HOST PATHOGEN INTERACTIONS .36. PARTIAL-PURIFICATION AND CHARACTERIZATION OF HEAT-LABILE MOLECULES SECRETED BY THE RICE BLAST PATHOGEN THAT SOLUBILIZE PLANT-CELL WALL FRAGMENTS THAT KILL PLANT-CELLS

Heat-labile factors capable of killing plant cells are secreted by the rice pathogen Magnaporthe grisea when grown on rice cell walls. Inhibition of [14C]-leucine incorporation into maize cells (Zea mays cv. Black Mexican Sweet) was shown to be as reliable as the vital dyes 2,3,5-triphenyltetrazolium chloride and fluorescein diacetate for assessing cell viability. The heat-labile factors responsible for killing plant cells were partially purified by CM-Sephadex and Superose 12 chromatography. A combination of four of the Superose 12 column fractions synergistically killed the plant cells; the killing activity of the combined fractions was 2·5 times as high as that obtained by the sum of the four fractions assayed individually. We purified to apparent homogeneity pectin lyase (PL), pectin methylesterase (PME), and xylanase from the fungal culture filtrate. When these enzymes were tested in various combinations and at the same concentrations as they were found in the culture filtrate, they did not kill plant cells. The same enzymes were not able to release fragments that killed plant cells from isolated maize cell walls, whereas fractions containing the partially purified heat-labile killing activity rapidly released heat-stable maize cell wall fragments that killed maize cells. The results of this study indicate that a heat-labile killing activity secreted by M. grisea, which probably consists of two or more factors (presumably proteins), solubilizes from maize cell walls heat-stable fragments (presumably carbohydrates) that kill maize cells. Furthermore, although pectic enzymes may prove to be necessary for killing, the pectic enzymes in the culture filtrate of M. grisea do not, by themselves, kill maize cells. © 1990.