Dominique Mazau

Hydroxyproline-rich glycoprotein accumulation in the cell walls of plants infected by various pathogens

Twenty-five samples of tissue from 16 different host—pathogen systems were analysed for changes in the hydroxyproline (Hyp) content of their cell walls and for the glycosylation pattern of the Hyp. Hydroxyproline and the corresponding Hyp-rich glycoproteins (HRGP) increased in all infected dicotyledons studied but the extent varied according to the host species. Fungal, bacteria! and viral infections triggered accumulation of the glycoprotein. This response occurred in relation to resistant as well as susceptible reactions. No significant increases were found in infected monocotyledons. This work confirmed that the cell wall HRGP of dicotyledons are very different from those of monocotyledons. A hitherto unreported Hyp-oligosaccharide was identified in the cell walls of melon, bean and cucumber.

Time-course study of the accumulation of hydroxyproline-rich glycoproteins in root cells of susceptible and resistant tomato plants infected by Fusarium oxysporum f. sp. radicis-lycopersici

The accumulation of hydroxyproline-rich glycoproteins (HRGPs) in cell walls of dicotyledonous plants is thought to be involved in the defense response to pathogens. An antiserum raised against deglycosylated HRGPs from melon was used for studying the subcellular localization of these glycoproteins in susceptible and resistant tomato (Lycopersicon esculentum Mill.) root tissues infected by Fusarium oxysporum f.sp. radicis-lycopersici. A time-course of HRGP accumulation revealed that these glycoproteins increased earlier and to a higher extent in resistant than in susceptible cultivars. In the compatible interaction, increase in HRGPs was largely correlated with pathogen invasion and appeared to occur as a result of wall damage. In the incompatible interaction, HRGPs accumulated in the walls of uninvaded cells, thus indicating a possible role in the protection against fungal penetration. The occurrence of substantial amounts of HRGPs in papillae, known to be physical barriers formed in response to infection, and in intercellular spaces provides additional support to the concept that such glycoproteins play an important role in disease resistance.

Differential accumulation of hydroxyproline-rich glycoproteins in bean root nodule cells infected with a wild-type strain or a C4-dicarboxylic acid mutant of Rhizobium leguminosarum bv. phaseoli.

An antiserum raised against deglycosylated hydroxyproline-rich glycoproteins (HPGPs) from melon (Cucumis melo L.) was used to study the relationship between Rhizobium infection and induction of HRGPs in bean (Phaseolus vulgaris L.) root nodule cells infected with either the wild-type or a C4-dicarboxylic acid mutant strain of Rhizobium leguminosarum bv. phaseoli. In effective nodules, where fixation of atmospheric dinitrogen is taking place, HRGPs were found to accumulate mainly in the walls of infected cells and in peribacteroid membranes surrounding groups of bacteroids. Internal ramifications of the peribacteroid membrane were also enriched in HRGPs whereas the peribacteroid space as well as the bacteroids themselves were free of these glycoproteins. In mutant-induced root nodules, HRGPs were specifically associated with the electron-dense, laminated structures formed in plastids as a reaction to infection by this mutant. The presence of HRGPs was also detected in the host cytoplasm. The aberrant distribution of HRGPs in infected cells of mutant-induced nodules likely reflects one aspect of the altered host metabolism in relation to peribacteroid-membrane breakdown. The possibility that the antiserum used for HRGP localization may have cross-reacted with ENOD 2 gene products is discussed in relation to amino-acid sequences and sites of accumulation.

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.

Accumulation of hydroxyproline-rich glycoprotein mRNAs in infected or ethylene treated melon plants

Abstract Infection of melon plants by Colletotrichum lagenarium induces the production of five mRNA species (1·4, 1·65, 2·15, 3·2 and 4·5 kB) which hybridize to a genomic clone of hydroxyproline rich glycoproteins (HRGP) from carrot. Induction takes place 3 to 4 days after inoculation, which is consistent with the time course of accumulation of HRGP in the cell wall upon infection. mRNAs are expressed in leaves, stems, and roots. All transcripts, except the 4·5 kb species, are present among the cytosine-rich RNA isolated by affinity chromatography on an oligo(dG) cellulose column. Treatment of melon plants with ethylene leads to early induction of the 1·4 and 1·65 species and, to a lesser extent, the other three transcripts.

