Role of the putative membrane-bound endo-1,4-beta-glucanase KORRIGAN in cell elongation and cellulose synthesis in Arabidopsis thaliana.

A temperature-sensitive, elongation-deficient mutant of Arabidopsis thaliana was isolated. At the non-permissive temperature of 31 degrees C, the mutation impaired tissue elongation; otherwise, tissue development was normal. Hypocotyl cells that had established cell walls at 21 degrees C under light-dark cycles ceased elongation and swelled when the mutant was shifted to 31 degrees C and darkness, indicating that the affected gene is essential for cell elongation. Analysis of the cell walls of mutant plants grown at 31 degrees C revealed that the cellulose content was reduced to 40% and the pectin content was increased to 162% of the corresponding values for the wild type grown at the same temperature. The increased amounts of pectin in the mutant were bound tightly to cellulose microfibrils. No change in the content of hemicellulose was apparent in the 31 degrees C-adapted mutant. Field emission-scanning electron microscopy suggested that the structure of cellulose bundles was affected by the mutation; X-ray diffraction, however, revealed no change in the crystallite size of cellulose microfibrils. The regeneration of cellulose microfibrils from naked mutant protoplasts was substantially delayed at 31 degrees C. The recessive mutation was mapped to chromosome V, and map-based cloning identified it as a single G-->A transition (resulting in a Gly(429)-->Arg substitution) in KORRIGAN, which encodes a putative membrane-bound endo-1,4-beta-glucanase. These results demonstrate that the product of this gene is required for cellulose synthesis.

Callose synthesis in spirostanol treated carrot cells is not triggered by cytosolic calcium, cytosolic pH or membrane potential changes.

Carrot (Daucus carota L.) cell suspensions were treated with a spirostanol saponin from Yucca. This saponin is an elicitor of callose synthesis. Irrespectively of the mode of action of spirostanol on the callose synthase activity itself, the spirostanol-induced callose synthesis in carrot is not preceded by changes in membrane potential, cytosolic free calcium or cytosolic pH. The inability of modulators of cytosolic free calcium content (verapamil, nifedipine and Br-A23187), EGTA and a proton pump inhibitor (vandate) to inhibit or induce callose formation is consistent with a calcium- and pH-independent mechanism for callose deposition.