Reversible Dark-Induced Senescence of Soybean Root Nodules

Abstract

Nodule senescence was induced in intact soybean I Glycine max. (L.) Merr., cv Woodworthl plants by an 8-day dark treatment. Dark-induced senescence resulted in the complete loss of acetylene reduction activity, a 67% loss of total soluble protein, and an almost complete loss in total leghemoglobin of nodule extracts. Isoelectric focusing gels demonstrated a preferential loss of certain proteins, which was correlated with an increase in endoprotease specific activity toward azocasein. Nodules were completely green after the 8-day dark treatment. If plants were returned to a normal photoperiod after 8 days in the dark, nodules recovered from the dark treatment in 12 to 16 days. Acetylene reduction activity returned to normal, and both total soluble protein and leghemoglobin were resynthesized while protease activity against azocasein decreased to the level of control nodules. The nodule population that had turned green after 8 days in the dark exhibited a progressive increase in red color starting nearest the exterior of the nodule, and after 16 days of recovery nodules were indistinguishable from control nodules maintained under a normal photoperiod.and a senescent zone connected to the lateral root (9,16,19,27).Thus, a single pea or alfalfa nodule possesses zones of different age and symbiotic viability. Senescence of alfalfa nodules can be accelerated as a result of defoliation or prolonged darkness (4,26,27), and the capacity of a single nodule to fix nitrogen can be regained by virtue of reinfection of the meristematic tissue (27 Initial experiments in this laboratory suggested that the metabolic changes observed in soybean nodules as a result of foliar dark treatment, including loss of nitrogenase activity, could be reversed. This study was conducted to investigate the changes in nodular metabolism which occurred as a result of dark-induced senescence, and to document the reversal of this process in soybean nodules.Foliar dark treatment has been used to induce metabolic changes in legume nodules which are characteristic of nodule senescence (3,15,17, 28). Dark treatment of soybean plants caused an immediate decline in the level of nitrogenase activity, although at least 4 to 6 d of continuous darkness were required to completely inhibit N2 fixation (17, 28). Declining glucose concentrations were closely correlated to the complete inhibition of nitrogenase activity (17), and although poly-/8-hydroxybutyrate did not appear to be used as an alternate energy source for N2 fixation, it might serve as a source of four carbon intermediates for bacteroid respiration (28). Dark treatment of soybeans for 8 to 11 d resulted in a 50% decline in total Lb3 which indicated substantial changes occurring in host cell function. In contrast, no differences were detected in bacteroid nucleic acid content or size (17), which suggests that during dark-induced senescence of soybean nodules bacteroid integrity was maintained to a greater extent than host cell function.The morphology of senescing legume nodules has been m...

Proteolytic Activity in Soybean Root Nodules

“…In all assays, the enzyme activity was proportional to time throughout the period of incubation. The azocaseindigesting activity of nodule cytosol was determined as reported previously (16). The proteinase activity of the crude bacteroid extracts was measured using azocasein and hemoglobin as substrates.…”

“…protein standard. Total leghemoglobin in the nodule cytosol fraction was determined by the pyridine hemochrome assay (1) as described previously (16). The average mol wt of leghemoglobin was assumed to be 16,375 (4).…”

“…Nitrogen fixation was measured on intact plants using the acetylene reduction assay (7) as reported previously (16).…”

Developmental Regulation of Enzymes of Sucrose and Hexose Metabolism in Effective and Ineffective Soybean Nodules

“…Sucrose synthase was the only sucrose metabolic enzyme which was not expressed in ineffective nodules. To further determine the regulation of the sucrose metabolizing enzymes, nodulated plants were placed in the dark to reduce the transport of sucrose to nodules which results in reduced acetylene reduction activity (24). Although the protein content declined by 58%, sucrose synthase was the only enzyme to show a marked reduction in activity (Table II).…”

“…On d 20, one group of plants was placed in the dark for 8 d as described by Pfeiffer et al (24). The control group remained at the normal light/dark regime.…”