Interactions among Flavonoid <i>nod</i> Gene Inducers Released from Alfalfa Seeds and Roots

Hartwig, Ueli A.; Maxwell, Carl A.; Joseph, Cecillia M.; Phillips, Donald A.

doi:10.1104/pp.91.3.1138

Cited by 59 publications

(31 citation statements)

References 18 publications

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“…3), and those exuded by roots, which have 4',7-substitution patterns (20), may be important for understanding how rhizosphere chemistry influences R. meliloti and symbiotic N2 fixation in alfalfa. For example, combinations of luteolin, the dominant nod gene inducer from seeds, and 4,4'-dihydroxy-2'-methoxychalcone, the strong nod gene inducer from alfalfa root exudates, produced a synergistic increase in transcription of the nodABC-lacZ fusion in R. meliloti 102lpRmM57 (13). Since it is known that R. meliloti has three functional copies of nodD (12,15), that effect may operate through the capacity of these molecules to interact with different nodD genes (UA Hartwig, unpublished data).…”

Section: Discussionmentioning

confidence: 99%

Chrysoeriol and Luteolin Released from Alfalfa Seeds Induce nod Genes in Rhizobium meliloti

Hartwig¹,

Maxwell²,

Joseph³

et al. 1990

Plant Physiol.

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157

101

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Flavonoid signals from alfalfa (Medicago sativa L.) seed and root exudates induce transcription of nodulation (nod) genes in Rhizobium meliloti. The flavone luteolin previously was isolated from alfalfa seeds by other workers and identified as the first nod gene inducer for R. meliloti. Our recent study of 'Moapa 69' alfalfa root exudates found no luteolin but did identify three other nod gene inducers: 4,4'-dihydroxy-2'-methoxychalcone, 4',7-dihydroxyflavone, and 4',7-dihydroxyflavanone. The goal of the current study was to identify and quantify nod gene-inducing flavonoids that may influence Rhizobium populations around a germinating alfalfa seed. Aqueous rinses of Moapa 69 alfalfa seeds were collected and assayed for induction of a nodABClacZ fusion in R. meliloti. During the first 4 hours of imbibition, total nod gene-inducing activity was released from seeds at 100-fold higher rates than from roots of 72-hour-old seedlings. Five flavonoids were purified and identified by spectroscopic analyses (ultraviolet/visible absorbance, proton nuclear magnetic resonance, and mass spectroscopy) and comparison with authentic standards. Two very active nod gene-inducing flavonoids, chrysoeriol (3'-methoxyluteolin) and luteolin, were identified in seed rinses. Luteolin required a higher concentration (18 nanomolar) than chrysoeriol (5 nanomolar) for half-maximum induction of nodABC-IacZ in R. meliloti, and both were less active than 4,4'-dihydroxy-2'-methoxychalcone (2 nanomolar) from root exudates. Seeds exuded three other luteolin derivatives: luteolin-7-0-glucoside, 5-methoxyluteolin, and 3',5-dimethoxyluteolin. Their combined quantities were 24-fold greater than that of luteolin plus chrysoeriol. Most nod gene-inducing activity of these luteolin derivatives apparently is associated with degradation to luteolin and chrysoeriol. However, their presence in large quantities suggests that they may contribute significantly to nod gene-inducing activity in the soil. rial nodulation genes nodABC (22), rhizobial products induce root hair curling (17) and cortical cell divisions (8). The first molecule showing nod gene-inducing activity in R. meliloti was isolated from alfalfa seeds and identified as 3',4',5,7-tetrahydroxyflavone, a compound known as luteolin (23). Recent studies of root exudates from 72-h-old 'Moapa 69' alfalfa seedlings identified 4,4'-dihydroxy-2'-methoxychalcone, 4',7-dihydroxyflavone, and 4',7-dihydroxyflavanone as active nod gene inducers, but no luteolin was detected (20). Numerous flavonoids from various legumes have been reported as active nod gene inducers (4,9,18,25,26,29), but qualitative and quantitative differences between compounds actually released from seeds and roots have not been described.The presence of particular flavonoids inside plants cannot be taken as evidence of their release into exudates. Yelton et al. (28) observed that extracts, but not exudates, ofsome plant species induced nod genes in R. meliloti. Presumably, host plant factors controlling the synthesis and release off...

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Section: Discussionmentioning

confidence: 99%

Chrysoeriol and Luteolin Released from Alfalfa Seeds Induce nod Genes in Rhizobium meliloti

Hartwig¹,

Maxwell²,

Joseph³

et al. 1990

Plant Physiol.

Self Cite

157

101

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“…Flavonoids that do not stimulate nod gene induction can act as inhibitors that are capable of antagonizing nod gene expression (13,84). In the soil, rhizobia are exposed to a variety of flavonoids, and so nod gene expression is likely affected by a combination of stimulatory and inhibitory interactions (12,13,15,30).No direct biochemical evidence has been reported for flavonoid binding to NodD, but several lines of genetic evidence indicate that NodD is directly involved in flavonoid perception. First, a functional nodD gene is both necessary and sufficient to mediate nod gene induction in the presence of flavonoids (49,83).…”

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confidence: 99%

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Diverse Flavonoids Stimulate NodD1 Binding tonodGene Promoters inSinorhizobium meliloti

