CELL-WALL DEFICIENCY IN SLIME STRAINS OF NEUROSPORA-CRASSA - OSMOTIC INHIBITION OF CELL-WALL SYNTHESIS AND BETA-D-GLUCAN SYNTHASE ACTIVITY

1. The RCP-3 S/H mutant of Neurospora crassa was obtained by vegetativ e selection in medium of high osmolarity of a mycelial form of an fz, sg, os-1 (''slime''-like) segregant. The mutant exhibits spheroplast-h yphal dimorphism conditioned by the osmolarity of the culture medium ( Pietro et al. (1990). Journal of General Microbiology, 136: 121-129), The carbohydrate composition of the cell wall of the mutant was differ ent from that of the wild type in the absence of an alkali-soluble gal actosaminoglycan polymer. Furthermore the mutant cell wall had a somew hat lower content of beta-glucan relative to that of chitin. 2. Increa sing concentrations of sorbitol in the culture medium of the mutant in hibited by 10-fold the formation of cell wall relative to total biomas s. The cell wall of the mutant cultured in the presence of sorbitol la cked mannose- and galactose-containing polymers, and also showed progr essively lower amounts of B-glucan relative to chitin. 3. The activity of membrane-bound (1-3)-beta-D-glucan synthase from the mutant grown in the absence of sorbitol shared several properties with the wild typ e enzyme (i,e., Km app., Vmax, stability at 30 degrees C, activation b y GTP gamma S, and dissociability by treatment with NaCl and Tergitol NP-40 into a membrane-bound catalytic center and a GTP-binding activat ing protein). On the other hand, the enzyme from the mutant but not th at from the wild type was inactivated by about 15% by treatment with N aCl and detergent. 4. At high concentrations of sorbitol (1.0 M) the R CP-3 S/H mutant exclusively produced spheroplasts devoid of(1-3)-beta- D-glucan synthase activity. The defect was at the level of the membran e-bound catalytic center. The activity of the GTP-binding activating f actor was apparently normal in these cells.