微小核酸對大岩桐花朵對稱性的調控

Abstract

花朵性狀的多樣發育變化形成開花植物在地球上繁盛出近三十萬種的多樣性，其中，演化出花朵兩側對稱性的物種，具有相異的背腹側花瓣形態，不但使花形變異更特化，花朵傳粉性狀特化也促進植物與昆蟲共演化。然而，對於建立背腹側花瓣極性的分子機制，目前學界尚未完全知曉。MicroRNAs (miRNAs)為一種內生的單股、非編碼的小片段RNA，長度約20-24個鹼基對。許多研究發現，miRNAs參與生物的基因表現調控，但目前尚未有研究在背腹側花瓣中探討miRNAs的調控。本實驗以野生型的大岩桐原種 (Sinningia speciosa) 兩側對稱花為材料，結合次世代定序與生物資訊分析，發現157個miRNAs表現於背腹側花瓣中。其中，不僅包含尚未在苦苣苔科發表過的conserved miRNAs，同時也有95個未曾在其他物種發表的novel miRNAs。有趣的是，我們發現miR157與miR390在背腹側花瓣有差異性表現，兩者在腹側花瓣皆偵測到較高的表現量。透過降解體定序，我們分別確認miR157與miR390的目標基因為SPLs與TAS3。SPLs曾被報導透過調控auxin-responsive genes來決定花器的大小；TAS3則是藉由產生ta-siRNAs，降解ARF3, ARF4，同樣調控auxin-responsive genes，以建立葉片上下表面的極性。轉錄體定序資料顯示，大岩桐的SPLs與ARF3同樣也具有背腹側表現差異。接著，我們進行stem-loop qPCR，再次驗證了定序資料的結果。綜合以上，本研究顯示miR157/SPLs與miR390/ARF3的調控路徑參與大岩桐花瓣發育。同時，此二路徑藉由調控下游的auxin-responsive genes，可能促使背腹側花瓣相異的形態發生。進一步，我們的發現也顯示，原先參與葉片上下表面極性的分子調控，可能擴展適應到建立花原基背腹側的極性。本實驗由轉譯後層次探討植物背腹側花瓣的差異分化，及其對於後續兩側對稱性的貢獻，有助於我們對於植物體花朵對稱性有更完整且全面的了解。

Establishment of dorsi-ventral polarity is essential for zygomorphic flower development, which contributes to the evolution of specialized pollination syndrome thus enhances species richness. But the mechanisms underlying the polarity establishment remain unclear. MicroRNAs (miRNAs) are embedded in the regulatory networks of plant development, but direct evidence for miRNAs regulating dorsi-ventral petal morphogenesis has been lacking. Here, we investigated the dorsi-ventrally expressed miRNAs in the zygomorphic flower of Sinningia speciosa, an economically important ornamental plant. Using a computational pipeline, we identified 157 miRNAs in developing petals, including both conserved miRNAs that are reported for the first time in S. speciosa, as well as 95 novel miRNAs that are not found in other plants. Interestingly, we discovered that miR157 and miR390 appeared more abundant in ventral petal than dorsal part. A combination of PARE analysis allowed us to validate their targets, respectively. SPLs, which are reported to be involved in regulating the floral organ size by controlling auxin-responsive genes, and TAS3, which have been invoked in the control of the specification of abaxial/adaxial polarity of leafs by generating ta-siRNAs, which in turn cause degradation of the AUXIN RESPONSE FACTOR 3 (ARF3) and ARF4 mRNAs. Intriguingly, our transcriptome data indicated that both SPLs and ARF3 show dorsi-ventral expression pattern. A similar result was also observed when we experimentally validated the expression patterns by qPCR. Our findings indicated that the regulation of miR157/SPLs may regulate the auxin-responsive genes, thus contribute to the different morphogenesis of dorsal and ventral petals. As for miR390/ARF3, the establishment of the ta-siRNA polarity is similar to the findings in leaves, which is a crucial determinant of abaial/adaxial polarity development. As a result, our finding raises an intriguing possibility of co-option in the development of leaf abaxial-adaxial polarity into floral dorsi-ventral zygomorphy.