2013-08-012024-05-18https://scholars.lib.ntu.edu.tw/handle/123456789/703954摘要：在高鹽逆境下，植物通過調整信號傳導路徑對鹽逆境產生反應，此包括離層酸依存及獨立的途徑。在前人的研究中發現bZIP 轉錄因子參與鹽或滲透逆境的反應和抗性及離層酸信號傳導，然而，其分子網絡及調控機制仍有大部分是未知的。我們希望找尋尚未被發現之受鹽分調控之bZIP 基因，並探求其分子調控機制。此計畫以阿拉伯芥為材料，最終目標是要闡明bZIP 基因參與鹽或滲透逆境的反應及抗性機制，和在離層酸信號傳導中的作用機制。35S 啟動子驅動的AtbZIP 過量表達植株將被建立，並測試受到鹽、滲透壓力或離層酸處理之反應。我們將使用微陣列晶片、chromation immunoprecipitation 及蛋白質組學分析AtbZIP 失去功能及得到功能突變株在鹽、滲透壓力或離層酸處理後下游受調控之基因及蛋白。此外，我們將更進一步純化bZIP 之蛋白複合體，並利用質譜分析bZIP 互動蛋白之身分，並希望利用轉錄活化法分析互動蛋白對轉錄因子之轉錄活性調控。另一方面，我們也希望研究bZIP 及AP2/ERF轉錄因子是否及如何協同參與鹽或滲透逆境的反應。初步結果中發現一些 AtbZIP 基因的過度表達之阿拉伯芥轉植株是耐鹽的，其中包括ABF3 及之前尚未被發現之bZIP 基因。此外，經雙分子螢光互補系統分析發現AtABF3 蛋白與14-3-3 蛋白有交互作用。我們希望這項研究能讓我們更好地瞭解bZIP 基因的作用機轉。<br> Abstract: Under salinity stress, crop yield is significantly reduced. Plants respond to salt stress by adjusting various signaling transduction pathways including abscisic acid (ABA)-dependent and ABA-independent pathways. In previous studies, bZIP transcription factor was found to be involved in salt or osmotic stress response and tolerance. In addition, bZIP was found to be involved in ABA signaling pathway. However, the molecular network and mechanism is still largely unknown. We aim to elucidate the mechanism of bZIP gene in salt or osmotic stress response, and involve in ABA signaling pathway in planta. In this proposed project we will use model organism Arabidopsis thaliana as plant materials. The ultimate goal of this project is to elucidate the function of AtbZIP in salt or osmotic stress response, and ABA signal transduction pathway. Over-expression line of HA-tagged bZIP driven by 35S promoter will be generated and subjected to phenotyping to observe salt, osmotic stress, or ABA response. In order to identify downstream genes or proteins affected by salt or osmotic stress, transcriptomic and proteomic analysis will be introduced. We will use microarray analysis and chromatin immunoprecipitation (ChIP) to reveal the up-regulated and down-regulated genes using T-DNA knockout mutant and over-expression line under salt, osmotic stress, or ABA treatment. A 2D-gel based and gel-free proteomic analysis will be introduced to compare protein accumulation in wild type and mutant line under salt, osmotic stress, or ABA treatment. In addition, we will utilize affinity column to purify AtbZIP protein complex in vivo followed by mass spectrometry analyses for protein identification to identify bZIP interacting proteins. We will also study the role of interacting protein in the transcriptional regulation by transactivation. Moreover, we will study if and how bZIP and AP2/ERF synergistically involve in salt or osmotic stress response. In our preliminary results, we found that at least 8 independent AtbZIP gene overexpression lines are salt-tolerant. These included known abiotic stress and ABA-regulated gene ABF3 and some uncharacterized bZIP genes. Moreover, ABF3 can interact with 14-3-3 protein in the nucleus based on our transient assay using bimolecular fluorescence complementation (BiFC). We hope that this study will lead to a better understanding of the function of bZIP gene in salt or osmotic stress and ABA response. We hope we can eventually apply this to crop (i.e. rice) to improve salt stress resistance and improve crop yield.bZIP鹽滲透壓離層酸14-3-3bZIPsaltosmoticABA14-3-3