LE-ACS4, a fruit ripening and wound-induced 1-aminocyclopropane-1-carboxylate synthase gene of tomato (Lycopersicon esculentum). Expression in Escherichia coli, structural characterization, expression characteristics, and phylogenetic analysis

Abstract

ACC (1-aminocyclopropane-1-carboxylic acid) synthase is the key regulatory enzyme in the biosynthetic pathway of the plant hormone ethylene and is encoded by a highly divergent multigene family in tomato (Rottmann, W. H., Peter, G. F., Oeller, P. W., Keller, J. A., Shen, N. F., Nagy, B. P., Taylor, L. P., Campbell, A. D., and Theologis, A. (1991) J. Mol. Biol. 222, 937-961). Two members of the family, LE-ACS2 and LE-ACS4, are induced during fruit ripening and upon treatment of mature green fruits with exogenous ethylene (C2H4) in a dose-dependent manner. Both genes are superinduced by wounding of pericarp tissue during various stages of ripening. The wound-induced accumulation of LE-ACS2 mRNA is more rapid and greater than that of LE-ACS4. Both mRNAs accumulate in the absence of protein synthesis, suggesting that their induction is a primary response to the inducer. The LE-ACS4 gene was isolated and structurally characterized. The function of the LE-ACS4 protein (53,509 Da, pI 5.4) was verified by expression experiments in Escherichia coli. The promoters of LE-ACS2 and LE-ACS4 contain potential cis-acting regulatory elements responsible for induction by ethylene, wounding, and anaerobiosis. In addition, elements for binding the transcriptional factors EmBP1, GBF-1, and OCSBF-1 are also present. Phylogenetic analysis of 20 ACC synthases from dicots and monocots indicate that the LE-ACS2 and LE-ACS4 proteins belong to an unique sublineage that includes an additional member of the tobacco family, NT-ACS1. The divergence of this sublineage is a relatively recent event in the evolution of ACC synthase protein.