Membranes in lupin root nodules: II. preparation and properties of peribacteroid membranes and bacteroid envelope inner membranes from developing lupin nodules

Abstract

Peribacteroid membranes and bacteroid envelope inner membranes have been isolated from developing lupin nodules. Isolation of the peribacteroid membranes was achieved by first preparing membrane-enclosed bacteroids free from other plant organelles or membranes. The peribacteroid membranes were then released by osmotic shock and purified by centrifugation to equilibrium on sucrose gradients. The bacteroids were broken in a pressure cell and the bacteroid envelope inner membranes were isolated using sucrose gradient fractionation of the bacteroid total envelope preparation. The density of the peribacteroid membranes decreased during the period of development of N2-fixation in lupin nodules from 1.148 g/ml for nodules from 12-day plants to 1.137 g/ml for nodules from 18-day plants. The density of the bacteroid envelope inner membranes from nodules from 18-day plants was 1–153 g/ml. The identity and homogeneity of the isolated membranes was established, by comparison with membranes in intact nodules, using phosphotungstic acid and silver staining of thin sections and particle densitites on faces of freeze-fracture replicas of the membranes. Analyses for NADH oxidase and succinate dehydrogenase, spectral analyses and gel-electrophoretic analysis of proteins were also used to characterize the membrane and soluble protein fractions from the nodules. The ratio of lipid to protein was 6.1 for the peribacteroid membranes and 2.5 for the bacteroid envelope inner membranes. Leghaemoglobin was localized in the plant cytoplasm in lupin nodules and not in the peribacteroid space.

Leghemoglobin biosynthesis in soybean root nodules. Characterization of the nascent and released peptides and the relative rate of synthesis of the major leghemoglobins

“…We could not detect any immunoreactive material inside the isolated membrane envelopes enclosing the bacteroids (Verma et al, 1978). Robertson et al (1978) showed by isolating the bacteroids enclosed in the membrane envelope from Lupin nodules in the presence of ferritin that these envelopes do not open and reseal during fractionation, thereby leaking all their Lb contents. These data, along with analyses of the primary translation product of Lb mRNA, conclusively prove that Lb is a cytoplasmic protein of the host cell and it does not cross the membrane envelope enclosing the bacteroids (see Verma & Bal 1976.…”

RPG interacts with E3-ligase CERBERUS to mediate rhizobial infection in Lotus japonicus

“…Rhizobia colonize the ITs which grow through root cells, ultimately reaching the nodule primordium. The bacteria are then budded off surrounded by a plant-made membrane into plant cells in which they fix nitrogen using carbon supplied by the plant [ 5 ]. Genetic studies in Lotus japonicus and Medicago truncatula have identified several genes required for IT initiation and formation.…”

RPG interacts with E3-ligase CERBERUS to mediate rhizobial infection in Lotus japonicus

“…Rhizobia attach to root hairs, triggering root-hair curling that entraps rhizobia which induce localized cell-wall degradation and rearrangement of the plant cytoskeleton, contributing to the formation of plant-made tunnel-like structures called the infection threads (ITs) (Gage, 2004). Rhizobia colonize the ITs which grow through root cells, ultimately reaching the nodule primordium; the bacteria are then budded off surrounded by a plant-made membrane into plant cells in which they fix nitrogen using carbon supplied by the plant (Robertson et al, 1978). Genetic studies in Lotus japonicus and Medicago truncatula have identified several genes required for IT initiation and formation.…”