阿拉伯芥BASIC LEUCINE-ZIPPER 6 (bZIP6) 參與在茉莉酸和乙烯的訊息傳遞路徑去調控ETHYLENE RESPONSE FACTOR 1(ERF1)的基因表現

植物在遭遇非生物逆境時，AP2/ERF蛋白扮演著非常重要的角色來調控植物的生長與發育，目前已知ETHYLENE RESPONSE FACTOR 1 (ERF1) 在阿拉伯芥裡被分類在第IX族群的轉錄因子會參與許多不同荷爾蒙的傳遞訊息路徑。在前人的研究中，當植物遇到necrotrophic pathogens，ERF1基因受到茉莉酸 (JA) 與乙烯 (ET) 共同誘導而表現，賦予植物抵抗生物逆境的耐受性。先前的研究顯示， ERF1基因受到非生物逆境的誘導而表現，賦予植物抵抗生物逆境的耐受性。ERF1基因受到JA和ET的誘導表現一旦同時處理離層酸 (ABA) 則強烈的受到抑制，暗示著ABA可能直接或是間接透過JA或ET訊息傳遞來調控ERF1基因表現。在本研究中，利用原生質體去處理ABA、JA和ET之後進行轉錄活性的分析實驗，證明ERF1啟動子上唯一ABRE (ABA-response element) 序列是具有生物功能性的。另外透過酵母菌單雜交的實驗找到BASIC LEUCINE-ZIPPER 2 (bZIP2)、bZIP6和bZIP7可以結合到ERF1啟動子上游的ABRE序列。此外，利用原生質體進行轉錄分析的實驗顯示除了bZIP2之外，bZIP6和bZIP7可以結合到ERF1啟動子上游的ABRE序列。從轉錄活性的分析實驗顯示bZIP6可以活化LUCIFERASE表現，而bZIP7可以抑制LUCIFERASE表現，說明bZIP6是一個活化因子而bZIP7是一個抑制因子。利用膠體電泳位移分析和染色質免疫沉澱法驗證了bZIP6會結合到ERF1啟動子上游的ABRE序列。bZIP6基因不僅會受到乾旱、滲透壓和鹽逆境的誘導且會受到JA和ET的誘導表現，但bZIP6基因不受ABA的影響。在35S::bZIP6-GFP轉植株的ERF1的基因表現相對於野生型而言明顯較高。總結bZIP6參與在JA或ET的訊息傳遞路徑中會扮演一個新的角色直接調控ERF1的基因表現。

關鍵字

bZIP6 ； ERF1 ；非生物逆境；離層酸；茉莉酸；乙烯

並列摘要

AP2/ERF proteins play crucial roles in plant growth and development and in response to environment stress. ETHYLENE RESPONSE FACTOR 1 (ERF1) transcription factor belongs to group IX in ERF family which is involved in many different hormones signaling pathways. ERF1 gene expression was synergistically activated by the jasmonate (JA) and ethylene (ET) when plant was attacked by necrotrophic pathogens to confer biotic stress tolerance. Previous study also showed that ERF1 can be induced by various abiotic stresses to confer abiotic stress tolerance. ERF1 gene expression induced by JA and ET was strongly repressed by abscisic acid (ABA) treatment implying ABA may regulate ERF1 gene expression directly or JA/ET signaling indirectly. In this study, using protoplast transactivation assay I demonstrated the biological function of ABRE (ABA-response element) cis-element in ERF1 promoter under ABA, JA and ET treatment. I found BASIC LEUCINE-ZIPPER 2 (bZIP2), bZIP6 and bZIP7 could bind to the only ABRE cis-element in the promoter of ERF1 gene using yeast one-hybrid assay. Furthermore, protoplast transactivation assay showed bZIP6 and bZIP7 could interact with ABRE cis-element but bZIP2 couldn’t. Transactivation assay revealed that bZIP6 could activate the expression of LUCIFERASE while bZIP7 could repress it, indicating bZIP6 is an activator and bZIP7 is a repressor. Moreover, bZIP6 binding to the ABRE-cis element was verified by EMSA (electrophoretic mobility shift assay) and ChIP (chromatin immunoprecipitation) assay. The bZIP6 gene expression can be induced by drought, mannitol and salt stress treatment, and also can be induced by JA or ET but not respond to ABA. In 35S::bZIP6-GFP overexpression line, ERF1 expression was greater when compare with WT plant. In conclusion, bZIP6 might be a new player involved in JA or ET signaling pathway to directly regulate ERF1 gene expression.

並列關鍵字

bZIP6 ； ERF1 ； abiotic stress ； ABA ； JA ； ET

參考文獻

Anderson JP, Badruzsaufari E, Schenk, PM, Manners JM, Desmond OJ, Ehlert C, Maclean DJ, Ebert PR, Kazan K (2004) Antagonistic interaction between abscisic acid and jasmonate-ethylene signaling pathways modulates defense gene expression and disease resistance in Arabidopsis. Plant Cell 16: 3460–3479

Berrocal-Lobo M, Molina A, Solano R (2002) Constitutive expression of ETHYLENE-RESPONSE-FACTOR1 in Arabidopsis confers resistance to several necrotrophic fungi. Plant J 29: 23–32

Binkert M, Kozma-Bognar L, Terecskei K, De Veylder L, Nagy F, Ulm R (2014) UV-B-responsive association of the Arabidopsis bZIP transcription factor ELONGATED HYPOCOTYL5 with target genes, including its own promoter. The Plant Cell 26: 4200-4213

Cao WH, Liu J, He XJ, Mu RL, Zhou HL, Chen SY, Zhang JS (2007) Modulation of ethylene responses affects plant salt-stress responses. Plant Physiol 143: 707–719

Cao YR, Chen SY, Zhang JS (2008) Ethylene signaling regulates salt stress response: an overview. Plant Signal Behav 3: 761–763

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阿拉伯芥BASIC LEUCINE-ZIPPER 6 (bZIP6) 參與在茉莉酸和乙烯的訊息傳遞路徑去調控ETHYLENE RESPONSE FACTOR 1(ERF1)的基因表現

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