CHARACTERIZATION OF 1,3-BETA-D-GLUCAN (CALLOSE) SYNTHESIS IN ROOTS OF TRITICUM-AESTIVUM IN RESPONSE TO ALUMINUM TOXICITY

To develop a sensitive marker for aluminum (Al) toxicity, synthesis of 1,3-beta-D-glucan (callose) in 5-mm root tips of Triticum aestivum was examined using spectrofluorometry. In a time course study where an Al-sensitive cultivar, Neepawa, was exposed to 75 mu M Al, a rapid phase of callose synthesis was observed in the first 6-12 h, followed by a slower linear phase with no saturation up to 48 h. Treatment with Al increased accumulation of callose by 86% within 30 min and by 3821% after 48 h. In experiments comparing genotypes, more callose accumulated in roots of Al-sensitive cultivars than in Al-resistant cultivars/lines after 2-h and 24-h exposure. Accumulation of callose increased at concentrations as low as 5 mu M Al and continued to increase with saturation occurring between 250 and 500 mu M Al. callose synthesis was negatively correlated with root growth. An Al-resistant experimental line, PT 741, and an Al-sensitive cultivar, Neepawa, accumulated callose to a similar extent when faced with concentrations of Al that produced equal reductions in root growth. Because accumulation of callose reflects cumulative exposure to Al, a second technique based upon labeling of callose with [C-14]glucose was developed to measure current synthesis. Treatment with Al increased incorporation of C-14 into laminarinase (a 1,3(1,3; 1,4)-beta-D-glucan 3 (4)-glucanohydrolase, EC 3.2.1.6) digestion products from cell-wall material isolated from both the Al-sensitive Neepawa and Al-resistant PT 741. Greater incorporation of C-14 was observed in Neepawa over a broad range of Al concentrations (0-250 mu M). Results of both spectrofluorometric and C-14 labeling studies support the use of callose synthesis as a rapid and sensitive marker for Al-induced injury.