A phytotoxic protein-lipopolysaccharide complex produced by Verticillium dahliae

Abstract

A phytotoxic protein-lipopolysaccharide complex (PLPC) was purified to electrophoretic homogenetiy from seven-day-old culture filtrates of Verticillium dahliae by means of acetone precipitation, gel exclusion chromatography, and preparative agarose electrophoresis with a yield of 4.5 mg 1 −1 culture filtrate. The PLPC consists of 15.7% protein, 13.0% lipid, 0.4% phosphate and 70% carbohydrate. The isoelectric point is 3.8 and the amino acid composition of the protein fraction was determined. The complex has a M r of 197 000 and can be dissociated into five protein-containing components, with M r s of 78 000, 62 000, 48 000, 32 0000 and 28 000. Polygalacturonase and cellulase enzyme activities were identified in the PLPC by means of enzyme staining techniques, and were found to be associated with the 28 000 and 48 000 M r complexes, respectively. 1,3-β-Glucanase activity was associated with the 32 000 and 28 000 M r complexes. Treatment of cotton seedlings with concentrations from 2.5 μg ml −1 PLPC resulted in symptoms of wilting and necrosis and was accompanied by inhibition of the H +- ATPase activity of plasma membranes and the rapid transient elicitation of phenylalanine ammonia-lyase activity. Significant differences were found between resistant and susceptible cultivars. When applied to disks of mature cotton leaves, the PLPC elicited the accumulation of pathogenesis-related proteins in the intercellular spaces.