Nucleotide sequence of a cDNA clone encoding 1-aminocyclopropane-1-carboxylic acid synthase from broccoli.

Ethylene regulates many processes in plant growth, development, and senescence. Two enzymes, ACC synthase and ACC oxidase, are responsible for ethylene biosynthesis from its precursor S-adenosyl Met (Yang and Hoffman, 1984), and blocking the synthesis of these enzymes in antisense RNA experiments results in reduced ethylene synthesis (Hamilton et al., 1990; Oeller et al., 1991). Functional identification of cDNA clones encoding ACC synthase has been achieved by expression studies in Escherichia coli and yeast and by sequence comparisons to peptides purified from ACC synthase protein (Sato and Theologis, 1989; Yip et al., 1990; Dong et al., 1991). Broccoli florets are composed of both vegetative and floral tissues that senesce rapidly after harvest. The most visible change is Chl degradation in the sepals, which begins within 48 h of harvest (tissues held in darkness at ZO'C), and is associated with increases in ACC oxidase mRNA and ethylene synthesis from within the florets (Tian et al., 1994; B. J. Pogson, unpublished data). We report the sequence of a cDNA clone (pbroc3, Table I) isolated from a cDNA library in AZapII (Stratagene) constructed from poly(A+) " A from broccoli florets held at 2OoC in darkness for 48 h after harvest. The cDNA is 1734 bp long and encodes an open reading frame of 481 amino acids. The nucleotide and predicted amino acid sequences show high identity with ACC synthase from tomato (LEACC2G) and apple (pAASZ), respectively (Dong et al., 1991; Rottmann et al., 1991). T h e predicted peptides of the broccoli, tomato, and apple clones were aligned (Devereux et al., 1984), and regions of high identity were found to correspond to conserved regions of other ACC synthases (Park et al., 1992). The pbroc3 cDNA contained a region with 83% amino acid identity (100% similarity based on conservative changes) with the peptide sequence identified as the active site of ACC synthase (Yip et al., 1990).