Proteinaceous hexagonal inclusions in hyphae of Whetzelinia sclerotiorum and Neurospora crassa

Abstract

The chemical nature of the hexagonal inclusions occurring in microbodies of Whetzelinia sclerotiorum and in the cytoplasm of Neurospora crassa was investigated. These inclusions were considered to be ergosterol or protein. The major β-3-hydroxyl-sterol present in W. sclerotiorum was identified as ergosterol. When glutaraldehyde (GA) fixed hyphae of both W. sclerotiorum and N. crassa were treated with sterol-extracting solvents, the hexagonal inclusions were still observed in thin sections. Sodium methoxide treatment of thin sections of hyphae fixed with GA and OsO4 removed the lipid bodies, but not the hexagonal inclusions. Various proteases, subtilisin, Sigma protease type VI, and pepsin, digested the inclusions from thin sections of hyphae fixed only with GA. Inclusions in thin sections of hyphae fixed with 2% KMnO4 were extracted by 1.0 N NH4OH, 0.1 M Na3PO4, or 0.16 N NaOH (about pH 12), but not by 0.16 N NaOH adjusted to pH 8.8 with HCl. These results indicate that the hexagonal inclusions in W. sclerotiorum and N. crassa are primarily proteinaceous in nature and are similar in composition to the Woronin bodies observed in other Ascomycetes.