水稻在淹水逆境中Sub1A-1下游基因的轉錄調控

Abstract

Sub1A-1是一個乙烯反應轉錄因子(ethylene response transcription factor, ERF)，當處於淹水逆境之時，它會受到乙烯的增量調控(up-regulated)並且幫助植物存活下來。帶有Sub1A-1對偶基因(Sub1A allele)的基因型之水稻，例如FR13A和Pokkali (這兩個品種皆為秈稻)能夠忍受長時間的淹水逆境；IR29和IR64雖然也同為秈稻但卻帶有Sub1A-2且無法抵抗淹水逆境；另外一個秈稻品種，Swarna不具有Sub1A allele且不耐淹水逆境。本實驗希望能夠找到Sub1A-1下游調控的基因、其可能的調控方式以及植物生理適應的變化。透過FR13A、IR29和Pokkali的基因微陣列分析(Microarray)並且用Swarna、Swarna(Sub1)、IR64、IR64(Sub1)做定量即時聚合酶鏈鎖反應（Quantitative real time polymerase chain reaction）進行檢測，我們找出了27個基因會受到Sub1A-1誘導，其中三個基因參與糖解作用，它們分別是Aldolase C-1、GAPC和PPDK。另外，我們也找出了兩個未知功能的基因會受到Sub1A-1抑制。 從電泳速率變化分析實驗(Electrophoretic Mobility Shift Assay, EMSA)可以發現，Sub1A-1可能藉由和GCC box結合來直接調控Aldolase C-1、GAPC和PPDK的表現。在生理適應方面，ATP隨著淹水逆境時間增加會逐漸減少，但在FR13A和IR64(Sub1)是相對多於IR29和IR64；當比較碳水化合物於淹水逆境的含量時，蔗糖、葡萄糖和果糖都會隨著淹水時間增加而減少，蔗糖在帶有Sub1A-1基因的品系中有較高的含量。這些結果顯示，Sub1A-1藉由調控其下游基因使植物保留較多的ATP和蔗糖，並對淹水逆境有較高的耐受性。

Sub1A-1, an ethylene response transcription factor (ERF), is up-regulated by ethylene and helps plants' survival under submergence. Rice whose genotype carrying Sub1A-1 allele like FR13A and Pokkali (both of them are O. sativa ssp. indica cultivar) can tolerate prolonged submergence. The intolerant cultivars such as IR29 and IR64 possess Sub1A-2, and Swarna does not have Sub1A allele. However, the downstream genes under Sub1A-1 regulation remain to be investigated. By using FR13A, IR29 and Pokkali to do microarray analysis and further confirmed with Swarna, Swarna(Sub1), IR64 and IR64(Sub1), we validated several genes that were under Sub1A-1 regulation. There are 27 genes that were up-regulated by Sub1A-1 under submergence, including 9 transcription factors, 12 functional genes and 6 genes with unknown functions. On the other hand, there are only two genes with unknown function that were down-regulated by Sub1A-1. Among the genes that were up-regulated by Sub1A-1, 3 genes were involved in the glycolytic pathway, including Aldolase C-1, Glyceraldehyde-3-phosphate dehydrogenase (GAPC) and Pyruvate phosphate dikinase (PPDK). From the Electrophoretic mobility shift assay (EMSA) experiments, Sub1A-1 might directly regulate its downstream targets by binding to the GCC box on their promoters. The genes that are regulated by Sub1A-1 also contribute to the energy and metabolic adaption in plants. The ATP levels in Sub1 haplotypes such as FR13A and IR64(Sub1) were relative higher than IR29 and IR64. Sucrose was more abundant in IR64(Sub1) than IR64 after submergence. This indicated that the energy and carbohydrate reservation were important for survival under submergence.