O'Neill S D, Kumagai M H, Majumdar A, Huang N, Sutliff T D, Rodriguez R L
Department of Genetics, University of California, Davis 95616.
Mol Gen Genet. 1990 Apr;221(2):235-44. doi: 10.1007/BF00261726.
Two cDNA clones, pOS103 and pOS137, were isolated which code for distinct alpha-amylase isozymes in germinating rice seeds. Sequence analysis indicated that the clones encode polypeptides of approximately 48 kDa, both of which possess a signal peptide involved in directing secretion of the protein. Comparison of the two rice alpha-amylase amino acid sequence showed that they are 76% similar to each other, while showing 85% to 90% similarity with other cereal alpha-amylases. A comparison of eleven cereal alpha-amylases also revealed three new conserved regions (I', II', and IV') not previously identified in the animal, bacterial, and fungal alpha-amylases. Regions I' and IV' are sites for intron splicing while region II' is probably involved in calcium binding. One of the rice alpha-amylase cDNAs, pOS103, encodes a protein that has two potential N-glycosylation sites, one in the signal peptide and the other in the mature portion of the protein. The cDNA clone, pOS137, encodes an alpha-amylase with a single glycosylation site in the signal peptide, suggesting that the mature OS137 isozyme is not glycosylated. Analysis of the expression of these genes in germinating rice seeds indicated that mRNA corresponding to pOS103 and pOS137 could be detected throughout a 48 h period of seed imbibition. RNA levels, however, were dramatically stimulated by treatment of embryoless half-seeds with exogenous GA3. Our results demonstrate that at least two forms of alpha-amylase are expressed in germinating rice seeds and that the expression of these genes is regulated by the phytohormone GA3.
分离出了两个cDNA克隆，pOS103和pOS137，它们编码萌发水稻种子中不同的α-淀粉酶同工酶。序列分析表明，这些克隆编码约48 kDa的多肽，二者都具有参与指导蛋白质分泌的信号肽。两种水稻α-淀粉酶氨基酸序列的比较表明，它们彼此之间有76%的相似性，而与其他谷物α-淀粉酶的相似性为85%至90%。对11种谷物α-淀粉酶的比较还揭示了三个新的保守区域（I'、II'和IV'），这些区域在动物、细菌和真菌α-淀粉酶中未曾被鉴定出来。区域I'和IV'是内含子剪接位点，而区域II'可能参与钙结合。水稻α-淀粉酶cDNA之一，pOS103，编码一种具有两个潜在N-糖基化位点的蛋白质，一个在信号肽中，另一个在蛋白质的成熟部分。cDNA克隆pOS137编码一种在信号肽中有单个糖基化位点的α-淀粉酶，这表明成熟的OS137同工酶没有糖基化。对这些基因在萌发水稻种子中的表达分析表明，在种子吸胀的48小时内都能检测到与pOS103和pOS137相对应的mRNA。然而，用外源GA3处理无胚半种子可显著刺激RNA水平升高。我们的结果表明，至少有两种形式的α-淀粉酶在萌发的水稻种子中表达，并且这些基因的表达受植物激素GA3的调控。
