Isolation by ConA binding of haustoria from different rust fungi and comparison of their surface qualities

Abstract

Summary. Rust haustoria isolated from infected leaf tissue strongly bind to ConA. This property was exploited to purify them by affinity chromatography on a ConA-Sepharose macrobead column. Haustoria were obtained with more than 90% purity and yields of up to 50%. Binding of haustoria to the column was partially inhibited by a ConA-specific sugar, methyl ct-D-mannopyranoside. Compared to ConA, Lens culinaris agglutinin and wheat germ agghitinin were less efficient affinity ligands. Using ConA-Sepharose, rust haustoria from a variety of sources could be isolated with equal efficiency, indicating that they have similar carbohydrate surface properties. The haustoria maintained their typical shape after the isolation procedure, which suggests a rather rigid wall structure. The morphology of haustoria was characteristic both for a given species and the nuclear condition of the rust mycelium. Electron microscopy of isolated haustoria revealed an intact haustorial wall surrounded by a fibrillar layer presumably derived from the extrahaustorial matrix. The matrix thus appears to represent a layer with geMike properties which is rich in ConA-binding carbohydrates and connected to the haustorial wall but not to the host-derived extrahaustorial membrane.

The Melampsora lini AvrL567 Avirulence Genes Are Expressed in Haustoria and Their Products Are Recognized inside Plant Cells

“…We used affinity chromatography with a Sepharose concanavalin A column (Hahn and Mendgen, 1992) to purify haustoria from L. usitatissimum leaves infected with rust CH5. The presence of intact haustoria in these samples was confirmed by microscopy and immunolabeling with a monoclonal antibody that detects polysaccharide epitopes present on the haustorial surface ( Figure 5C; Murdoch et al, 1998).…”

“…Haustoria were isolated from L. usitatissimum leaves 6 d after inoculation with rust strain CH5 by affinity chromatography as described by Hahn and Mendgen (1992), except that an 11-mM pore size nylon mesh was used to remove plant cell material from the crude preparation. An affinity column was prepared by covalently attaching concanavalin A (Pharmacia Biotech, Uppsala, Sweden) to cyanogen bromide-activated Sepharose 6MB (Pharmacia Biotech) as described in the manufacturer's protocol.…”

High Level Activation of Vitamin B1 Biosynthesis Genes in Haustoria of the Rust Fungus Uromyces fabae

“…RNA isolations from rust-infected leaves, U. fabae infection structures, and isolated haustoria (Hahn and Mendgen 1992), and Northern (RNA) blot hybridization with digoxigenin-labeled DNA probes ), were performed as described.…”

Plasma Membrane H+-ATPase Activity in Spores, Germ Tubes, and Haustoria of the Rust FungusUromyces viciae-fabae

“…Haustoria were isolated by binding on a ConASepharose macrobead column as described by Hahn and Mendgen (1992) with the exception that BSA 2 was omitted from the homogenization and the suspension buffers.…”

“…A new method to isolate rust haustoria from infected plant material (Hahn and Mendgen, 1992) has opened the possibility to characterize ATPase in hyphae and haustoria and to study the haustorial function.…”