Zhu Q, Dabi T, Beeche A, Yamamoto R, Lawton M A, Lamb C
Plant Biology Laboratory, Salk Institute for Biological Studies, La Jolla, CA 92037, USA.
Plant Mol Biol. 1995 Nov;29(3):535-50. doi: 10.1007/BF00020983.
Phenylalanine ammonia-lyase (PAL) genomic sequences were isolated from a rice (Oryza sativa L.) genomic library using a PCR-amplified rice PAL DNA fragment as a probe. There is a small family of PAL genes in the rice genome. The nucleotide sequence of one PAL gene, ZB8, was determined. The ZB8 gene is 4660 bp in length and consists of two exons and one intron. It encodes a polypeptide of 710 amino acids. The transcription start site was 137 bp upstream from the translation initiation site. Rice PAL transcripts accumulated to a high level in stems, with lower levels in roots and leaves. Wounding of leaf tissues induced ZB8 PAL transcripts to a high level. In rice suspension-cultured cells treated with fungal cell wall elicitors, the ZB8 PAL transcript increased within 30 min and reached maximum levels in 1-2 h. The transcription of the ZB8 gene was investigated by fusing its promoter to the reporter gene beta-glucuronidase (GUS) and transforming the construct into rice and tobacco plants, as well as rice suspension-cultured cells. High levels of GUS activity were observed in stems, moderate levels in roots and low levels in leaves of transgenic rice and tobacco plants. Histochemical analysis indicated that in transgenic rice the promoter was active in root apical tips, lateral root initiation sites, and vascular and epidermal tissues of stems and roots. In rice flowers, high GUS activity was observed in floral shoots, receptacles, anthers and filaments, occasionally GUS activity was also detected in lemma and awn tissues. In tobacco flowers, high GUS activity was detected in the pink part of petals. Consistent with the activity of endogenous PAL transcripts, wounding of rice and tobacco leaf tissues induced GUS activity from low basal levels. Tobacco mosaic virus (TMV) infection of tobacco leaves induced GUS activity to a high level. Fungal cell wall elicitors strongly induced GUS activity and GUS transcripts to high levels in transgenic rice suspension-cultured cells. We demonstrated that the promoter of ZB8 gene is both developmentally regulated and stress-inducible.
利用PCR扩增的水稻苯丙氨酸解氨酶（PAL）DNA片段作为探针，从水稻（Oryza sativa L.）基因组文库中分离出PAL基因组序列。水稻基因组中存在一个小的PAL基因家族。测定了一个PAL基因ZB8的核苷酸序列。ZB8基因长度为4660 bp，由两个外显子和一个内含子组成。它编码一个由710个氨基酸组成的多肽。转录起始位点位于翻译起始位点上游137 bp处。水稻PAL转录本在茎中积累水平较高，在根和叶中水平较低。叶片组织受伤诱导ZB8 PAL转录本高水平积累。在用真菌细胞壁激发子处理的水稻悬浮培养细胞中，ZB8 PAL转录本在30分钟内增加，并在1-2小时内达到最高水平。通过将ZB8基因的启动子与报告基因β-葡萄糖醛酸酶（GUS）融合，并将构建体转化到水稻和烟草植株以及水稻悬浮培养细胞中，研究了ZB8基因的转录。在转基因水稻和烟草植株的茎中观察到高水平的GUS活性，在根中为中等水平，在叶中为低水平。组织化学分析表明，在转基因水稻中，启动子在根尖、侧根起始位点以及茎和根的维管组织和表皮组织中具有活性。在水稻花中，在花茎、花托、花药和花丝中观察到高GUS活性，偶尔在稃片和芒组织中也检测到GUS活性。在烟草花中，在花瓣的粉红色部分检测到高GUS活性。与内源性PAL转录本的活性一致，水稻和烟草叶片组织受伤从低基础水平诱导GUS活性。烟草花叶病毒（TMV）感染烟草叶片诱导GUS活性达到高水平。真菌细胞壁激发子在转基因水稻悬浮培养细胞中强烈诱导GUS活性和GUS转录本达到高水平。我们证明了ZB8基因的启动子在发育上受到调控且对胁迫有诱导性。
