NolL of Rhizobium sp. Strain NGR234 Is Required for O -Acetyltransferase Activity

Abstract

Following (iso)flavonoid induction, nodulation genes of the symbiotic nitrogen-fixing bacterium Rhizobium sp. strain NGR234 elaborate a large family of lipooligosaccharidic Nod factors (NodNGR factors). When secreted into the rhizosphere of compatible legumes, these signal molecules initiate root hair deformation and nodule development. The nonreducing glucosamine residue of NodNGR factors are N acylated, N methylated, and mono- or biscarbamoylated, while position C-6 of the reducing extremity is fucosylated. This fucose residue is normally 2-O methylated and either sulfated or acetylated. Here we present an analysis of all acetylated NodNGR factors, which clearly shows that the acetate group may occupy position C-3 or C-4 of the fucose moiety. Disruption of the flavonoid-inducible nolL gene, which is preceded by anod box, results in the synthesis of NodNGR factors that lack the 3-O- or 4-O-acetate groups. Interestingly, the nodulation capacity of the mutant NGRΩnolL is not impaired, whereas introduction of thenod box::nolL construct into the related strain Rhizobium fredii USDA257 extends the host range of this bacterium to Calopogonium caeruleum,Leucaena leucocephala, and Lotus halophilus. Nod factors produced by a USDA257(pnolL) transconjugant were also acetylated. The nodbox::nolL construct was also introduced into ANU265 (NGR234 cured of its symbiotic plasmid), along with extra copies of the nodD1 gene. When permeabilized, these cells possessed acetyltransferase activity, although crude extracts did not.

Flavonoids induce temporal shifts in gene‐expression of nod‐box controlled loci in Rhizobium sp. NGR234

“…1). NB4, which regulates the O‐acetyltransferase encoded by nolL (Berck et al ., 1999), is more active than NB7 that is upstream of the sulphuryltransferase encoded by noeE (Fig. 2D and G) (Hanin et al ., 1997).…”

“…data). pMP220 constructs containing nod ‐boxes ♯2 (pMP‐NB2), ♯4 (pMP‐NB4), ♯7 (pMP‐NB7), ♯8 (pMP‐NB8) and ♯12 (pMP‐NB12) correspond to plasmids pMP220‐NB (Fellay et al ., 1995), pNB nolL (Berck et al ., 1999), pNB noeE (Hanin et al ., 1997), pNBA (Fellay et al ., 1998), and pA27 (Lewin et al ., 1990) respectively. The size and position of the cloned nod ‐box promoters in relation to their predicted protein‐coding regions on pNGR234 a (complete DNA sequence GenBank accession ♯ NC_000914) are shown in Fig.…”

High‐resolution transcriptional analysis of the symbiotic plasmid of Rhizobium sp. NGR234

“…Among these were nolL and noeE (Fig. 1, SH, panel hA to hJ), which are responsible for the acetylation and sulphatation of NodNGR factors, respectively (Hanin et al, 1997;Berck et al, 1999), as well as a series of ORFs that comprise fixF to y4hA (Fig. 2, SH, segment G and H).…”

Reduction of bacterial genome size and expansion resulting from obligate intracellular lifestyle and adaptation to soil habitat.

“…At least in part, it re sults from NGR234 unique abil ity to pro duce a much higher num ber of var i ous Nod fac tors (mostly dif fer ing at their re duc ing end) than any other rhizobium sp. (Berck et al, 1999;Perret et al, 2000;Price et al, 1992;Quesada-Vincens et al, 1998). Nev er the less, this broad host range must be de ter mined by some un rec og nized fac tors as well, since the closely re lated S. fredii (shares > 95% sequence iden tity with nodulation genes of NGR234) nodulates many spe cies of Leguminosae, even though it pro duces only one or two types of Nod fac tors.…”

“…Proba bly, most le gumes nodulated by these two rhizobium spe cies are pro mis cu ous plants that tol er ate var i ous NFs. Nev er the less, nodulation of cer tain le gumes re quires the pres ence of in trin sic mod i fi ca tions con ferred by host spec i fic ity genes, which are pres ent ex clu sively in NGR234 (Berck et al, 1999;Hanin et al, 1997).…”