Induction of (1→3,1→4)-β-D-glucan hydrolases in leaves of dark-incubated barley seedlings

When seedlings of barley (Hordeum vulgare L.) were transferred from a natural light/dark cycle into darkness, (1→3,1→4)-β-D-glucan endohydrolase (EC 3.2.1.73) activity in leaf extracts increased 3- to 4-fold after 2 days. Activity decreased to normal levels within a day if the light/dark cycle was restored. Although there are two (1→3,1→4)-β-D-glucan endohydrolase isoenzymes in barley, the increased enzyme activity in dark-grown seedlings was attributable entirely to increases in isoenzyme EI. Northern hybridization analyses confirmed that mRNA transcripts encoding (1→3,1→4)-β-D-glucan endohydrolase isoenzyme EI accumulated in the leaves of dark-incubated seedlings; no isoenzyme EII mRNA was detected. Activity of β-D-glucan glucohydrolases also increased 10-fold after 2 days of dark treatment. The latter, broad-specificity enzymes release glucose from (1→3,1→4)-β-D-glucans and from β-D-oligoglucosides released by (1→3,1→4)-β-D-glucan endohydrolases. Consistent with the activity patterns of these enzymes, the (1→3,1→4)-β-D-glucan content of leaf cell walls decreased by about 30% when barley seedlings were transferred into darkness. Soluble sugars in the leaves decreased by about 60% during the same period. Because no measurable leaf elongation was detected during the various light/dark treatments, the enzymes were unlikely to be participating in wall loosening and cell elongation. Instead, the results suggest that cell wall (1→3,1→4)-β-D-glucans can be re-mobilized in the non-elongating, dark-incubated leaves and the glucose so generated could serve as an energy source under conditions of sugar depletion.