Involvement of a Soybean ATP-Binding Cassette-Type Transporter in the Secretion of Genistein, a Signal Flavonoid in Legume-Rhizobium Symbiosis

Abstract

Legume plants have an ability to fix atmospheric nitrogen into nutrients via symbiosis with soil microbes. As the initial event of the symbiosis, legume plants secrete flavonoids into the rhizosphere to attract rhizobia. Secretion of flavonoids is indispensable for the establishment of symbiotic nitrogen fixation, but almost nothing is known about the membrane transport mechanism of flavonoid secretion from legume root cells. In this study, we performed biochemical analyses to characterize the transport mechanism of flavonoid secretion using soybean (Glycine max) in which genistein is a signal flavonoid. Plasma membrane vesicles prepared from soybean roots showed clear transport activity of genistein in an ATP-dependent manner. This transport activity was inhibited by sodium orthovanadate, a typical inhibitor of ATP-binding cassette (ABC) transporters, but was hardly affected by various ionophores, such as gramicidin D, nigericin, or valinomycin, suggesting involvement of an ABC transporter in the secretion of flavonoids from soybean roots. The K m and V max values of this transport were calculated to be 158 mM and 322 pmol mg protein 21 min 21 , respectively. Competition experiments using various flavonoids of both aglycone and glucoside varieties suggested that this ABC-type transporter recognizes genistein and daidzein, another signaling compound in soybean root exudates, as well as other isoflavonoid aglycones as its substrates. Transport activity was constitutive regardless of the availability of nitrogen nutrition. This is, to our knowledge, the first biochemical characterization of the membrane transport of flavonoid secretion from roots.Plant root exudates secreted into the soil mediate complex interactions between soil-born microorganisms and plants. These interactions occur in the small region around the plant roots, which is called the rhizosphere, where plants protect root tissues from attack by bacteria and fungi and symbiotic interactions with rhizobia and arbuscular mycorrhiza are also involved in a cooperative context. Examples of root exudates responsible for the protective actions include cyclic hydroxamic acids, such as 2,4-dihydroxy-7-methoxy-1, 4-benzoxazin-3-one and the naphthoquinone-derivative shikonin [5,8-dihydroxy-2-(1-hydroxy-4-methylpent-3-en-1-yl)-1,4-naphthoquinone], which were reported in wheat (Triticum aestivum) and Lithospermum erythrorhizon, respectively (Niemeyer, 1988;Brigham et al., 1999). The well-known phenomenon called allelopathy, in which organic compounds in the root exudates or secondary metabolites produced in the aerial part of the plant cause deleterious effects on neighboring plants, thus suppressing the growth of other plants, is an example of interactions between plants in the rhizosphere (Weir et al., 2004). In contrast to the negative interactions between plants and other organisms, positive interactions between plants and microbes also occur in the rhizosphere. Flavonoids in root exudates of legume plants activate nod gene expression of rhizobia (Peters et a...

Spatio Temporal Influence of Isoflavonoids on Bacterial Diversity in the Soybean Rhizosphere

“…However, rhizodeposition of isoflavonoids occurs through mechanisms other than root exudation as well, e.g., root border cells (Hawes et al 2000). Therefore, silencing components of root exudation machinery (e.g., ABC transporters [Brechenmacher et al 2009;Sugiyama et al 2007]) might not result in efficient depletion of isoflavonoids in rhizodeposits. Similarly, the use of adsorbents might disrupt quorum signals between bacteria and cause nonspecific modifications in root-surface microbial communities and exert unpredicted, indirect impacts on nontarget organisms, including the plant producing the flavonoids (Hassan and Mathesius 2012).…”

Flavonoids in plant rhizospheres: secretion, fate and their effects on biological communication

“…Using plasma membrane vesicles prepared from soybean roots, these roots were shown to possess an ATPdependent transporter that functions in the secretion of genistein into the rhizosphere (Sugiyama et al 2007). Biochemical transport assays using various inhibitors have suggested that an ABC transporter is the primary candidate for this process.…”

Soy, phytoestrogens and their impact on reproductive health

“…Isoflavones play important roles as signal molecules for inhibiting pathogen attacks (Lozovaya et al, 2007;Zabala et al, 2006) and for plant-microbe symbiotic interactions (Subramanian et al, 2006;Sugiyama et al, 2007). Genistin and daidzin is initiated by the conversion of the essential amino acid L-phenylalanine into p-Coumaroyl-CoA.…”