Hexose Metabolism in Effective and Ineffective Soybean Nodules1

Soybean (Glycine max) nodules formed by inoculation with either an effective strain or an ineffective (noninvasive, noduleforming) strain of Bradyrhizobium japonicum were assayed for changes in developmental patterns of carbon metabolic enzymes of the plant nodule cells. Of the enzyme activities measured, only sucrose synthase, glutamine synthetase, and alcohol dehydrogenase were altered in the ineffective nodules relative to the effective nodules. Sucrose synthase and glutamine synthetase activities were greatly reduced, whereas alcohol dehydrogenase activity was elevated. Dark-induced senescence severely affected sucrose synthase but had little, if any, effect on the other enzymes measured. The developmental pattems of the anaerobically induced enzymes, aldolase and alcohol dehydrogenase, were different from those expected, implying that their development is not regulated solely by oxygen deprivation. However, anaerobic treatment of nodules resulted in responses similar to those enzymes in maize. The developmental profiles of the carbon metabolic enzymes suggest that carbohydrates are metabolized via the sucrose synthase and pentose phosphate pathways. This route of carbon metabolism, compared to glycolysis, would reduce the requirement of ATP for carbohydrate catabolism, generate NADPH for biosynthetic reactions, and provide intermediates for plant secondary metabolism. The establishment ofan effective N2 symbiosis on the roots of leguminous plants requires infection by the appropriate species ofRhizobium. The Rhizobium/leguminous plant nodule provides a unique environment where photosynthetically derived carbon compounds synthesized by the plant are exchanged for ammonium derived from the reduction of atmospheric N2 catalyzed within the bacterial endophytes. Nodule differentiation results in a variety of morphological (22) and biochemical (15, 18, 27) changes in the plant nodule cells to accommodate the energy needs of the N2 fixation process and to supply carbon skeletons to transport the ammonium formed via nitrogenase. 'Supported by the U.S. Department of Agriculture competitive Grant No. 85-CRCR-1-1734, the Frasch Foundation, and the Missouri Soybean Merchandising Council. This is contribution No. 10489 of the Agricultural Experiment Station of the University of Missouri. 2Present address: Sungene Technologies Corporation, 330 Hillview Ave., Palo Alto, CA 94304. Plant proteins specifically formed as a result of the plantmicrobe interaction are termed nodulins and have been identified in the nodules of several leguminous plants (7, 10, 19, 23). The functions of a few nodulins in various legume hosts has been established: leghemoglobin (2), uricase (27), glutamine synthetase (16), and sucrose synthase (26). However, the roles of most nodulins are not known. Ineffective strains of rhizobia have been used to monitor the expression of nodulins of unknown function in order to determine their role and ontogeny in nodule development (7, 10, 19, 23). Bacterial differentiation and N2 fixation are not essential for expression of some nodulins, as they may be expressed in nodules formed by inoculation with a variety of mutant rhizobia. Mutants of Rhizobium meliloti (10, 20), R. trifolii (8), Rhizobium sp. strain NGR234 (9), and Bradyrhizobium japonicum (19) which exhibit altered exopolysaccharide composition and/or content, uncouple the infection process from nodule development. The nodules formed by these mutants are essentially devoid of rhizobia. Thus, these strains are capable of eliciting the signal to induce plant cell morphogenesis but are unable to gain entry into the plant cells. During development ofnodules formed by inoculation with an effective strain of Rhizobium, a number of enzymes of carbon metabolism increase in activity (17). These increases in activity may be due to nodule-specific forms or simply to elevation of preexisting species to sustain the nodule. Here, we present the developmental changes, relative to those of known nodulins, of a number of enzymes of carbon metabolism within the plant cells of soybean nodules induced by an effective and an ineffective strain of B. japonicum. MATERIALS AND METHODS