植物熱休克反應之分子基礎研究-熱休克訊息傳遞相關分子之鑑定與氧化逆境交叉路徑之探討

Molecular Bases of the Heat Shock Response in Plants- Identification of Elements Involved in Heat Shock Transduction Pathway and in the Cross Talk between Heat Shock and Oxidative Stress

吳慧珍(Hui-Chen Wu)

指導教授 ： Florence Vignols ； 靳宗洛

國立臺灣大學/生命科學院/植物科學研究所/博士(2010年)

https://doi.org/10.6342/NTU.2010.00680

摘要

植物生活史中常遭遇環境之脅迫，如熱、乾旱及鹽害等逆境，當其改變幅度超過適合植物正常生命活動範圍時，對其生命活動則造成不利之影響。尤其以溫度增加所引起之熱逆境，為普遍存在的農業問題，導致作物產量大幅的削減。另外，熱逆境會誘導活性氧大量產生，造成植物體內氧化逆境之形成；因此，活性氧之清除也是避免細胞在熱逆境下受到傷害之重要機制。探討逆境信號在植物體內傳導途徑及基因表達調控等分子層次上之調節，可提供具潛力發展之基因工程策略，以改進植物對逆境之耐性，在理論及生產上提供重要之研究基礎與實踐意義。
博士論文第一部分，主要以水稻及大豆為模式植物探討「熱逆境誘導外源鈣離子與細胞壁果膠甲基酯酶，參與植物細胞壁重建及訊息傳導之調控」。植物能大量累積低分子量熱休克蛋白質(small HSP, sHSP)，扮演著抗熱逆境之重要角色；然本研究發現植物在熱休克 (heat shock, HS) 誘導下，藉由細胞壁果膠甲基酯酶 (pectin methylesterase, PME) 活性之調節，及激發細胞壁結構性鈣之移動，一面參與細胞壁之重建，增強細胞壁結構及細胞間之黏結作用；另一面，誘導外源鈣離子進入細胞質中，提高鈣訊號之震盪幅度及頻率，由鈣調蛋白 (calmodium, CaM) 接收並將信息傳達至下游，誘導低分子量熱休克蛋白質之表現，而提升植物抗熱逆境之能力。
論文第二部分以阿拉伯芥為模式植物，探討「硫氧還蛋白與分子伴護蛋白系統之功能性基因體與蛋白質體之交互作用」。植物中特有的類硫氧還蛋白(thioredoxin-like)稱Tetratricoredoxin (TDX)，兼具氧化還原活性中心及分子伴護之特性。利用酵母菌雙雜交系統 (Y2H) 及雙分子螢光互補技術 (BiFC)，在活细胞內證實TDX與阿拉伯芥高分子量熱休克蛋白質70 (HSP70) 發生交互作用，推測TDX能夠穩定HSP70與基質之結合，間接參與變性蛋白構型及活性之恢復。並且TDX受氧化逆境之誘導，轉移並累積至細胞核中，推測其功能與氧化逆境訊息之傳遞相關；藉由TDX基因缺失突變株，對氧化及熱逆境敏感性下降之外表型，並提高逆境相關基因之表現，推測TDX位於逆境訊號路徑之上游，扮演著訊號接受的角色，此過程可能與HSP70間之交互作用有關。此研究首次發現植物硫氧還蛋白在氧化與熱逆境中，同時參與訊息傳遞及伴護蛋白之功能，對於蛋白質在逆境間之交互作用有實質的貢獻。

關鍵字

二價鈣離子 ； 分子伴護蛋白 ； 細胞壁果膠甲基酯酶氧化逆境 ； 硫氧還蛋白 ； 鈣調蛋白 ； 熱休克蛋白質 ； 熱逆境

並列摘要

While being unable to escape their lands, plants are continuously submitted to the modifications of their environment, and need to adjust proper physiological processes in response to various stimuli. During this work, I devoted my studies on two major stresses affecting plant development, heat shock (HS) and oxidative stresses (OS), focusing on key elements in these pathways (HS chaperons and HS-related thioredoxins) in order to bring news elements of knowledge and interconnexion of these pathways.
Using rice and soybean as mono- and dicotyledonous plant systems, I show how HS leads to calcium release from plant cell apoplast to the cytosol in a typical “calcium signature”, conferring cell wall rigidity and enhancing HS signaling pathway. I also identify pectin methylesterase (PME) as required in this pathway for cell wall remodeling and plasma membrane integrity. I further investigate how plant sense temperature increases and how they transmit the HS signal to downstream elements. Using systematic analyses of calmodulin (CaM) and small heat shock protein (sHSP) gene expression, I identify one CaM as a coordinator of HS response, which I characterize as involving specific cytosolic/nuclear isoforms of the sHSP family.
I latter perform the molecular analysis of TDX, a thioredoxin suspected to be involved in heat shock response. I show that TDX interacts with nucleo-cytoplasmic members of the HSP70 family in a redox dependent manner, both HS and OS inducing its nuclear relocation, and that TDX is required for both acquired thermotolerance and OS signaling.
I finally discuss the data brought by this work and propose models with cross-talks between HS and OS signaling.

並列關鍵字

Calcium ； Chaperone ； Calmodium ； Heat shock proteins ； Oxidative stress ； Pectin methylesterase ； Tetratricoredoxin

參考文獻

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