Department of Botany, Graduate School of Science, Kyoto University, Kyoto, Japan.
Faculty of Agriculture, Ryukoku University, Otsu, Japan.
J Exp Bot. 2021 Feb 24;72(4):1260-1270. doi: 10.1093/jxb/eraa519.
Hydathodes are typically found at leaf teeth in vascular plants and are involved in water release to the outside. Although morphological and physiological analysis of hydathodes has been performed in various plants, little is known about the genes involved in hydathode function. In this study, we performed fluorescent protein-based imaging and tissue-specific RNA-seq analysis in Arabidopsis hydathodes. We used the enhancer trap line E325, which has been reported to express green fluorescent protein (GFP) at its hydathodes. We found that E325-GFP was expressed in small cells found inside the hydathodes (named E cells) that were distributed between the water pores and xylem ends. No fluorescence of the phloem markers pSUC2:GFP and pSEOR1:SEOR1-YFP was observed in the hydathodes. These observations indicate that Arabidopsis hydathodes are composed of three major components: water pores, xylem ends, and E cells. In addition, we performed transcriptome analysis of the hydathode using the E325-GFP line. Microsamples were collected from GFP-positive or -negative regions of E325 leaf margins with a needle-based device (~130 µm in diameter). RNA-seq was performed with each single microsample using a high-throughput library preparation method called Lasy-Seq. We identified 72 differentially expressed genes. Among them, 68 genes showed significantly higher and four genes showed significantly lower expression in the hydathode. Our results provide new insights into the molecular basis for hydathode physiology and development.
水孔通常位于维管植物的叶片齿上，参与水向外部的释放。尽管已经在各种植物中对水孔进行了形态和生理分析，但对参与水孔功能的基因知之甚少。在这项研究中，我们在拟南芥水孔中进行了荧光蛋白的成像和组织特异性 RNA-seq 分析。我们使用了已报道在其水孔中表达绿色荧光蛋白（GFP）的增强子捕获系 E325。我们发现 E325-GFP 在水孔内的小细胞（称为 E 细胞）中表达，这些细胞分布在水孔和木质部末端之间。在水孔中未观察到韧皮部标记物 pSUC2:GFP 和 pSEOR1:SEOR1-YFP 的荧光。这些观察结果表明，拟南芥水孔由三个主要组成部分组成：水孔、木质部末端和 E 细胞。此外，我们使用 E325-GFP 系对水孔进行了转录组分析。使用基于针的装置（直径约 130 µm）从小叶片边缘的 GFP 阳性或阴性区域采集微样本。使用称为 Lasy-Seq 的高通量文库制备方法对每个单一样本进行 RNA-seq。我们鉴定了 72 个差异表达基因。其中，有 68 个基因在水孔中表达显著升高，有四个基因表达显著降低。我们的结果为水孔生理学和发育的分子基础提供了新的见解。
