IDD5的O-GlcNAc糖基化修饰在赤霉素信号转导通路中的作用

Protein modification by O-linked N-acetylglucosamine (O-GlcNAc) is a very import posttranslational modification in all eukaryotes. Extensive research in metazoan has demonstrated that, like phosphorylation, O-GlcNAc modification plays key roles in cellular regulation processes such as signal transduction and transcriptional regulation. O-GlcNAc modification has been shown to regulate various proteins by altering their subcellular localization, degradation, protein-protein interaction, and activities of transcription factors. There have been, however, very few studies of O-GlcNAc modification in plants. Recent studies revealed that O-GlcNAc modification may play important roles in Gibberellins (GA) signaling. From our previous proteomic study, we found that INDETERMINATE DOMAIN (IDD) prorotein family is O-GlcNAcylated. IDDs have been reported as a DELLA interacting transcription factors in GA signaling and regulate GA signaling responsive genes. Taken together, we hypothesize that O-GlcNAcylation of IDD5 may alter interaction between DELLA and IDD5 and/or binding affinity of IDD5 to its target promoters. In this proposed study, we will figure out what is the role of O-GlcNAcylation of IDD5 in GA signaling and how O-GlcNAcylation of IDD5 is regulated. Like phosphorylation, O-GlcNAc modification is believed as fine-tune factor for delicate, rapid, and proper response to cellular and environmental cues. This study will elucidate how GA signaling is precisely regulated to response various and complex signals by post-translational modifications.

氧连氮-乙酰葡糖胺(O-GlcNAc)糖基化修饰与磷酸化修饰类似，在细胞信号转导、基因表达等过程中发挥重要功能。O-GlcNAc修饰可改变蛋白的亚细胞定位、稳定性、蛋白间相互作用，及结合DNA的亲和性。研究表明O-GlcNAc对转录调控因子的修饰在赤霉素信号转导中发挥重要功能。我们研究组首次用质谱手段鉴定到植物中大量受O-GlcNAc修饰的蛋白以及其修饰位点；其中包括已知参与赤霉素信号转导的IDD家族转录因子。赤霉素诱导DELLA蛋白的降解来激活受DELLA抑制的转录因子，进而改变基因表达和细胞生长。而IDD5是受DELLA直接抑制的转录因子，直接参与对部分赤霉素响应基因的调控。O-GlcNAc修饰可能影响IDD5与RGA的相互作用、结合启动子的能力，或转录活性。本研究将通过遗传和生化手段阐明IDD5的O-GlcNAc修饰在GA信号通路中的作用，以及影响IDD5功能的分子机制。

氧连氮-乙酰葡糖胺(O-GlcNAc)糖基化修饰是一种动态的蛋白质翻译后修饰，在调控蛋白的亚细胞定位、稳定性及蛋白的相互作用方面发挥着重要的作用。在植物中，O-GlcNAc也被发现在众多的生物学过程中发挥着重要的作用。前期的研究表明参与赤霉素信号转导的IDD5蛋白上存在O-GlcNAc修饰，但是具体的生物学意义和机制不明。本项目将IDD5蛋白与糖基化修饰转移酶SPY与SEC共转化，发现O-GlcNAc修饰不影响转录因子IDD5的核定位，但是能够增加IDD5蛋白的稳定性。在IDD5蛋白存在时，共表达SEC还能够增加RGA蛋白的稳定性。通过诱导干扰两个OGT基因的表达，严重影响了多个GA参与的植物的生长发育过程，如下胚轴伸长、根的生长、叶柄伸长等。RNA-seq分析也发现当O-GlcNAc修饰缺失时，多个GA信号通路的基因的转录水平受到影响。利用免疫沉淀及质谱技术，发现IDD5也是SPY蛋白的相互作用蛋白，过量表达IDD5表现出植株矮小、叶柄变短的GA缺失或者不敏感的表型。本项目初步解析了IDD5的O-GlcNAc对自身蛋白稳定性的影响，过量表达IDD5对GA相关表型起到了负调控的作用，这些结果为进一步分析IDD5蛋白的功能以及O-GlcNAc修饰对GA信号转导影响提供了实验证据，对后续解析O-GlcNAc修饰在植物中的作用具有重要的意义。