Genetics and expression of pectinmethylesterase, endo-β-glucanase, and polygalacturonase genes in Valencia orange

The genetics and expression of pectinmethylesterase (PME), endo-beta -glucanase (EBG), and polygalacturonase (PG) were studied in Valencia orange in relation to juice quality and cell separation, particularly abscission. We previously isolated two unique PME genes in Valencia orange. One of these PME genes, CsPME1, was constitutively expressed in tissues of Valencia orange. The other gene, CsPME3, was ethylene regulated. Both genes were expressed in ethylene-induced abscission zones of Valencia, but expression of CsPME3 was markedly pronounced. Transient expression of the CsPME3 open reading frame (ORF) in tobacco demonstrated that a large fragment of the variable N-terminus of CsPME3 was cleaved during processing. Western blots of whole cell extracts contained the large (58 kDa) translation product of the expressed CsPME3 construct and the smaller (36 kDa) processed product. The mature protein encoded by the more conserved C terminus of the translated product was associated with the cell wall fraction. The large translated product of the CsPME3 construct was absent in cell wall fractions. These results suggest that processing of PME in Valencia orange is a rapid, perhaps single post-translational cleavage event. In addition to PME, two unique abscission-related EBG genes were isolated from an ethylene-induced mature fruit abscission zone cDNA library. One gene, CsCel-a1, encoded a 51-kDa acidic protein, whereas the other, CsCel-b1 encoded a 51-kDa basic protein. Both genes were expressed in abscission zones. Differential expression of CsCel-a1 and CsCel-b1 in ethylene-induced calyx abscission zones was demonstrated in mature and immature fruit. Two PG genes were also isolated from the abscission zone library. One gene, CsPG1, encoded a 45.7-kDa basic protein. The other gene, CsPG2, is being further characterized. Both genes were expressed in floral, leaf, and fruit abscission zones of Valencia orange but not in other vegetative or reproductive tissues.