Down-regulation of tomato β-galactosidase 4 results in decreased fruit softening

Transcript abundance of the gene encoding beta-galactosidase II, a beta-galactosidase/exo-galactanase (EC 3.2.1.23) present during tomato (Lycopersicon esculentum) fruit ripening, was suppressed by expression of an antisense tomato beta-galactosidase 4 (TBG4) cDNA driven by the cauliflower mosaic virus 35S promoter. RNA gel-blot analysis was used to evaluate TBG4 mRNA levels in transgenic fruit. All of the antisense lines had attenuated TBG4 mRNA levels in turning stage fruit; however, TBG4 mRNA suppression was unstable, and mRNA levels varied in red-ripe fruit among the lines. Suppression of TBG4 mRNA levels in antisense fruit was correlated with a reduction in extractable exo-galactanase activity against a lupin galactan. All of the antisense lines had reduced free galactose levels at mature green stage 4, but levels comparable with controls during ripening. Total cell wall galactosyl contents in the antisense fruit were not significantly different from control fruit. Whole-fruit firmness was measured using a texture analyzer and the means of the peak force measurements for four of six antisense lines were significantly higher than control fruit. One antisense line had red-ripe fruit that were 40% firmer than controls. Fruit from this antisense line also had the lowest TBG4 mRNA and exo-galactanase levels and the highest wall galactosyl content during the early stages of ripening, implicating an involvement of this gene product in cell wall modification leading to fruit softening.