Molecular processes of cell wall construction mediated by endo-xyloglucan transferase

根据我们从赤豆上胚轴质外体液中纯化的EXT蛋白的氨基酸序列信息，我们从赤豆和拟南芥等6种植物中克隆了EXT-cDNA。EXTs的氨基酸序列在6种植物中高度保守，表明该酶在高等植物中普遍存在。RNA凝胶印迹分析表明，EXT-mRNA广泛转录在生长中的V.angularis和A.thaliana茎，特别是在表现出高活性的细胞壁构建的组织。赤霉素和吲哚-3-乙酸显著提高了从V.angularis幼苗上胚轴切下的组织切片中EXT-mRNA的水平。此外，50 mM蔗糖显着增加了在茎组织中的EXT-mRNA水平。以拟南芥ext-cDNA克隆为分子探针，我们分离到了一个含有拟南芥ext基因的基因组DNA克隆。对该基因启动子区的序列分析表明，存在一个类似于谷物淀粉酶基因中发现的“赤霉酸反应元件”（GARE）的核苷酸序列。

Besed on the amino acid sequence information of the purified EXT portein, which we had purified from apoplastic fluid of epicotyls of Vigna angularis, we have cloned EXT-cDNAs from 6 plant species including V.angularis and Arabidopsis thaliana. The amino acid sequence of the EXTs are highly conserved in the six species, suggesting that this enzyme is ubiquitous among higher plants. RNA gel blot analyzes have revealed that EXT-mRNA is extensively transcribed in growing stem of V.angularis and A.thaliana, particularly in tissues which exhibit high activities for the cell wall construction. Gibberellic acid and indole-3-acetic acid substantially increased EXT-mRNA levels in tissue sections excised from epicotyls of V.angularis seedlings. Also, 50 mM sucrose significantly increased the EXT-mRNA levels in the stem tissue. Using A.thaliana EXT-cDNA clone as a molecular probe, we have isolated a genomic DNA clone containing the ext gene of A.thaliana. Sequence analysis of the promoter region of the gene has revealed the presence of a nucleotide sequence that resembles "gibberellic acid responsive element" (GARE) found in cereal amylase genes.