Starch content and activities of starch-metabolizing enzymes in effective and ineffective root nodules of soybean SHARON I. FORREST,' DESH PAL S. VERMA,' AND RAJINDER S . DH1NDSA3 Department of Biology, McGill University, 1205 Avenue Docteur Penfield, Montreal, Que., Canada H3A 1B1 Received September 10, 1990 FORREST, S. I., VERMA, D. P. S., and DHINDSA, R. S. 1991. Starch content and activities of starch-metabolizing enzymes in effective and ineffective root nodules of soybean. Can. J. Bot. 69: 697-701. Starch content and activities of some enzymes of starch metabolism were determined in wild-type, N2-fixing (fix') nodules and in two non-N2-fixing (fix-) nodules induced by Bradyrhizobium japonicum mutant strains, T5-95 and T8-1, on soybean (Glycine max L.) roots. The T5-95 nodules are similar to wild type in ultrastructure, but the T8-1 nodules are different in that the bacteroids are not released from the infection thread. After initial accumulation to relatively high concentration, starch was depleted during nitrogen fixation in fix' nodules. However, in fix- nodules, the accumulated starch was not metabolized. The activity of starch-bound starch synthase (EC 2.4.1.21) declined in fix' nodules but remained high in fix- nodules. The activity of a-amylase (EC 3.2.1.1) was only slightly higher than wild type in T5-95 but was four times higher than wild type in T8-1 nodules. The activity of starch phosphorylase (EC 2.4.1.1) increased in all nodule types from 14 to 21 days postinfection. A positive correlation was observed between the capacity of nodules to fix N2 and their capacity to degrade starch. Collectively, these results support the concept that starch accumulated during early stages of nodule devel- opment is metabolized to supply energy for nitrogen fixation and to meet the metabolic demands of bacteroids. Key words: nitrogen fixation, starch content, effective and ineffective nodules, starch synthase, starch phosphorylase, a-amylase. FORREST, S. I., VERMA, D. P. S., et DHINDSA, R. S. 1991. Starch content and activities of starch-metabolizing enzymes in effective and ineffective root nodules of soybean. Can. J. Bot. 69 : 697-701. Les auteurs ont dCterminC le contenu en amidon et I'activitC de certaines enzymes du mCtabolisme de I'amidon chez des nodules fixateurs d'azote (fix') de type sauvage ainsi que chez les nodules non-fixateurs d'azote (fix-) produits par des souches mutantes du Bradyrhizobium japonicum, T5-95 et T8-1, sur racines de feve soja (Glycine max L.). Les nodules de type T5-95 ont une ultrastructure semblable, mais different en ce que les bactCroi'des ne quittent pas le cordon d'infection. Aprbs une pCriode initiale d'accumulation pour atteindre des concentrations relativement ClevCes, I'amidon disparait par la suite au cours de la fixation de I'azote par les nodules fix'. Cependant, chez les nodules fix- I'amidon accumulC n'est pas mCtabolisC. Chez les nodules fix', les activitCs de la synthase de I'amidon like i I'amidon (EC 2.4.1.21) diminuent mais demeurent ClevCes dans les nodules fix-. L'activitC de I'a-amylase (EC 3.2.1.1) n'est que lkgbrement plus forte que chez le type sauvage dans les nodules T5-95, mais est quatre fois plus ClevCe que chez le type sauvage dans le cas des nodules T8-1. L'activitC de la phosphorylase de l'amidon (EC 2.4.1.1) augmente dans tous les types de nodules, du 14 i 21 jours aprbs la colonisation. On observe une corrClation positive entre la capacitC des nodules i fixer I'azote et leur capacitk i dCgrader I'amidon. Mis ensemble, ces rCsultats supportent I'idCe que I'amidon accumulC pendant les premiers stades du dCveloppement du nodule est mCtabolisC pour fournir 1'Cnergie nkcessaire i la fixation de l'azote et pour rencontrer les besoins mCtaboliques des bactCroTdes. Mots clts : fixation de I'azote, contenu en amidon, nodules efficaces et non efficaces, synthase de l'amidon, phosphorylase - - de l'amidon, a-amylase. [Traduit par la rCdaction] Introduction Wild-type Bradyrhizobium japonicum induces effective (capable of fixing N,) nodulation on roots of soybean. Pho- tosynthates, primarily sucrose, are translocated to the nodule and support the energy requirements of the bacteroids, and the excess photosynthates are stored as starch (19, 20). Although much work has been canied out on symbiotic N, fixation, little attention has been paid to the carbohydrate metabolism within the nodule. We know that one of the genes involved in nodule metabolism, nodulin-100, has been identified as a nodule- specific form of sucrose synthase (19). Regulation of this enzyme by heme appears to play a role in controlling the flow of carbon to bacteroids. A high activity of this enzyme during early stages of nodule development facilitates accumulation of starch. There is evidence that photosynthate supply to the nodule affects its level of N,-fixing activity (2, 7, 23). Thus starch might be expected to play an important role as a source of energy for N, fixation. There have been studies on the distri- bution of starch between infected and uninfected cells within the nodule (5). However, different cell types within an effec- tive nodule may not be comparable, since they are likely to exhibit a metabolic division of labour. Although enzyme activ- ities involved in nitrogen fixation, glycolysis, and amino acid synthesis have been studied (6), those related to starch metab- olism have received little attention. Herein we compare the 'Present address: Environment Canada, Division of Hydrology and Networks, Centre Sainte-Laurent, 105 McGill St. Suite 400, Mont- starch content and the activities of some starch-metabolizing real, Que., Canada H2Y 2E7. enzymes between effective (fix') nodules induced by wild- 2Present address: Department of Molecular Genetics and the Ohio type strain 61A76 of B. japonicum and two types of ineffective State Biotechnology Center, Ohio State University, Columbus, OH (fix-) nodules induced by mutant strains, T5-95 andT8-1. The 43210-1002, U.S.A. latter were obtained through Tn5 mutagenesis of the wild-type 3Author to whom correspondence should be addressed. strain (13). Nodules induced by T5-95 are similar to wild-type Printed in Canada / Imprim6 au Canada Can. J. Bot. 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