SEQUENCE OF A CDNA CODING FOR A 1-AMINOCYCLOPROPANE-1-CARBOXYLATE OXIDASE HOMOLOG FROM APPLE FRUIT
SEQUENCE OF A CDNA CODING FOR A 1-AMINOCYCLOPROPANE-1-CARBOXYLATE OXIDASE HOMOLOG FROM APPLE FRUIT
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DOI：
10.1104/pp.98.4.1530
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发表时间：
1992-04-01
期刊：
影响因子：
7.4
通讯作者：
YANG, SF
中科院分区：
文献类型：
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作者：
DONG, JG;OLSON, D;YANG, SF
在跃变型果实中，乙烯在启动果实成熟中的重要作用已得到充分证实。在高等植物中，乙烯通过S-腺苷甲硫氨酸和ACC 2由甲硫氨酸生物合成（1）。ACC氧化酶（也称为乙烯形成酶）催化ACC氧化为乙烯，这是该生物合成途径的最后一步。与ACC合酶是限速酶的营养组织不同，果实组织中与成熟相关的乙烯产生受ACC合酶和ACC氧化酶活性的调节（4）。为了研究苹果果实成熟过程中乙烯生物合成的分子调控，必须分离和鉴定ACC合酶和ACC氧化酶基因（表I）。我们先前从苹果果实中克隆了编码苹果ACC合酶的cDNA（2）。分离了两个同源的番茄ACC氧化酶cDNA克隆，并在酵母（3）和非洲爪蟾卵母细胞（6）中进行了功能性表达。另外，从鳄梨果实（5）和香石竹花（7）中分离到两个同源序列。在比较了番茄ACC氧化酶与其他已报道的同源序列后，我们构建了两个简并寡核苷酸引物，分别对应于ACENWGF和KENAKEP的保守序列（图1中下划线部分）。从成熟苹果果实中分离mRNA，构建了3 '端加poly（dA）尾、5'端加poly（dC）尾的cDNA文库（2）。以苹果cDNA文库为模板，以poly（dC）、poly（dT）和简并寡核苷酸为引物，进行PCR扩增。对PCR产物进行测序，并将三个PCR产物之间的重叠序列组合并融合以产生命名为pAE 12的全长cDNA。pAE 12的序列长1199个碱基对，含有314个氨基酸的开放阅读框（图1）。核苷酸和推导的氨基酸序列高度同源，
In climacteric fruits, the essential role of ethylene in initi-ating fruit ripening has been well established. Ethylene is biosynthesized from methionine via S-adenosylmethionine and ACC2 in higher plants (1). ACC oxidase (also known as the ethylene-forming enzyme) catalyzes the oxidation of ACC to ethylene, the final step ofthis biosynthetic pathway. Unlike vegetative tissues, in which ACC synthase is the rate-limiting enzyme, ripening-associated ethylene production in fruit tis-sues is regulated by both ACC synthase and ACC oxidase activities (4). To study the molecular regulation of ethylene biosynthesis during apple fruit ripening, it is essential to isolate and characterize the ACC synthase and ACC oxidase genes (Table I). We previously cloned a cDNA encoding apple ACC synthase from apple fruit (2). Two homologous tomato ACC oxidase cDNA clones were isolated and functionally expressed in yeast (3) and Xenopus oocytes (6). In addition, two homol-ogous sequences were isolatedfrom avocado fruit (5) and carnation flowers (7). After comparing the tomato ACC oxidases with the other reported homologous sequences, we constructed two degenerate oligonucleotide primers, corre-sponding to the conserved sequences of ACENWGF and KFQAKEP (underlined in Fig. 1). A cDNA library made from mRNA isolated from ripe apple fruit was constructed with poly (dA) tailing at the 3'end andpoly (dC) tailing at the 5'end (2). PCR-based amplifications were carried out using the apple cDNA library as the template and poly (dC), poly (dT), and the degenerate oligonucleotides as the primers. The PCR products were sequenced, and the overlapping sequences between three PCR products were combined and fused to generate a full-length cDNA designated as pAE 12. The sequence of pAE12 is 1199 base pairs long and contains an open reading frame of 314 amino acids (Fig. 1). The nucleotide and the deduced amino acid sequences are highly