Intracellular characterization of bean yellow mosaic virus-induced inclusions by differential enzyme digestion

Abstract

The composition of crystalline inclusions and dense bands in cells of broad bean leaves infected with bean yellow mosaic virus was studied by differential enzymatic digestion. Frozen thick sections were prefixed in glutaraldehyde and exposed to proteinases and nucleases, after which ultrathin sections were prepared for electron microscopy. Examination revealed that the crystals were completely digested by pepsin in 30 min, whereas the dense bands remained intact for the first 20 min, and could not be found after longer periods of digestion. When ultrathin sections of tissues embedded in glycol methacrylate were incubated with the enzymes, pepsin digested the crystals; this left only a filamentous residue which did not disappear after further exposure to pepsin or to RNase. Trypsin had the same effect, but was slower and less consistent. The dense bands were entirely digested in thin sections by pepsin and trypsin. Neither inclusion was affected by RNase or DNase in thick or thin sections. These results demonstrate that the crystals and dense bands are composed entirely or primarily of protein, but there is no evidence that they contain nucleic acid.