKE（CsSCL）靶标的表达水平，并激活了细胞分化所需的应激反应相关生物过程和代谢过程。然而，在 SE 诱导过程中，OE 愈伤组织中 CsSCLs 的上调激活了胚胎发生进展所需的细胞分裂和发育过程。我们的结果验证了 miR171 在 SE 调控中的功能，并揭示了 miR171 在柑橘中引发的可能促进 SE 的生物学反应。
