Altered Gene Expression, Leaf Senescence, and Fruit Ripening by Inhibiting Ethylene Synthesis with EFE-Antisense Genes

Abstract

Genes involved in changes in colour, flavour, texture, and aroma are expressed during ripening of fleshy fruits. In tomato, and other climacteric fruits, ethylene synthesised at the outset of ripening is involved in regulating expression of some of these ripening genes. Many ripening-related mRNAs have been cloned and several, including polygalacturonase, pectinesterase, ACC synthase, ethyleneforming enzyme (EFE), and phytoene synthase have been identified and sequenced (for review see Gray et al., 1992). We have used an antisense-RNA strategy, developed originally for the polygalacturonase gene (Smith et al., 1988, 1990) to identify cDNA and genomic clones for EFE (Hamilton et al., 1990, 1991). Reducing EFE expression in transgenic tomatoes inhibits ethylene synthesis and slows ripening and over-ripening of fruits. Ripening can be accelerated by adding ethylene to detached fruit. In the EFEantisense plants, leaf senescence is delayed and wound-ethylene synthesis is also reduced. Inhibiting ethylene synthesis caused a reduction in carotenoid production in fruit, which was associated with reduced expression of phytoene synthase required for the production of β-carotene and lycopene (Bird et al., 1991). A naturally occurring tomato mutant deficient in carotenoid production has been characterised and shown to contain a mutation in the phytoene synthesis gene expressed in fruit. Constitutive over-expression of a wild-type sequence in mutant transgenic plants restores carotenoid production in ripening tomatoes and leads to unscheduled carotenoid synthesis in other cell types. The potential for using sense and antisense genes to identify and assign functions to ripening genes and to alter the physiology and biochemistry of fruits is discussed.