Evaluation of the role of the endo-beta-(1,4)-glucanase gene FaEG3 in strawberry fruit softening

In strawberry, Fragaria x ananassa Duch., fruit, two different endo-beta-1,4-glucanase genes, FaEG1 and FaEG3, also named Cel1 and Cel2, are expressed during the softening process that occurs during fruit ripening. It has also been suggested that FaEG3, which contains a putative cellulose-binding domain, could play a key role in fruit development, since previous attempts to down-regulate this gene through transgenesis have been unsuccessful. In this investigation, we obtained transgenic strawberry plants containing an antisense sequence of the FaEG3 gene under the control of the 35S promoter. Ripened fruit from transgenic lines (Acel lines) showed large variation in FaEG3 silencing, but fruit firmness was similar to control fruit in all the lines. Two Acel lines showing almost 95% reductions in FaEG3 mRNA levels were selected for further study. In these lines, FaEG3 down-regulation was high, from 78 to 95%, at all fruit developmental stages, whereas FaEG1 was only slightly suppressed. In spite of the high FaEG3 silencing achieved, EGase activity was not modified in ripe fruit. At the cell wall level, walls from transgenic ripe fruit contained a significantly higher amount of the 4 M KOH fraction, which is enriched in hemicellulosic polymers. The analysis of this fraction by size exclusion chromatography showed that transgenic cell walls contained a smaller amount of higher molecular mass polymers than controls. Altogether, these results indicate that FaEG3 does not play a key role either in fruit development or fruit softening. However, its silencing affects the amount and, in a minor way, the size of hemicellulosic polymers. (C) 2009 Elsevier B.V. All rights reserved.