Chloroplast and cytosolic glutamine synthetase are encoded by homologous nuclear genes which are differentially expressed in vivo.

S V Tingey,F Y Tsai,J W Edwards,E L Walker,G M Coruzzi

doi:10.1016/s0021-9258(19)81566-1

Abstract

We have shown that the individual members of the plant gene family for glutamine synthetase (GS) are differentially expressed in vivo, and each encode distinct GS polypeptides which are targeted to different subcellular compartments (chloroplast or cytosol). At the polypeptide level, chloroplast GS (GS2) and cytosolic GS (GS1 and GSn) are distinct and show an organ-specific distribution. We have characterized full length cDNA clones encoding chloroplast or cytosolic GS of pea. In vitro translation products encoded by three different GS cDNA clones, correspond to the mature GS2, GS1, and GSn polypeptides present in vivo. pGS185 encodes a precursor to the chloroplast GS2 polypeptide as shown by in vitro chloroplast uptake experiments. The pGS185 translation product is imported into the chloroplast stroma and processed to a polypeptide which corresponds in size and charge to that of mature chloroplast stromal GS2 (44 kDa). The 49 amino terminal amino acids encoded by pGS185 are designated as a chloroplast transit peptide by functionality in vitro, and amino acid homology to other transit peptides. The cytosolic forms of GS (GS1 and GSn) are encoded by highly homologous but distinct mRNAs. pGS299 encodes the cytosolic GS1 polypeptide (38 kDa), while pGS341 (Tingey, S. V., Walker, E. L., and Coruzzi, G. M. (1987) EMBO. J. 6, 1-9) encodes a cytosolic GSn polypeptide (37 kDa). The homologous nuclear genes for chloroplast and cytosolic GS show different patterns of expression in vivo. GS2 expression in leaves is modulated by light, at the level of steady state mRNA and protein, while the expression of cytosolic GS is unaffected by light. The light-induced expression of GS2 is due at least in part to a phytochrome mediated response. Nucleotide sequence analysis indicates that chloroplast and cytosolic GS have evolved from a common ancestor and suggest a molecular mechanism for chloroplast evolution.

Highlights

We have shown that the individual members of the evolved from a common ancestor andsuggest a molecplant gene family for glutamine synthetase (GS) are ular mechanism for chloroplast evolution
Chloroplast GS (GSz) and cyto- Higher plant glutamine synthetase (GS,‘ EC 6.3.1.2), the solic GS (GS1and GS.) are distinct andshow an organ- major enzyme which catalyzes the assimilation of ammonia, specific distribution
In roots, where GS functions to assimilate the polypeptide which corresponds in size and charge to ammonia fixed by bacteroids, the levels of certain cytosolic that of mature chloroplast stromal GSz (44 kDa)

Summary

EXPERIMENTAL PROCEDURES

Ll Recipient of National Science Foundation Postdoctoral Fellow- were imbibed and germinated in a Conviron environmental chamber ship in Plant Biology DMB8710630. With a day length of 16 h, illumination of 1000 microeinsteins.m-*. 11 National Institutes of Health Predoctoral Trainee, supported by s-l (1 einstein = 1mol of photons), at a day/night cycle of 21/18 “C. ** To whom correspondence should be addressed Laboratory of ‘The abbreviations used are: GS, glutamine synthetase;SDS-. Plant Molecular Biology,The RockefellerUniversity, 1230 York Ave., PAGE, sodium dodecyl sulfate-polyacrylamide gel electrophoresis; New York, NY 10021-6399.

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DISCUSSION