Background and Aims Aquilegia produce elongated, three-dimensional petal spurs that fill with nectar to attract pollinators. Previous studies have shown that the diversity of spur length across the Aquilegia genus is a key innovation that is tightly linked with its recent and rapid diversification into new ranges, and that evolution of increased spur lengths is achieved via anisotropic cell elongation. Previous work identified a brassinosteroid response transcription factor as being enriched in the early developing spur cup. Brassinosteroids are known to be important for cell elongation, suggesting that brassinosteroid-mediated response may be an important regulator of spur elongation and potentially a driver of spur length diversity in Aquilegia. In this study, we investigated the role of brassinosteroids in the development of the Aquilegia coerulea petal spur. Methods We exogenously applied the biologically active brassinosteroid brassinolide to developing petal spurs to investigate spur growth under high hormone conditions. We used virus-induced gene silencing and gene expression experiments to understand the function of brassinosteroid-related transcription factors in A. coerulea petal spurs. Key Results We identified a total of three Aquilegia homologues of the BES1/BZR1 protein family and found that these genes are ubiquitously expressed in all floral tissues during development, yet, consistent with the previous RNAseq study, we found that two of these paralogues are enriched in early developing petals. Exogenously applied brassinosteroid increased petal spur length due to increased anisotropic cell elongation as well as cell division. We found that targeting of the AqBEH genes with virus-induced gene silencing resulted in shortened petals, a phenotype caused in part by a loss of cell anisotropy. Conclusions Collectively, our results support a role for brassinosteroids in anisotropic cell expansion in Aquilegia petal spurs and highlight the brassinosteroid pathway as a potential player in the diversification of petal spur length in Aquilegia.

中文翻译：

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油菜素类固醇通过控制细胞伸长来调节耧斗菜的花瓣刺长度

背景和目的 Aquilegia 产生细长的三维花瓣刺，其中充满花蜜以吸引传粉者。先前的研究表明，芫荽属的刺长度的多样性是一项关键创新，这与其最近快速多样化到新的范围密切相关，并且增加的刺长度的演变是通过各向异性细胞伸长来实现的。以前的工作确定了一种油菜素内酯反应转录因子在早期发育的支杯中富集。已知油菜素内酯对细胞伸长很重要，这表明油菜素内酯介导的反应可能是刺伸长的重要调节剂，并可能是耧斗菜中刺长度多样性的驱动因素。在这项研究中，我们调查了油菜素内酯在蓝耧斗菜花瓣刺发育中的作用。方法 我们将具有生物活性的油菜素内酯油菜素内酯外源应用于发育中的花瓣骨刺，以研究在高激素条件下的花刺生长。我们使用病毒诱导的基因沉默和基因表达实验来了解油菜素内酯相关转录因子在 A. coerulea 花瓣刺中的功能。关键结果 我们总共鉴定了 BES1/BZR1 蛋白家族的三个 Aquilegia 同源物，并发现这些基因在发育过程中在所有花组织中普遍表达，然而，与之前的 RNAseq 研究一致，我们发现其中两个旁系同源物被富集在早期发育的花瓣中。由于各向异性细胞伸长和细胞分裂增加，外源应用油菜素内酯增加了花瓣刺长度。我们发现用病毒诱导的基因沉默靶向 AqBEH 基因会导致花瓣缩短，这是一种表型，部分原因是细胞各向异性的丧失。结论 总的来说，我们的研究结果支持油菜素类固醇在耧斗菜花瓣刺中各向异性细胞扩增中的作用，并强调油菜素类固醇途径是耧斗菜花瓣刺长度多样化的潜在参与者。