Primary structure and differential expression of glutamine synthetase genes in nodules, roots and leaves of Phaseolus vulgaris

Abstract

In plants, glutamine synthetase (GS) is the enzyme primarily responsible for the assimilation of ammonia into organic nitrogen. In Phaseolus vulgaris a number of isoenzymic forms of GS are found, each of which consists of eight subunits of mol. wt 41 000‐45 000. The GS subunits of P. vulgaris have previously been shown to be encoded by a small multigene family and a partial cDNA clone for a nodule‐specific GS subunit has been obtained. We report here the isolation and nucleotide sequencing of two essentially full‐length GS cDNA clones (pR‐1 and pR‐2) from a root cDNA library and the deduced amino acid sequences of the corresponding GS subunits (355 amino acid residues each). The coding sequences of pR‐1 and pR‐2 are closely related (80% nucleotide homology, 88% amino acid homology), but their 5′‐ and 3′‐untranslated regions have diverged almost completely. Both pR‐1 and pR‐2 are related to, but distinct from, the nodule GS clone, pcPvNGS‐01 (or pN‐1). Hybridization to genomic Southern blots showed that the three GS mRNAs are encoded by three seperate genes and indicated the existence of a fourth class of GS gene. An S1 nuclease protection assay demonstrated the presence of R‐1 and R‐2 mRNA in both roots and leaves and confirmed that expression of the N‐1 gene is nodule‐specific. Expression of the R‐1 and R‐2 genes in the roots did not change significantly during nodulation. However, only the R‐1 gene is expressed in the nodules themselves, indicating that the R‐2 gene is specifically repressed during nodule development.

Novel Expression Pattern of Cytosolic Gln Synthetase in Nitrogen-Fixing Root Nodules of the Actinorhizal Host, Datisca glomerata

“…napus-3, cyt (root) Temple et al, 1995;Nicotiana plumbaginifolia, cp, M19055, Tingey and Coruzzi, 1987;Nicotiana sylvestris, cp, X66940, Becker et al, 1992; Nicotiana tabacum-1, cyt (seedling), X95932, Dubois et al, 1996; N. tabacum-2, cyt (seedling), X95933, Dubois et al, 1996; Oryza sativa, cyt (shoot), X14245, Sakamoto et al, 1989; O. sativa, cyt (derived from genomic sequence), AB037595, Kojima et al, 2000;O. sativa, cp, X14246, Sakamoto et al, 1989; Pinus sylvestris, cyt (seedling), X74429, Elmlinger et al, 1994; Pisum sativum-1, cyt (nodule), X05515, ; P. sativum-2, cyt (nodule), X04763, ; P. sativum 3 cyt (nodule) M20663, Tingey et al, 1988; P. sativum cp, M20664, Tingey et al, 1988; Phaseolus vulgaris-1, cyt (root), X04001, Gebhardt et al, 1986; P. vulgaris-2, cyt (root), X04002, Gebhardt et al, 1986;P. vulgaris, cp, X12738, Lightfoot et al, 1988; P. vulgaris gamma, cyt (nodule) X14605, ; Populus alba 3 Populus tremula cyt AF236074, Wu, D. and Kirby, E.G.…”

“…The placement of DgGS1-1 within this assemblage is moderately well supported with a decay value of three. This phylogenetic analysis also includes legume nodule GS sequences that group according to a, b, and g subclasses identified previously (Gebhardt et al, 1986;Morey et al, 2002). Based on this analysis, the legume b and g subclasses appear to be most closely related to each other, and DgGS1-1 is most closely associated with the a subclass of legume GS sequences.…”

Cell-specific expression in transgenic plants reveals nonoverlapping roles for chloroplast and cytosolic glutamine synthetase.

“…The role of chloroplast GS2 in the reassimilation of photorespiratory ammonia is supported by the analysis of mutants in barley (7) and is substantiated by gene expression studies in pea (17). For cytosolic GS, molecular studies have revealed the presence of a number of distinct isoforms in several plant species (12,14,16,20). In pea it has been shown that two classes of genes encode homologous but distinct cytosolic GS isoforms (13).…”

“…Early biochemical data revealed that GS functions in the assimilation of ammonia generated by numerous plant processes, which include seed germination (4,5), photorespiration (6,7), nitrite reduction (8), nitrogen-fixation in root nodules (9,10), and primary ammonia assimilation from the soil (11). An analysis of the GS genes in several species has revealed a strong correlation of individual GS gene expression with specific aspects of plant development (12)(13)(14)(15)(16)(17)(18). Recent sequence analysis of GS cDNAs from Pisum sativum and Phaseolus vulgaris has shown that chloroplast and cytosolic GS are encoded by separate but similar nuclear genes (12,13,19).…”

Subunit Composition of Glutamine Synthetase Isozymes from Root Nodules of Bean (Phaseolus vulgaris L.)

“…A small source is in the infected cells, and a large source is in the uninfected cells, which for the purpose of this discussion are all nodule cells except the infected ones. The (3 polypeptide is found in the leaves, roots, and nodules (7,13). Its gene appears to be constitutively expressed (7).…”