Distribution of O-acetyl groups in the exopolysaccharide synthesized by Rhizobium leguminosarum strains is not determined by the Sym plasmid

H.C Cremers,M Batley,J.W Redmond,A.H Wijfjes,B.J Lugtenberg,C.A Wijffelman

doi:10.1016/s0021-9258(18)92856-5

Abstract

The patterns of O-acetylation of the exopolysaccharide (EPS) from the Sym plasmid-cured derivatives of Rhizobium leguminosarum bv. trifolii strain LPR5, R. leguminosarum bv. trifolii strain ANU843 and R. leguminosarum bv. viciae strain 248 were determined by 1H and 13C NMR spectroscopy. Beside a site indicative of the chromosomal background, these strains have one site of O-acetylation in common, namely residue b of the repeating unit. The O-acetyl esterification pattern of EPS of the Sym plasmid-cured derivatives of strains LPR5, ANU843, and 248 was not altered by the introduction of a R. leguminosarum bv. viciae Sym plasmid or a R. leguminosarum bv. trifolii Sym plasmid. The induction of nod gene expression by growth of the bacteria in the presence of Vicia sativa plants or by the presence of the flavonoid naringenin, produced no significant changes in either amount or sites of O-acetyl substitution. Furthermore, no such changes were found in the EPS from a Rhizobium strain in which the nod genes are constitutively expressed. The substitution pattern of the exopolysaccharide from R. leguminosarum is, therefore, determined by the bacterial genome and is not influenced by genes present on the Sym plasmid. This conclusion is inconsistent with the suggestion of Philip-Hollingsworth et al. (Philip-Hollingsworth, S., Hollingsworth, R. I., Dazzo, F. B., Djordjevic, M. A., and Rolfe, B. G. (1989) J. Biol. Chem. 264, 5710-5714) that nod genes of R. leguminosarum bv. trifolii, by influencing the acetylation pattern of EPS, determine the host specificity of nodulation.

Highlights

Introduction of theSym plasmidof another cross-inocula- moters in front of the inducible nod genes present on the R.tion group, namelythe R. leguminosarum bv. trifolii Sym leguminosarum bv. uiciae Sym plasmidpRLlJI in thaebsence plasmid pSym5, didnotinfluencethesiteoramount of of flavonoids (Spaink et al, 1989a)
In order to study the influence of expression of nodulation genes on the structureof EPS synthesized by R. leguminosarum biovar strains, we first established the basic chemical structure of the EPS isolated from Sym plasmid3-cured derivatives of strains LPR5, ANU843
All the shiftswere 'H (Fig. 3B) and 13CNMR spectra (Fig. 2C) of the repeating less than 0.05 ppm and were temperature dependent (Fig. 5) unit of theEPS isolated from R. leguminosarum strain and were too small for the acetylation to be on ANU845 contained all the peaks assigned to the presence of residue a

Summary

MATERIALS AND METHODS’

Attention has been paid tothe possibility that rhizobia1exopolysaccharide (EPS)’is involved in nodulation. Several authors described the isolation of mutants affected in the synthesis of EPS thaatlso showed defects in nodulation ability. Leigh et al (1985) described numerous Exo- mutants of Rhizobium meliloti that all induce nonnitrogen-fixing nodules. Diebold and Noel (1989) described the isolation of Exo- mutants of R. leguminosarumbv.

RESULTS

DISCUSSION

Materials and Methods

Results