探究SlWRKY15轉錄因子在番茄基礎防禦反應扮演的角色

Abstract

WRKY為植物重要轉錄因子 (transcription factor)，常以基因族 (gene family) 型式存在於植物體中，參與多種生物 (biotic) 及非生物 (abiotic) 逆境反應之分子調控，故為非常重要的轉錄調控因子。番茄為世界性重要的經濟作物，瞭解番茄因應病原菌而啟動的基因表現以及其後續所引發的各種抗病反應為當前重要的議題，而WRKY轉錄因子在這樣的生物逆境反應很可能也扮演舉足輕重的角色，值得我們深入探究。本實驗室先前以番茄生物晶片所進行的研究發現，某些WRKY基因在番茄被疫病菌 (Phytophthora parasitica) 感染後會被誘導表現，另外以semi-quantitative reverse transcriptase PCR所做的分析也發現特定之WRKY 基因的表現確實會因應疫病菌侵染而啟動，將其命名為SlWRKY15。SlWRKY15屬於WRKY IId族群，並且位於細胞核內。基因表現分析發現，SlWRKY15會被疫病菌誘導表現，但不會被青枯病菌 (Ralstonia solnacearum)及細菌性斑點病菌(Pseudomonas syringae)誘導表現；此外，SlWRKY15之表現受到植物荷爾蒙水楊酸以及乙烯的調控。以agroinfection在番茄植株大量表現SlWRKY15基因能使植株在疫病菌及青枯病菌感染下呈現較佳的耐病性。相對而言，利用VIGS (virus-induced gene silencing) 抑制SlWRKY15基因表現，則增加番茄對上述兩種病原菌的感病性。由於大量表現SlWRKY15所引發的耐病並無專一性，顯示SlWRKY15基因可能在番茄的先天性防禦反應扮演重要的角色；實驗也顯示疫病菌PAMP，包括parA1及pep13，能誘導菸草中SlWRKY15之同源性基因的表現。進一步的研究發現，SlWRKY15並不能與LeMPK2 及LeMPK3等PTI調控相關蛋白交互作用，但當SlWRKY15大量表現時，會降低部分抗病相關基因的表現，這些結果顯示SlWRKY15透過複雜的調控引起番茄的耐病性，其詳細分子機制有待進一步釐清。

WRKY transcription factors (WRKYs) are encoded by large gene families in all higher plants. They are known to play pivotal roles in plant-specific processes, including plant responses to biotic and abiotic stresses. While analyzing tomato genes which are differentially expressed in response to infection by Phytophthora parasitica, we found a gene that is up-regulated and encodes a putative WRKY transcription factor, herein named SlWRKY15. The predicted amino acid sequence of SlWRKY15 contains a WRKY domain, a zinc-finger motif, nuclear localization signal, and a calmodulin binding domain. Analysis by semi-quantitative reverse transcriptase polymerase chain reaction indicated that SlWRKY15 is expressed in all parts of the tomato plant, with the messages in the flowers and stems being more abundant than those in other parts. Furthermore, expression of SlWRKY15 is induced following treatment of the plants with salicylic acid and ethylene. As well, expression of the homologs of SlWRKY15 in tobacco is induced by PAMPs from Phytophthora, including parA1 and pep13. To investigate its function, SlWRKY15 overexpressing plants were conducted by agroinfection and then inoculated the plants with P. parasitica and Ralstonia solnacearum. The results indicated that disease symptoms caused by either P. parasitica or R. solnacearum on plants overexpressing SlWRKY15 were less severe than those on the control plants. In contrast, down regulation of SlWRKY15 by virus-induced gene silencing enhanced the plant susceptibility s to these two pathogens. In plant overexpressing SlWRKY15, the expression of defense related genes (ACO1, PAL5, and PIN2) was down regulated. These results suggest a potential role of SlWRKY15 in the plant basal defense response, but how is it involved in the regulation of defense response awaits further investigation.