Flavone-enhanced accumulation and symbiosis-related biological activity of a diglycosyl diacylglycerol membrane glycolipid from Rhizobium leguminosarum biovar trifolii

Abstract

Rhizobium leguminosarum bv. trifolii is the bacterial symbiont which induces nitrogen-fixing root nodules on the leguminous host, white clover (Trifolium repens L.). In this plant-microbe interaction, the host plant excretes a flavone, 4',7-dihydroxyflavone (DHF), which activates expression of modulation genes, enabling the bacterial symbiont to elicit various symbiosis-related morphological changes in its roots. We have investigated the accumulation of a diglycosyl diacylglycerol (BF-7) in wild-type R. leguminosarum bv. trifolii ANU843 when grown with DHF and the biological activities of this glycolipid bacterial factor on host and nonhost legumes. In vivo labeling studies indicated that wild-type ANU843 cells accumulate BF-7 in response to DHF, and this flavone-enhanced alteration in membrane glycolipid composition was suppressed in isogenic nodA::Tn5 and nodD::Tn5 mutant derivatives. Seedling bioassays performed under microbiologically controlled conditions indicated that subnanomolar concentrations of purified BF-7 elicit various symbiosis-related morphological responses on white clover roots, including thick short roots, root hair deformation, and foci of cortical cell divisions. Roots of the nonhost legumes alfalfa and vetch were much less responsive to BF-7 at these low concentrations. A structurally distinct diglycosyl diacylglycerol did not induce these responses on white clover, indicating structural constraints in the biological activity of BF-7 on this legume host. In bioassays using aminoethoxyvinylglycine to suppress plant production of ethylene, BF-7 elicited a meristematic rather than collaroid type of mitogenic response in the root cortex of white clover. These results indicate an involvement of flavone-activated nod expression in membrane accumulation of BF-7 and a potent ability of this diglycosyl diacylglycerol glycolipid to perform as a bacterial factor enabling R. leguminosarum bv. trifolii to activate segments of its host's symbiotic program during early development of the root nodule symbiosis.