Glutamine synthetase genes of pea encode distinct polypeptides which are differentially expressed in leaves, roots and nodules.

Abstract

We have characterized the distinct polypeptides, primary translation products and mRNAs encoding glutamine synthetase (GS) in the various organs of pea. Western blot analysis of soluble protein has identified five distinct GS polypeptides which are expressed at different relative levels in leaves, roots and nodules of pea. Of the two GS polypeptides in leaves (44 and 38 kd), the 44‐kd GS polypeptide is predominant and is localized to the chloroplast stroma. In roots, the predominant GS polypeptide is 38 kd. Upon Rhizobium infection of roots, three 37‐kd GS polypeptides increase in abundance in the nodules relative to uninfected roots. cDNA clones encoding three different GS mRNAs have been characterized. Hybrid‐select translation has identified three different GS primary translation products (49, 38 and 37 kd). Two cDNA clones (pGS134 and pGS341) are homologous to GS mRNAs most abundant in nodules which encode the 38‐ and 37‐kd GS primary translation products. A third cDNA (pGS197) corresponds to a larger GS mRNA species specific to leaf poly(A) RNA, which encodes a 49‐kd putative precursor to the mature chloroplast GS polypeptide. cDNA sequence analysis and Southern blot analysis of pea nuclear DNA identifies at least three genes encoding GS in pea which are related but distinct in structure and in vivo pattern of expression.

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

“…"Sparkle" (Rogers Brothers Seed, Twin Falls, ID) constructed in "Lambda Dash" (Stratagene). Complete sequence analysis of each genomic clone (unpublished data) revealed that the genomic clone for chloroplast GS2 (GS2ct) corresponds to the GS185 cDNA (13), whereas the genomic clone for cytosolic GS (GS3ACY) corresponds to the GS341 cDNA (12,18).…”

“…1B). pGS3AcY-GUS contains 88 nucleotides of the 5' untranslated leader of GS3A mRNA and encodes 6 amino acids of the cytosolic GS protein (12).…”

“…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).In pea, the single nuclear gene for chloroplast GS2 is expressed predominantly in leaves in a light-dependent fashion (13,17). 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).…”

“…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).…”