cDNA Cloning of Carrot Extracellular ,&Fructosidase and Its Expression in Response to Wounding and Bacterial lnfection

We isolated a full-length cDNA for apoplastic (extracellular or cell wall-bound) 8-fructosidase (invertase), determined its nucleotide sequence, and used it as a probe to measure changes in mRNA as a result of wounding of carrot storage roots and infection of carrot plants with the bacterial pathogen Erwinia carotovora. The derived amino acid sequence of extracellular 8-fructosidase shows that it is a basic protein (pl 9.9) with a signal sequence for entry into the endoplasmic reticulum and a propeptide at the N terminus that is not present in the mature protein. Amino acid sequence comparison with yeast and bacterial invertases shows that the overall homology is only about 28%, but that there are short consenred motifs, one of which is at the active site. Maturing carrot storage roots contain barely detectable levels of mRNA for extracellular 8-fructosidase and these levels rise slowly but dramatically after wounding with maximal expression after 12 hours. lnfection of roots and leaves of carrot plants with E. carotovora results in a very fast increase in the mRNA levels with maximal expression after 1 hour. These results indicate that apoplastic 8-fructosidase is probably a new and hitherto unrecognized pathogenesis-related protein [Van Loon, L.C. (1985). Plant MOI. Biol. 4,111-1 161. Suspension-cultured carrot cells contain high levels of mRNA for extracellular 0-fructosidase and these levels remain the same whether the cells are grown on sucrose, glucose, or fructose.