Endo-1,4-β-glucanase gene expression and cell wall hydrolase activities during abscission in Valencia orange

The physiological and molecular events of ethylene-induced abscission in mature fruit calyx, laminar and floral abscission zones of cv. Valencia orange were examined. Continuous exposure of fruit explants to 5 mu l 1(-1) ethylene for 2 to 40 h resulted in marked increases in endo-1,4-beta-glucanase (cellulase) and polygalacturonase (PG) activities in calyx abscission zones. Two abscission-related cellulases and one PG were found. The major peak of cellulase activity corresponded to a pI of 8.0 and molecular weight of 51 kDa, whereas the minor cellulase peak had a pI of 5.5. The abscission polygalacturonase had a pI of 5.5. Calyx abscission zone RNA was amplified with degenerate primers based on sequence of the purified Valencia orange calyx abscission cellulase, and cloned. The two partial cellulase cDNA clones were 59% identical at the nucleotide level. Genomic Southern analysis suggested that Valencia orange contained two groups of cellulase genes. A full-length cDNA clone from each group was isolated from a cDNA library prepared from ethylene-induced calyx abscission zcne mRNA. Both genes were expressed in ethylene-induced calyx, laminar and floral abscission zones, but were not expressed in non-induced abscission zones or mature leaves treated with or without ethylene, young bark or young fruit of Valencia.