College of Horticulture, Nanjing Agricultural University, Nanjing, 210095, China.
Department of Agriculture and Food Technology, Karakoram International University, Gilgit, 15100, Pakistan.
Mol Genet Genomics. 2018 Dec;293(6):1333-1354. doi: 10.1007/s00438-018-1462-1. Epub 2018 Jun 25.
SPL is a plant-specific transcription factor family. Many researchers reported that SPL members targeted by miR156s could play crucial roles in the modulation of plant growth and development. Although there are similar reports on grapes, till now little is known about grape berry development and ripening. To gain more insight into how grape miR156s (Vv-miR156s) modulated the above given processes of grape berries by mediating their target gene Vv-SPLs, here we identified the precise sequences of Vv-miR156s in 'Giant Rose' grape berries, predicted their potential targets, and revealed that the matching degree of various Vv-miR156: Vv-SPL pairs exhibited some discrepancy, implying the divergence of their interaction. Subsequently, we also discovered similar motifs such as ABRE, CGTCA and ERE, which are more specific to berry development and ripening, within the promoters of both Vv-MIR156s and Vv-SPLs. With berry development and ripening, meanwhile, Vv-miR156a, b/c/d, e and f/g/i exhibited an overall increasing expression trend, while their targets showed opposite trends at the corresponding stages. Additionally, exogenous ABA and NAA application promoted or curbed the expression of Vv-miR156s to some extent, before grape berry ripening stage. The cleavage products, sites and frequencies of Vv-miR156a, b/c/d, e, f/g/i and their respective targets (Vv-SPL2, 9, 10, 16) during grape berry development and ripening process were validated by our developed PPM-RACE and modified RLM-RACE together with qRT-PCR, which demonstrated that Vv-miR156s can be involved in the modulation of grape berry development and ripening process by mediating the expression of Vv-SPL2, 9, 10, 16. Our findings lay an important foundation for further recognizing their functions in grape berries, and enrich the knowledge of the regulatory mechanism of miRNA-mediated grape berry development and ripening.
SPL 是一个植物特有的转录因子家族。许多研究人员报告说，miR156s 靶向的 SPL 成员可以在植物生长和发育的调节中发挥关键作用。尽管在葡萄上也有类似的报道，但到目前为止，人们对葡萄浆果的发育和成熟过程知之甚少。为了更深入地了解葡萄 miR156s（Vv-miR156s）如何通过调节其靶基因 Vv-SPLs 来调节葡萄浆果的上述过程，我们在这里鉴定了‘巨玫瑰’葡萄浆果中 Vv-miR156s 的精确序列，预测了它们的潜在靶标，并发现了各种 Vv-miR156：Vv-SPL 对的匹配程度存在一些差异，这表明它们的相互作用存在分歧。随后，我们还在 Vv-MIR156s 和 Vv-SPLs 的启动子中发现了类似的基序，如 ABRE、CGTCA 和 ERE，这些基序更特定于浆果发育和成熟。随着浆果的发育和成熟，Vv-miR156a、b/c/d、e 和 f/g/i 的表达呈整体上升趋势，而它们的靶标在相应阶段则呈相反趋势。此外，在葡萄浆果成熟之前，外源 ABA 和 NAA 的应用在一定程度上促进或抑制了 Vv-miR156s 的表达。通过我们开发的 PPM-RACE 和改良的 RLM-RACE 以及 qRT-PCR 验证了 Vv-miR156a、b/c/d、e、f/g/i 及其各自靶标（Vv-SPL2、9、10、16）在葡萄浆果发育和成熟过程中的切割产物、位点和频率，这表明 Vv-miR156s 可以通过调节 Vv-SPL2、9、10、16 的表达来参与葡萄浆果发育和成熟过程的调节。我们的研究结果为进一步认识它们在葡萄浆果中的功能奠定了重要基础，并丰富了 miRNA 介导的葡萄浆果发育和成熟调控机制的知识。
