Involvement of nodS in N-Methylation and nodU in 6-O-Carbamoylation of Rhizobium sp. NGR234 Nod Factors

Abstract

Although Rhizobium sp. NGR234 and Rhizobium fredii USDA257 share many traits, dysfunctional nodSU genes in the latter prohibit nodulation of Leucaena species. Accordingly, we used R. fredii transconjugants harboring the nodS and nodU genes of NGR234 to study their role in the structural modification of the lipo-oligosaccharide Nod factors. Differences between the Nod factors mainly concern the length of the oligomer (three to five glucosamine residues in USDA257 and five residues only in NGR234) and the presence of additional substituents in NGR234 (N-linked methyl, one or two carbamoyl groups on the non-reducing moiety, acetyl or sulfate groups on the fucose). R. fredii(nodS) transconjugants produce chitopentamer Nod factors with a Nlinked methyl group on the glucosaminyl terminus. Introduction of nodU into USDA257 results in the formation of 6-O-carbamoylated factors. Co-transfer of nodSU directs N-methylation, mono-6-O-carbamoylation, and production of pentameric Nod factors. Mutation of nodU in NGR234 suppresses the formation of bis-carbamoylated species. Insertional mutagenesis of nodSU drastically decreases Nod factor production, but with the exception of sulfated factors (which are partially N-methylated and mono-carbamoylated), they are identical to those of the wild-type strain. Thus, Nod factor levels, their degree of oligomerization, and N-methylation are linked to the activity encoded by nodS.

Molecular Basis of Symbiotic Promiscuity

“…In contrast, the NGR⌬nodSU mutant, which secretes only 1/10 the amount of Nod-factor produced by the wild type, is incapable of nodulating L. leucocephala (Table 5). One interpretation of these data is that higher levels of Nod factors (produced when a functional NodS or NodU is present) (129), not the presence of N-methylated or 6-O-carbamoylated LCOs, is necessary for nodulation of Leucaena.…”

“…NodS is an N-methyltransferase (93,94,129), while NodU and NolO control carbamoylation at C-6 and C-3 (or C-4), respectively, on the nonreducing N-acetyl-D-glucosamine (57,129,131). Inactivation of nodS in A. caulinodans, strain NGR234, and R. tropici abolishes the nodulation of L. leucocephala and P. vulgaris (158,276).…”

“…Introduction of either nodS or nodU into R. fredii extends its host range to include Leucaena spp. (129,144).…”

Functional redundancy of genes for sulphate activation enzymes in Rhizobium sp. BR816

“…On the non-reducing end, the nods (Geelen et al, 1993), nodL (Downie, 1989) and nodU (Jabbouri et al, 1995) genes have been shown to be responsible for N-methyl (Price et a!., 19921, 0-acetyl (Carlson et al, 1993) or 0-carbamoyl (Price et al, 1992 substitutions, respectively. On the reducing end, sugar modifications such as a fucosyl residue (Price et al, 1992;Sanjuan et al, 1992), an arabinosyl residue (Mergaert et al, 1993) or a mannosyl moiety (Folch-Mallol et al, 1996) can be present (for an overview of Nod factor structures and nodulation genes, see Denarie et al, 1996).…”

Sulphation of Rhizobium sp. NGR234 Nod factors is dependent on noeE, a new host‐specificity gene

“…Elution was performed with 9:1 (v/v) methanol/5.5 M ammonia. Separation of LCOs by HPLC was performed as described previously (Jabbouri et al, 1995). Mass spectra were recorded on an Autospec instrument (Fisons, VG-Analytical) fitted with a caesium-ion gun working at 20 kV.…”