一個新穎的植物特有基因BIG GRAIN 2透過促進細胞增殖調控水稻穀粒大小

Abstract

世界作物產率即將達到飽和，因應世界人口增加的食物需求，高產量作物的育種日漸重要。植物基因工程提供一個有效提高作物產量的途徑。利用正向遺傳學的方法，我們從台灣水稻T-DNA突變庫找到一個表現出大穀粒性狀的T-DNA活化表現株BG2act。插入點上下游有數個基因被T-DNA上的強化子活化表現。BIG GRAIN 2 (BG2)轉譯一個新穎的植物特有未知蛋白，胺基酸序列在開花植物裡具有保守性，確認其活化表現與大穀粒性狀有關。以玉米ubiquitin啟動子大量表現BG2，轉殖株呈現多效性的影響，包含植株高度、花穗長度、穀粒大小與分糵角度的增加。產量分析結果顯示，大量表現BG2亦增加了穀粒重量，總產量卻輕微減少，指出到處表現BG2對於增產有些負面影響。組織學分析顯示，BG2大量表現呈現的大穀粒是因為總細胞數目的增加而非細胞大小改變的影響，指出BG2可能參與細胞週期的調控。降低BG2表現的植株性狀近似於野生型，顯示BG2具有功能重複性。BG2廣泛表現於各發育時期，尤其在地上部年輕組織中表現最多。次細胞內定位分析結果顯示，BG2主要表現在近細胞膜的位置，在細胞質與細胞核內也有較低量表現。BG2的表現可被生長素、油菜素類固醇及激勃素誘導表現。生長素及油菜素類固醇生合成與訊息傳導的代表性基因在BG2大量表現植株中有增加與降低表現，幾個細胞週期調控基因也有表現增加情形。大量表現油菜素類固醇生合成訊息傳導途徑中正向調控子常引起植株高度與穀粒大小的增加，綜合以上結果我們發現BG2可能是一個新的油菜素類固醇訊息傳導途徑之正向調控子，透過促進細胞增殖控制水稻穀粒大小。BG2是否透過油菜素類固醇訊息傳導控制細胞週期以及是否涉及多種植物荷爾蒙交互條控仍待進一步的實驗證明。BG2的研究連結了油菜素類固醇等植物荷爾蒙與細胞週期對穀粒大小的調控，並可能成為一個新的分子育種標的基因。

The world’s crop productivity has reached a plateau in recent years. Breeding of high-yielding rice is crucial for meeting the food demand of increasing world population. Plant genetic engineering offers an efficient alternative for increase in crop productivity. Using the forward genetic approach, a T-DNA activation tagged mutant BG2act exhibiting big grain phenotype was identified from our T-DNA the Taiwan Rice Insertional Mutagenesis (TRIM) population. Several genes flanking the T-DNA insertion site are activated by the enhancer in the T-DNA construct. BIG GRAIN 2 (BG2), a novel gene encodes an unknown function protein which is conserved in Angiosperms, is validated responsible for the big grain phenotype. Overexpression of BG2 by maize ubiquitin promoter causes pleiotropic effects in transgenic rice, including increase in plant height, panicle length, grain size, and tiller angle. Yield analysis of BG2OX shows increase in grain weight but slightly decrease in total yield, indicating other negative influences caused by ubiquitous expression of BG2. Histological analysis results shows that cell number but not cell size accounts for the increase in grain length in BG2OX and BG2act lines, indicating that BG2 may be involved in the cell cycle regulation in grain size regulation. Knockdown expression of BG2 displays similar phenotype as wild-type plants, suggesting that BG2 has functional redundancy. BG2 is expressed ubiquitously during rice development, especially in shoot and young tissues. BG2 is mainly localized in plasma membrane and connected with the cell wall, and also localized in cytoplasm and nucleus at lower level. Expression of BG2 can be induced by auxin, brassinosteroids (BRs), and gibberellin (GA). Genes involved in biosynthesis and signaling of auxin and BR are up-regulated or down-regulated in BG2OX transgenic plant. Several genes involved in cell cycle also detected up-regulated. As overexpression of BRs biosynthesis genes and positive regulators usually lead to increase of plant height and grain size, these results indicate that BG2 may act as a novel positive regulator of BR response, increasing grain size by promoting cell proliferation. Crosstalk of hormones and other grain size control genes in grain size regulation will be further demonstrated. This study suggests that BG2 may provide a possible connection between plant hormones such as BRs and cell cycle regulation in grain size control, potentially as the target applied for molecular breeding.