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Journal of Bacteriology, February 2001, p. 854-864, Vol. 183, No. 3
0021-9193/01/$04.00+0   DOI: 10.1128/JB.183.3.854-864.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

Enhanced Symbiotic Performance by Rhizobium tropici Glycogen Synthase Mutants

Silvia Marroquí,1,2 Angeles Zorreguieta,1 Carmen Santamaría,3 Francisco Temprano,3 Mario Soberón,4 Manuel Megías,2 and J. Allan Downie1,*

John Innes Centre, Norwich NR4 7UH, United Kingdom1; Departamento de Microbiología y Parasitología, Universidad de Sevilla,2 and CIFA "Las Torres y Tomejil," Alcalá del Río,3 Sevilla, Spain; and Departamento de Biología Molecular de Plantas, Instituto de Biotecnología, UNAM, Cuernavaca, Morelos, México4

Received 28 July 2000/Accepted 7 November 2000

We isolated a Tn5-induced Rhizobium tropici mutant that has enhanced capacity to oxidize N,N-dimethyl-p-phenylendiamine (DMPD) and therefore has enhanced respiration via cytochrome oxidase. The mutant had increased levels of the cytochromes c1 and CycM and a small increase in the amount of cytochrome aa3. In plant tests, the mutant increased the dry weight of Phaseolus vulgaris plants by 20 to 38% compared with the control strain, thus showing significantly enhanced symbiotic performance. The predicted product of the mutated gene is homologous to glycogen synthases from several bacteria, and the mutant lacked glycogen. The DNA sequence of the adjacent gene region revealed six genes predicted to encode products homologous to the following gene products from Escherichia coli: glycogen phosphorylase (glgP), glycogen branching enzyme (glgB), ADP glucose pyrophosphorylase (glgC), glycogen synthase (glgA), phosphoglucomutase (pgm), and glycogen debranching enzyme (glgX). All six genes are transcribed in the same direction, and analysis with lacZ gene fusions suggests that the first five genes are organized in one operon, although pgm appears to have an additional promoter; glgX is transcribed independently. Surprisingly, the glgA mutant had decreased levels of high-molecular-weight exopolysaccharide after growth on glucose, but levels were normal after growth on galactose. A deletion mutant was constructed in order to generate a nonpolar mutation in glgA. This mutant had a phenotype similar to that of the Tn5 mutant, indicating that the enhanced respiration and symbiotic nitrogen fixation and decreased exopolysaccharide were due to mutation of glgA and not to a polar effect on a downstream gene.

* Corresponding author. Mailing address: Department of Genetics, John Innes Centre, Colney Lane, Norwich NR4 7UH, United Kingdom. Phone: 44 1603 450000. Fax: 44 1603 450045. E-mail: allan.downie{at}bbsrc.ac.uk.

Journal of Bacteriology, February 2001, p. 854-864, Vol. 183, No. 3
0021-9193/01/$04.00+0   DOI: 10.1128/JB.183.3.854-864.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.


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