Xyloglucan (amyloid) mobilisation in the cotyledons of Tropaeolum majus L. seeds following germination

The levels of cell-wall xyloglucan (amyloid) in nasturtium (Tropaeolum majus L.) cotyledons were monitored during a 28-d period covering seed imbibition, germination and early seedling development. The activities of the following enzymes capable of hydrolysing the glycosidic linkages in the xyloglucan were assayed in cotyledon extracts over the same period: endo — (1→4)-β-glucanase (EC 3.2.1.4), β-glucosidase (EC 3.2.1.21), α-xylosidase and β-galactosidase (EC 3.2.1.23). The endo-β-glucanase was assayed viscometrically using xyloglucan as substrate, and the three glycosidases using appropriate p-nitrophenylglycosides. Alpha xylosidase and β-galactosidase, the enzymes which would be expected to hydrolyse the side-chains from the xyloglucan molecule, were also assayed using xyloglucan as substrate. Under our culture conditions, xyloglucan levels remained constant at 30 mg per cotyledon pair for 7 d, that is until 3 d after germination: thereafter, the amount of xyloglucan diminished to zero in a 12-d period. The most rapid period of depletion was between days 9 and 13. The mobilisation of all reserve substances from the cotyledons resulted in a weight-loss of 92 mg: xyloglucan, therefore, is an important storage substance, representing 33% by weight of the seed's substrate reserves. It is a cell-wall storage polysaccharide. Xyloglucan mobilisation was accompanied by a 17-fold increase in endo-β-glucanase activity, a 7-fold increase in β-galactosidase and an 8-fold increase in α-xylosidase activities, all determined using xyloglucan as substrate. All three activities began to increase at day 5, peaked at days 12—14 when the most rapid phase of xyloglucan breakdown was over, and had declined to zero by days 22—25. The levels of theses enzymes have been shown to be consistent with their being responsible for xyloglucan hydrolysis in vivo. Nitrophenyl-β-galactosidase activity increased up to day 3, remained constant and then increased again 2.5-fold from day 5, peaking at day 11. Nitrophenyl-β-glucosidase remained relatively constant up to day 16 and then decreased to zero by day 25. Nitrophenyl-α-xylosidase activity was not detected.