The interaction in planta of polygalacturonases from Botrytis cinerea with a cell wall-bound polygalacturonase-inhibiting protein (PGIP) in raspberry fruits

The activities of four fungal polygalacturonases (endo-PGI,endo-PGII, exo-PGI, exo-PGII), detected when Botrytis cinereawas grown on immature fruits of red raspberry (Rubus idaeus),were fractionated into soluble and wall-bound fractions. Westernblots and plate-trapped antigen ELISA showed that endo-PGI andendo-PGII were most abundant in the cell wall-bound fractionsof the host. Immuno-inhibition studies using a polyclonal antiserumagainst polygalacturonase-inhibiting protein (PGIP), purifiedfrom immature raspberry fruits, showed that the low level offungal PG activity detected in fractions containing endo-PGIwas due to the presence of PGIP. When a purified preparationof endo-PGI and endo-PGII from B. cinerea was allowed to reactin vitro with either a crude host cell wall preparation, orone which had previously been treated to remove cell wall-boundproteins, both endo-PG isozymes had a greater binding capacitytowards the former wall preparation. Endo-PGI and endo-PGIIalso had an affinity for fungal cell walls. Exo-PGI and exo-PGIIbound to both fungal and host cell walls. Greater quantitiesof fungal endo-PGs were detected by ELISA in fruits previouslyfrozen and thawed (‘freeze-thawed’) and inoculated,than in fresh inoculated fruit. This result paralleled the extentof fungal growth in these tissues as assessed by chitin assayand suggests that the resistance shown by raspberries is dependenton continual replacement of inhibitory substances or inducedresistance mechanisms. Key words: Polygalacturonase-inhibiting protein, Rubus idaeus, red raspberry, Botrytis cinerea, pectinases