2006

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NodD1 is a member of the NodD family of LysR-type transcriptional regulators that mediates the expression of nodulation (nod) genes in the soil bacterium Sinorhizobium meliloti. Each species of rhizobia establishes a symbiosis with a limited set of leguminous plants. This host specificity results in part from a NodD-dependent upregulation of nod genes in response to a cocktail of flavonoids in the host plant's root exudates. To demonstrate that NodD is a key determinant of host specificity, we expressed nodD genes from different species of rhizobia in a strain of S. meliloti lacking endogenous NodD activity. We observed that nod gene expression was initiated in response to distinct sets of flavonoid inducers depending on the source of NodD. To better understand the effects of flavonoids on NodD, we assayed the DNA binding activity of S. meliloti NodD1 treated with the flavonoid inducer luteolin. In the presence of luteolin, NodD1 exhibited increased binding to nod gene promoters compared to binding in the absence of luteolin. Surprisingly, although they do not stimulate nod gene expression in S. meliloti, the flavonoids naringenin, eriodictyol, and daidzein also stimulated an increase in the DNA binding affinity of NodD1 to nod gene promoters. In vivo competition assays demonstrate that noninducing flavonoids act as competitive inhibitors of luteolin, suggesting that both inducing and noninducing flavonoids are able to directly bind to NodD1 and mediate conformational changes at nod gene promoters but that only luteolin is capable of promoting the downstream changes necessary for nod gene induction.The soil bacterium Sinorhizobium meliloti and the leguminous plant alfalfa form a symbiotic relationship that results in the differentiation of S. meliloti into nitrogen-fixing bacteroids that reside in nodules formed on alfalfa roots. To initiate the symbiosis, alfalfa roots and seeds excrete a cocktail of nodulation-inducing molecules composed predominantly of flavonoids (15,34,60). In response, NodD proteins in S. meliloti activate transcription of nod genes, which encode the enzymes responsible for the synthesis of Nod factor, a lipochito-oligosaccharide signal required for symbiotic development in alfalfa (17). The genome of S. meliloti encodes three NodD polypeptides, NodD1, NodD2, and NodD3, that share greater than 77% amino acid identity (29,32,48,67). NodD1 and NodD2 require plant-derived inducers for activity (49), while NodD3, when overexpressed, is active in the absence of flavonoids (33,48).Each species of rhizobia establishes a symbiosis with a limited set of host plants depending in part on the cocktail of flavonoids in the plant exudate. For example, S. meliloti nodulates alfalfa, while Rhizobium leguminosarum bv. trifolii nodulates clover (64). High-performance liquid chromatography (HPLC) analysis demonstrates that most legume seed and root exudates contain approximately 10 different flavonoid compounds (59, 81). Flavonoids that do not stimulate nod gene induction can act as inhibitors that are capab...

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“…S. meliloti-expressing vector alone gave Ͻ16. 5 Miller units of ␤-galactosidase activity under all conditions.…”

Section: Screen For Nodd1 Alleles Showing Altered Responses To Luteolinmentioning

confidence: 93%

“…In response to flavonoids, members of the NodD family of transcriptional activators induce expression of the nod genes (2, 3). NodD1 and NodD2 require plant-derived inducers for activity (such as luteolin, dihydroxymethoxychalcone [MCh], and chrysoeriol for NodD1 and trigonelline, stachydrine, and MCh for NodD2), while NodD3 is constitutively active when overexpressed (3)(4)(5)(6)(7). The nod genes encode enzymes that synthesize lipochitooligosaccharide host-signaling compounds known as Nod factors (NFs) (8,9).…”

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Isolation and Characterization of Mutant Sinorhizobium meliloti NodD1 Proteins with Altered Responses to Luteolin

et al. 2013

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c NodD1, a member of the NodD family of LysR-type transcriptional regulators (LTTRs), mediates nodulation (nod) gene expression in the soil bacterium Sinorhizobium meliloti in response to the plant-secreted flavonoid luteolin. We used genetic screens and targeted approaches to identify NodD1 residues that show altered responses to luteolin during the activation of nod gene transcription. Here we report four types of NodD1 mutants. Type I (NodD1 L69F, S104L, D134N, and M193I mutants) displays reduced or no activation of nod gene expression. Type II (NodD1 K205N) is constitutively active but repressed by luteolin. Type III (NodD1 L280F) demonstrates enhanced activity with luteolin compared to that of wild-type NodD1. Type IV (NodD1 D284N) shows moderate constitutive activity yet can still be induced by luteolin. In the absence of luteolin, many mutants display a low binding affinity for nod gene promoter DNA in vitro. Several mutants also show, as does wild-type NodD1, increased affinity for nod gene promoters with added luteolin. All of the NodD1 mutant proteins can homodimerize and heterodimerize with wild-type NodD1. Based on these data and the crystal structures of several LTTRs, we present a structural model of wildtype NodD1, identifying residues important for inducer binding, protein multimerization, and interaction with RNA polymerase at nod gene promoters.

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Interactions among Flavonoid nod Gene Inducers Released from Alfalfa Seeds and Roots

Cited by 59 publications

References 18 publications

Chrysoeriol and Luteolin Released from Alfalfa Seeds Induce nod Genes in Rhizobium meliloti

Chrysoeriol and Luteolin Released from Alfalfa Seeds Induce nod Genes in Rhizobium meliloti

Diverse Flavonoids Stimulate NodD1 Binding tonodGene Promoters inSinorhizobium meliloti

Isolation and Characterization of Mutant Sinorhizobium meliloti NodD1 Proteins with Altered Responses to Luteolin

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