阿拉伯芥中FIN219透過與NPH3交互作用參與向光性反應之功能性研究

Abstract

在阿拉伯芥中，FIN219 (FAR-RED INSENSITIVE 219)會協助生合成具有生物活性的茉莉酸 (JA-Ile)，是植物茉莉酸參與防禦反應與調控植物生長發育中十分重要的角色。植物的向光性反應主要受到藍光的光接受者PHOTOTROPINS (PHOTs)與其下游的訊息傳遞者NON-PHOTOTROPIC HYPOCOTYL3 (NPH3)所調節。在先前研究發現缺乏FIN219似乎會影響阿拉伯芥的向光性反應。為了更加了解FIN219是如何參與向光性反應機制，因此首先透過fin219-2突變株在接收到側邊藍光的照射後，發現表現出向光性缺失的外表型。我們透過Semi-CoIP assay 和雙分子螢光互補系統(BiFC)實驗證明了，PHOT1與FIN219蛋白質之間在細胞質與細胞膜有著交互作用的關係。並且透過Pull-down assay、Co-IP與BiFC也證明了NPH3與FIN219蛋白質之間也有著交互作用的現象。有趣的是，透過西方墨點法(Western blot)實驗發現，phot1、phot1/2與nph3突變株中FIN219蛋白質的表現量有明顯降低，說明在藍光所調節的向光性反應中光接受者PHOT1與下游的訊息傳遞者NPH3會正向調控FIN219，而PHOT1會穩定NPH3與FIN219之間的交互作用。當缺乏FIN219時NPH3無法正常去磷酸化，進而出現向光性缺失的情況。總之，FIN219在PHOT1，NPH3引發的阿拉伯芥向光性反應中扮演重要的角色。

In Arabidopsis, FIN219 (FAR-RED INSENSITIVE 219) participates in the biosynthesis of biologically active jasmonic acid-isoleucine (JA-Ile), playing a crucial role in plant defense response and plant growth and development. The phototropic response in plants is mainly regulated by the blue light photoreceptors known as PHOTOTROPINS (PHOTs), and their downstream signaling component NON-PHOTOTROPIC HYPOCOTYL3 (NPH3). Previous studies indicated that the absence of FIN219 seems to affect the phototropic response in Arabidopsis. To gain further insights into the interaction between FIN219 and the phototropic response mechanism, we firstly observed a defect of phototropism phenotype in the fin219-2 mutant upon exposure to lateral blue light. Through Semi-CoIP assay and Bimolecular Fluorescence Complementation (BiFC), we demonstrated the interaction between PHOT1 and FIN219 protein in the cytoplasm and cell membrane. Furthermore, Pull-down assay, CoIP, and BiFC experiments showed the interaction between NPH3 and FIN219. Interestingly, protein immunoblot analysis (Western blot) revealed a significant reduction in the expression level of FIN219 protein in phot1, phot1/2, and nph3 mutants, suggesting that the upstream blue light photoreceptor PHOT1 and downstream signaling component NPH3 positively regulate FIN219. Moreover, PHOT1 stabilizes the interaction between NPH3 and FIN219. In the absence of FIN219, NPH3 fails to undergo proper dephosphorylation, resulting in a loss of phototropism phenotype. Taken together, FIN219 plays a vital role in phot1-NPH3-mediated phototropic response in Arabidopsis.