【作者】 张磊；

【导师】 高学文； 邵敏；

【作者基本信息】 南京农业大学 ， 植物病理学， 2011， 博士

【摘要】 干旱是粮食生产和粮食安全面临的最严重的影响因素,在缺乏农业用水的地区尤其如此。据估计,中国每年由于干旱造成的国民经济损失高达250亿美元。全球变暖和干旱发生频率的增加,使形势更加严峻。从1981年到2002年,小麦、玉米和大麦这三种作物每年由于干旱造成的产量损失达四千万吨,价值50亿美元。水稻是世界上最重要的粮食作物,很多人以大米为生,大米是他们的直接能量来源。据估计,水稻生产耗费了中国总用水量的大约一半。然而,干旱胁迫仍然是水稻生长最严重的制约因素,最主要是由于降水模式的年际变化和水稻生长季节降水量分布不均造成的。总之,面临全球的水资源短缺,除了改善灌溉措施外,提高作物耐旱性在提高作物产量上也具有巨大的潜力。植物病原革兰氏阴性细菌三型泌出通道（Type III protein secretion system, TTSS）分泌的Harpin能普遍地激发病原菌在寄主植物上的毒性,诱导在非寄主植物上的过敏性细胞死亡（hypersensitive cell death, HCD）,并激发多种植物反应。外源喷施Harpin能够激发SA,诱导植物SAR,也能激发JA、ET介导的防卫反应通路。Harpin编码基因的超表达增强了烟草、水稻、油菜和棉花等作物的抗病性。Erwinia amylovora分泌的Harpin蛋白HrpNEa激活ABI2依赖的ABA信号通路,诱导拟南芥的耐旱性。然而,至今仍不知道内源表达Harpin编码基因是否也能增强植物耐旱性。邵等（2008）曾报道,超表达Harpin编码基因hrf1增强了水稻对稻瘟病的非小种特异性抗性。本研究证实,异源表达Harpin编码基因hrf1能激发水稻ABA信号通路,增强水稻植物耐旱性。内源表达hrf1诱导水稻植株ABA浓度的升高,促进气孔关闭。转基因系的保水能力显著增强,脯氨酸和可溶性糖水平显著升高,抗氧化胁迫和清除活性氧的能力显著增强,OsLEA3-1、OsP5CS、Mn-SOD和NM₀01074345等四个干旱相关基因在干旱条件下的表达水平比对照明显上升。这些结果证明,hrf1基因赋予了转基因系对干旱的抗性,表明Harpin蛋白为农作物耐旱性改良提供了新的契机。本研究采用57k水稻cDNA芯片分析了超表达hrf1基因水稻植株NJH12的表达谱,共找到了225个差异表达基因,其中包括多个与抗病性和耐旱性相关的基因,如防卫反应相关基因、抗逆信号传导相关基因、细胞程序性死亡相关基因等。另外,在NJH12的表达谱中,还有一些富含亮氨酸的蛋白激酶和MAPK等受体蛋白激酶编码基因和能量代谢相关基因的表达量也出现了上调。通过对这些差异表达基因的分析发现,异源表达hrf1基因在水稻中调控的耐旱性通路和抗病性通路可能存在交叉调控。更多还原

【Abstract】 Drought is the most devastating factor that menaces food production and food security, especially in areas with inadequate agricultural water resources. The estimated annual loss from the national economy caused by drought alone exceeds $25 billion in China. Global warming and increasingly frequent occurrence of drought exacerbate the situation. Warming since 1981 has resulted in annual combined losses of these three crops, wheat, maize and barley, representing roughly 40 Mt or $5 billion per year, as of 2002. Globally, rice is the most important crop in terms of the number of people dependent on it as a direct source of calories. It has been estimated that rice production consumes about half of the total water consumption in China. Drought stress is still the most important constraint in rice production, mostly due to annual variation in the rainfall patterns and uneven distribution of rainfall in the rice growing season. Consequently, with the global shortage of water, enhancing the drought tolerance of crops has a potentially huge impact on annual productivity, in addition to improved water management practices.Harpin proteins, which are secreted through Type III protein secretion system of Gram-negative plant pathogenic bacteria, generally affect virulence in host plants and induce hypersensitive cell death in nonhost plants and elicit multiple plant responses. Exogenous applications of Harpins induce SAR in plants by the activation of defense pathways mediated by salicylic acid, jasmonic acid or ethylene. Transformation of Harpin-encoding genes improves disease resistance in tobacco, rice, rape and cotton, a, a Harpin protein secreted by Erwinia amylovora, induces drought tolerance in Arabidopsis by activating the AB12-dependent ABA signaling. So far it is still unknown whether genetic transformation of Harpin-encoding genes improves plant tolerance to drought. Shao et al. （2008） have reported that expression of hrfl, a Harpin-encoding gene, in rice conferred durable nonspecific resistance to Magnaporthe grisea. In this study, the results showed that overexpression of a Harpin-encoding gene hrfl in rice increased drought tolerance through abscisic acid （ABA） signaling, hrfl-expression induces the increase of ABA content and promotes stomata closure in rice. The hrfl transgenic rice lines exhibited significantly increase in water retention ability, levels of free proline （Pro） and soluble sugars, tolerance to oxidative stress, the reactive oxygen species （ROS）-scavenging ability, and expression levels of four stress-related genes, OsLEA3-1, OsP5CS, Mn-SOD and NM₀01074345, under drought stress. Our studies confirmed that hrfl conferred transgenic crops enhanced tolerance to drought stress. These results suggest that Harpins may offer new opportunities for generating drought resistance in other crops.The transcriptome profile of the transgenic rice line NJH12 expressing hrfl from Xanthomonas oryzae pv. oryzae was analysed using a customized 57 k rice cDNA microarray. A total of 225 cDNA transcripts were differentially expressed in the transgenic line NJH12, in which several differentially expressed genes were related to the rice biotic and abiotic stress, including the transcripts related to defence response, the transcripts related to signal transduction, and the transcripts related to programmed cell death. Furthermore, transcripts encoding various recepter kinases, such as leucine-rich protein kinases, and mitogen-activated protein kinases were up-regulated in the transgenic line NJH12 and expression of transcripts related to the energy producing and consuming pathway was also increased. By analysing the putative functions of these transcripts, it is speculated that the signaling between drought tolerance and disease resistance in hrf1-overexpressing rice might converge on some pathways.更多还原