Hydroxyproline-containing fragments in the cell wall of Phytophthora parasitica

Abstract The composition of a cell wall derived elicitor preparation from Phytophthora parasitica var. nicotianae which induced ethylene and proteinase inhibitor production in tobacco plants, was analysed. The peptide part which accounted for 77% of the dry mass was characterized by high levels of Glx, Ala, Thr, Asx, Ser and Hyp; the carbohydrate moiety was mainly mannose and glucose. Unlike most plants, in Phytophthora cell walls and elicitor, most of the Hyp residues were non-glycosylated. The glycosylated Hyp (7%) carried oligoarabinose side chains composed of three or four arabinose residues.

Elicitors and Ethylene Trigger Defense Responses in Plants

Infected plants often produce large amounts of ethylene. The possible role of this hormone in plant-microorganism interaction is poorly understood. We have previously demonstrated that ethylene protects melon plants against Colletotrichum lagenarium, the causal agent of anthracnose (1). This has led to the hypothesis that ethylene could be a message which triggers defense responses in plants. The increased synthesis of proteins associated to defense mechanisms during plant-microorganism interactions, and their elicitation by ethylene, or via ethylene by fungal elicitors, are successively reported. The 3 proteins considered in this study are cell wall hydroxyproline-rich glycoprotein (HRGP, which strengthens the plant cell wall), chitinase (which digests chitin-containing fungal cell wall), and proteolytic inhibitors (which neutralize the trypsin-like proteases of plant pathogens).

Cytosine-rich RNAs from infected melon plants and their in vitro translation products

Plant cell wall hydroxyproline-rich glycoproteins (HRGPs) are synthesized from cytosine-rich messenger RNA. As an approach to understanding the increased synthesis of HRGP in melon plants infected by Colletotrichum lagmarium, polyadenylated [poly(A)]RNA was isolated and subsequently enriched in cytosine-rich RNA by chromatography on oligoguanylic acid-cellulose [oligo(dG)cellulose]. In vitro translation of poly(A)RNA extracted from melon plants revealed several modifications of the peptide pattern during infection. In particular, the cytosine-rich RNA obtained coded for proline-rich peptides; among them, two peptides of molecular weight 54 500 and 56 000, were specifically coded by the cytosine-rich RNA of infected plants. The two peptides were characterized by a molecular weight similar to the peptide moiety of melon HRGP (55 000), and the time-course of their appearance in vitro was coincident with the in vivo accumulation of HRGP. The data strongly suggest that the two peptides are two HRGP precursors whose synthesis is increased as a result of an increase of translatable cytosine-rich RNA during infection.

Biochemical study of hydroxyproline-rich glycoproteins in plant-pathogen interactions

The cell surfaces of a host plant and of a pathogen play an important and complex role during pathogenesis. Firstly, recognition events are supposed to involve specific sites of the cell wall or plasmalemma of the two partners. Secondly, intense degradation of host plant cell walls by fungal bacterial hydrolases has often been reported during the course of infection; in view of the occurrence of chitinase and β-1,3 glucanase in plants [1,2, 3], there are no reasons to believe a priori that chitin or glucan containing pathogen cell surfaces are not degraded as well, at least to some extent. Thirdly, active defence responses take place at the cell wall level of host plants [4]. It is likely, although not much documented yet, that pathogens have the ability to develop their own protection.

Cellular and Molecular Approaches of Defense in Plants

The signals exchanged during plant-microorganism interactions account for a three-step phenomenon leading from signal generation to cell responses, via transduction pathways. Recent reviews on signalling molecules and on their effects have been published (Darvill and Albersheim, 1984; Ryan, 1988). Attempts to integrate general features and more specific data from our work are presented.