Fruit composition

Soluble solids content of tomato fruit is a major determinant of fruit quality, particularly for processing tomatoes, and the soluble solids are comprised predominantly of soluble sugars. Thus, approaches to altering the composition of ripe fruit using transgenic strategies have focused on carbohydrate composition. One of the first attempts in tomato was to suppress the expression of acid invertase in ripe fruit in order to increase sucrose levels. Other transgenic tomato lines have been designed to alter the ratio of the monosaccharides, glucose and fructose, in the fruit.

Acid invertase and sucrose synthase
Although tomatoes transport sucrose in the phloem, tomato fruit typically have very low levels of sucrose and approximately equal ratios of the hexose sugars, glucose and fructose. Interestingly, some wild relatives of tomato accumulate primarily sucrose in their fruit and these fruit have very high levels of total soluble sugars. Introgression of the locus controlling sucrose accumulation (succor) from the wild relative, L. chmielewskii, resulted in smaller fruit with increased soluble sugar levels (Chetela et al., 1995). Because the sucr locus from L.chmielewskii was determined to encode an inactive allele of acid invertase, it was reasoned that the same trait could be produced by transgenic suppression of invertase expression. Constitutive expression of an antisense gene encoding tomato soluble acid invertase resulted in tomato fruit with an increased concentration of sucrose and decreased concentrations of the hexoses, fructose and glucose (Klann et al., 1996). Fruit from the sucrose accumulating transgenic plants were approximately 30% smaller, presumably due to the osmotic effects of sucrose accumulation as compared to the hexose-accumulating non-transgenic control plants. Many of the characteristics of the transgenic plants with reduced invertase expression were similar to sucrose accumulating lines of tomato that had been derived by introgression sucr locus from the L. chmielewskii. However, transgenic plants engineered using the E8 promoter (Deikman et al., 1992) to suppress invertase gene expression only in ripening fruit, remained hexose accumulators. This suggested that expression of the invertase gene regulates sucrose to hexose conversion early in fruit development, before the developmental timing of expression specified by the E8 promoter.

Sucrose synthase has also been a target for genetic modification with the goal of enhancing sink ‘strength’ by increasing the capacity to metabolize imported sucrose. However, the fruit-specific antisense suppression of sucrose synthase did not produce any changes in the accumulation of starch or sugars in the fruit tissues even though the transient increase in sucrose synthase expression normally observed in early in fruit development was suppressed (Chengappa et al., 1999). Similar transgenic plants with suppressed expression of a fruit specific sucrose synthase also exhibited no change in hexose or starch accumulation (D’Aoust et al., 1999b). However, fruit set (e.g. number of fruit) on these plants was diminished and the sucrose unloading capacity of young fruit was significantly reduced.

Additional transgenic strategies have been employed to specifically alter source-sink relations in tomato by ectopic expression of sucrose phosphate synthase. When the Zea maize sucrose phosphate synthase gene regulated by the rubisco promoter was expressed in transgenic tomato foliar tissue sucrose partitioning was increased and this reduced limitations of photosynthesis (Micallef et al., 1995). Sucrose unloading in fruit also was increased by the over-expression of sucrose phosphate synthase (Nguyen-Quoc, et al. 1999).

Hexokinase and fructokinases
Over-expression of hexokinase in transgenic tomato demonstrated the regulatory role of this enzyme in photosynthetic tissues, particularly affecting senescence. However, in fruit from plants over-expressing an Arabidopsis hexokinase gene the quantity of starch in young fruit and of hexose in ripe fruit was reduced (Dai et al., 1999).

Because fructose is almost twice as sweet as glucose, modification of fructokinase expression in ripening tomato fruit has been attempted in an effort to increase the ratio of fructose and glucose and, thus, enhance fruit ‘sweetness’. Two genes encoding fructokinase are expressed in tomato fruit Frk1, and Frk2 (Kanayama et al., 1998). Frk2 expression is correlated with periods of starch accumulation, and Frk1 is expressed ubiquitously, although less abundantly. The potentially complex regulation of both the expression and activity of each of these fructokinase isoforms may confound attempts to increase fructose concentrations in fruit from transgenic plants with antisense genes for either or both fructokinases.