Correlation between infection by Rhizobium leguminosarum and lectin on the surface of Pisum sativum L. roots

Abstract

The lectin on the surface of 4- and 5-dold pea roots was located by the use of indirect immunofluorescence. Specific antibodies raised in rabbits against pea seed isolectin 2, which crossreact with root lectins, were used as primary immunoglobulins and were visualized with fluorescein- or tetramethylrhodamine-isothiocyanate-labeled goat antirabbit immunoglobulin G. Lectin was observed on the tips of newly formed, growing root hairs and on epidermal cells located just below the young hairs. On both types of cells, lectin was concentrated in dense small patches rather than uniformly distributed. Lectin-positive young hairs were grouped opposite the (proto)xylematic poles. Older but still-elongating root hairs presented only traces of lectin or none at all. A similar pattern of distribution was found in different pea cultivars, as well as in a supernodulating and a non-nodulating pea mutant. Growth in a nitrate concentration which inhibits nodulation did not affect lectin distribution on the surface of pea roots of this age. We tested whether or not the root zones where lectin was observed were susceptible to infection by Rhizobium leguminosarum. When low inoculum doses (consisting of less than 10(6) bacteria·ml(-1)) were placed next to lectin-positive epidermal cells and on newly formed root hairs, nodules on the primary roots were formed in 73% and 90% of the plants, respectively. Only a few plants showed primary root nodulation when the inoculum was placed on the root zone where lectin was scarce or absent. These results show that lectin is present at those sites on the pea root that are susceptible to infection by the bacterial symbiont.

Sugar-Binding Activity of Pea Lectin Expressed in White Clover Hairy Roots

“…In some experiments, we used Boehringer Mannheim chemiluminescence detection solutions as substrates for peroxidase, following protocols described by the manufacturer. Anti-PSL rabbit antibodies (RAL 439, raised against SDS-denaturated PSL; IKAL 80, raised against native PSL) were obtained as described by Díaz et al (1984) Localization of Root Hair PSL by lndirect lmmunofluorescence Initially, we tried to detect PSL on the surface of putatively psl-transformed white clover roots by indirect immunofluorescence using affinity-purified anti-PSL2 antibodies, as described by Díaz et al (1986). In some experiments, we also used RAL 439 that had been affinity-purified by passage through a protein-G column.…”

“…To obtain root hair extracts, we took advantage of the transition between the zone of growing root hairs (extending 0.5-2 cm from the root tip) and the zone of nonelongating, mature root hairs (extending 2-3 cm above the zone of growing root hairs), which can be easily observed on the roots of 4-d-old pea seedlings (Díaz et al, 1986). To break off root hairs, sections from the zones of growing root hairs and mature root hairs of 4-d-old pea seedlings were submerged in liquid nitrogen, as described by Díaz et al (1990).…”

“…Root hair PSL was first localized by indirect immunofluorescence using rat IgE as PSL ligand and goat anti-rat IgE coupled to fluorescein isothiocyanate (Nordic Immunochemical Laboratories) following the procedure described by Díaz et al (1986), except that incubations were performed at room temperature. A second method, designed to obtain increased sensitivity, included the following steps.…”

“…This step is a determinant of host plant specificity in some Rhizobium-host plant combinations (Horvath et al, processing, which ultimately yields an acidic isolectin that can be isolated from pea roots by affinity chromatography on dextran beads (Díaz et al, 1990;Hoedemaeker et al, 1994). As shown by indirect immunofluorescence microscopy, root PSL is targeted to the tips of emerging and growing pea root hairs in the root zone susceptible to infection by Xhizobiiim (Díaz et al, 1986). Clover root lectin has been localized at similar sites on white clover roots (Dazzo et al, 1978).…”

Comparison of protein patterns in non‐mycorrhizal and vesicular–arbuscular mycorrhizal roots of red clover

“…At the extracellular level, the root epidermal cells interact with the tip of the hypha when it forms the appressorium. This interaction may involve Iectins, as in the case of the Rhizobium-legume symbiosis (Diaz et al, 1986). The role of Iectins in this process may be related to the activity of hydrolytic enzymes in that lectin may act as a cofactor which enables key lytic activities (Gilboa-Garber & Garber, 1989).…”