Immunogold localization of hydroxyproline-rich glycoproteins in necrotic tissue of Nicotiana tabacum L. cv. Xanthi-nc infected by tobacco mosaic virus

Abstract

An antiserum raised against melon hydroxyproline-rich glycoprotein 2b was found to cross react specifically with deglycosylated hydroxyproline-rich glycoproteins from tobacco. This antiserum was used to study the subcellular localization of hydroxyproline-rich glycoproteins in Nicotiana tabacum cv. Xanthi-nc reacting hypersensitively to tobacco mosaic virus infection. The hypersensitive reaction was characterized by the formation of necrotic lesions around the point of virus entry. Leaf samples were collected 7 days after inoculation when lesions reached an average diameter of 3 mm. In the core of necrotic lesions, hydroxyproline-rich glycoproteins were localized in hemispherical excrescences formed along cell walls and in intercellular spaces filled with an electron-opaque material. However, they were not detected in host cell walls. In the halo surrounding the necrotic core, hydroxyproline-rich glycoproteins were found to be associated with an amorphous material frequently deposited in vascular parenchyma cells and with aggregated fibrils filling xylem vessels. In the green tissue bordering necrotic lesions, hydroxyproline-rich glycoproteins were nearly absent except in vessels which contained a fine network of intertwined fibrils. Farther away in the green tissue area, very low amounts of hydroxyproline-rich glycoproteins were detected. The accumulation of hydroxyproline-rich glycoproteins at strategic sites such as the intercellular spaces in the necrotic area is discussed in relation to plant stress and resistance.