阿拉伯芥轉錄因子 HSFA1a/b/d 及 HSFA2 對一群熱逆境記憶效應基因調控之研究

Abstract

植物有許多應對環境逆境的策略，其中之一是對逆境的記憶能力，過去的研究發現，植物可以對不同的環境刺激進行記憶，然而，植物是否對熱逆境也有記憶則並不清楚。利用微陣列技術分析基因表現後發現，阿拉伯芥中有一小群基因受到反覆的熱處理後，其基因表達量會與只受到單一熱處理後有顯著差異，此現象進一步被及時定量聚合酶連鎖技術確認，我們把這個現象稱為熱逆境記憶反應，分析後，發現其中的維生素C過氧化酶(APX2)及肌醇磷酸合成酶(MIPS2)，其基因表達量在HSFA2過量表現株中上升，而在HSFA1s四重剔除株中下降，於是我們試圖去闡明HSFA2與HSFA1s跟熱逆境記憶的關係，比較野生株及HSFA2剔除株後，發現HSFA2剔除株中的熱逆境記憶效應完全消失，證實了HSFA2在記憶效應的重要性。然而，ROF1及HSP90兩個已知會結合HSFA2的蛋白質，皆不影響熱逆境記憶效應。利用HSFA1s的三重及四重突變株實驗，證實HSFA1a/d也對熱逆境記憶效應十分重要。儘管過去研究指出HSFs會被SUMO化，但是比較E3 SUMO ligase突變株siz1後，並沒有發現其會影響熱逆境記憶效應。最後，存活率測試發現熱逆境記憶效應能極大的提升植物面對致死熱逆境之下的存活率，證實熱逆境記憶效應確實有其重要的生理功能。

Plants have many strategies to cope with environmental stresses, and stress memory is one of them. Previous studies show that plants can respond to and remember various stimuli. However, it was not clear how plants remember the experience of heat stress. In this study, the results of microarray analysis showed that a small group of genes in Arabidopsis were induced by heat treatment to higher or lower expression level in seedlings that had a prior exposure to high temperature three days ago than those that had not. This phenomenon was called heat stress memory response (HSMR) herein. The HSMR of two of these genes, ascorbate peroxidase 2 (APX2) and myo-inositol phosphate synthase 2 (MIPS2) were further confirmed by quantitative RT-PCR. Since APX2 and MIPS2 were shown to be up-regulated by overexpression of heat shock transcription factor A2 (HSFA2) and down-regulated in heat shock factor A1 quadruple knockout (KO) mutant, the functions of HSFA2 and HSFA1 in HSMR were investigated. In the HSFA2 KO mutant, HSMR was greatly diminished, suggesting that HSFA2 plays an important role in HSMR. However, rotamase FK506-binding proteins and heat shock protein 90 that bind to HSFA2 were dispensable for HSMR. Studies on the quadruple and triple KO mutants of the four HFSA1 genes suggest that HSFA1a and HSFA1d were the key players of HSMR. In eukaryotes, sumoylation regulates the activities of HSFs. However, HSMR of the T-DNA KO mutant of SIZ1, which encodes a small ubiquitin-like modifier E3 ligase, was not significantly altered. Finally, thermotolerance assay showed that HSMR was associated with significantly enhanced survival rates of seedlings under severe high temperature, which underscores the importance of this type of stress memory.