Alteration of <i>Arabidopsis SLAC1</i> promoter and its association with natural variation in drought tolerance

Natural variation for drought tolerance is a major issue in adaptation and geographic distribution of terrestrial plants. Despite the importance, little is known about the genes and molecular mechanisms that determine its naturally occurring diversity. We analyzed the intraspecific drought tolerance variation between 2 accessions of <i>Arabidopsis thaliana</i>, Columbia (Col)-0 and Wassilewskija (Ws)-2. Measurement of weight loss in detached seedlings demonstrated a clear difference between drought-tolerant Col-0 and drought-sensitive Ws-2. They also differed in their stomatal response under drought condition. Using a quantitative genetic approach, we found a significant quantitative locus on chromosome 1. Surveying in the locus, we extrapolated that the <i>SLAC1</i> gene, which is associated with stomatal closure, was likely responsible for the difference of drought tolerance. Comparison of their nucleotide and amino acid sequences revealed that there were few differences in regions encoding <i>SLAC1</i> protein but was a large deletion in <i>SLAC1</i> promoter of Ws-2. Histochemical GUS staining showed that the <i>SLAC1</i> expressed dominantly in guard cells of Col-0, but did less in guard cells of Ws-2. Quantitative PCR analysis also showed that transcript level of <i>SLAC1</i> in guard cells was higher in Col-0 than in Ws-2. The <i>SLAC1</i> transcription analyses indicate low accumulation of <i>SLAC1</i> in guard cells of Ws-2. When taken together, our results suggest that the low drought tolerance of Ws-2 was associated with the deletion of the promoter region of Ws-2 <i>SLAC1</i>.耐旱性的自然变异是陆生植物适应性进化与地理分布的关键影响因素之一。尽管其生物学意义重大，但目前对调控该自然变异的基因及分子机制仍知之甚少。本研究以2个拟南芥（Arabidopsis thaliana）种质株系Columbia（Col-0）与Wassilewskija（Ws-2）为材料，分析其种内耐旱性变异。通过对离体幼苗的失重量进行检测，发现耐旱型株系Col-0与干旱敏感型株系Ws-2之间存在显著表型差异；二者在干旱胁迫下的气孔响应也存在显著差异。本研究采用数量遗传学分析方法，在1号染色体上定位到一个显著的数量性状位点。通过对该位点区域进行筛查分析，我们推测调控气孔关闭的SLAC1基因极有可能是导致二者耐旱性差异的关键基因。对二者的核苷酸与氨基酸序列进行比对后发现，SLAC1蛋白编码区仅存在少量序列差异，但Ws-2的SLAC1基因启动子区域存在大片段缺失。组织化学GUS染色实验结果显示，Col-0的SLAC1基因在保卫细胞中优势表达，而Ws-2的SLAC1基因在保卫细胞中的表达量显著降低。实时荧光定量PCR分析结果同样证实，Col-0保卫细胞中SLAC1的转录水平显著高于Ws-2。上述SLAC1基因转录水平的分析结果均表明，Ws-2保卫细胞内SLAC1的转录本积累量较低。综合上述实验结果，本研究认为Ws-2较低的耐旱性与其SLAC1基因启动子区域的大片段缺失密切相关。