Metabolic Regulation of the Gene Encoding Glutamine-Dependent Asparagine Synthetase in Arabidopsis thaliana

Abstract

Here, we characterize a cDNA encoding a glutamine-dependent asparagine synthetase (ASN7) from Arabidopsis thaliana and assess the effects of metabolic regulation on ASNl mRNA levels. Sequence analysis shows that the predicted ASNl peptide contains a purFtype glutamine-binding domain. Southern blot experiments and cDNA clone analysis suggest that ASNl is the only gene encoding glutamine-dependent asparagine synthetase in A. thaliana. The ASNl gene is expressed predominantly in shoot tissues, where light has a negative effect on its mRNA accumulation. This negative effect of light on ASNl mRNA levels was shown to be mediated, at least in part, via the photoreceptor phytochrome. We also investigated whether light-induced changes in nitrogen to carbon ratios might exert a metabolic regulation of the ASNl mRNA accumulation. These experiments demonstrated that the accumulation of ASNl mRNA in dark-grown plants is strongly repressed by the presence of exogenous sucrose. Moreover, this sucrose repression of ASNl expression can be partially rescued by supplementation with exogenous amino acids such as asparagine, glutamine, and glutamate. These findings suggest that the expression of the ASNl gene is under the metabolic control of the nitrogen to carbon ratio in cells. This is consistent with the fact that asparagine, synthesized by the ASNl gene product, is a favored compound for nitrogen storage and nitrogen transport in dark-grown plants. We have put forth a working model suggesting that when nitrogen to carbon ratios are high, the gene product of ASNl functions to re-direct the flow of nitrogen into asparagine, which acts as a shunt for storage and/or long-distance transport of nitrogen.

Hexokinase as a sugar sensor in higher plants.

“…In higher plants, sugars affect growth and development throughout the life cycle, from germination to flowering to senescence (Steeves and Sussex, 1989;Brusslan and Tobin, 1992;Graham et al, 1992;Bernier et al, 1993;Sheen, 1994;Thomas and Rodriguez, 1994;Dangl et al, 1995). Recently, it has become apparent that sugars are physiological signals repressing or activating plant genes involved in many essential processes, including photosynthesis, glyoxylate metabolism, respiration, starch and sucrose synthesis and degradation, nitrogen metabolism, pathogen defense, wounding response, cell cycle regulation, pigmentation, and senescence (Chen et al, 1994;Knight and Gray, 1994;Lam et al, 1994;Sheen, 1994;Herbers et al, 1995;Mita et al, 1995;Reynolds and Smith, 1995).…”

Characterization and Expression of NAD(H)-Dependent Glutamate Dehydrogenase Genes in Arabidopsis

“…1995) findings in maize suggest that NAD(H)-GDH isoenzymes are encoded by more than one gene. Furthermore, the presence of multiple genes encoding for enzymes involved in nitrogen metabolism appears to be a common occurrence in A. thaliana, and small multigene families encoding for GS (Peterman and Goodman, 1991), GOGAT (Lam et al, 1995a), Asn synthetase (Lam et al, 1995a(Lam et al, , 1995b, and aspartate aminotransferase (Schultz and Coruzzi, 1995;Wilkie et al, 1995) have been reported. Our findings confirm the existence of a small gene family encoding NAD(H)-GDH.…”

Expression of an Arabidopsis Aspartate Kinase/Homoserine Dehydrogenase Gene Is Metabolically Regulated by Photosynthesis-Related Signals but Not by Nitrogenous Compounds1

“…This results in efficient biosynthesis of Gln, which is further transaminated into aspartate. However, these metabolic signals also repress the expression of the gene encoding Asn synthetase, blocking further conversion of aspartate into Asn Coruzzi, 1990, 1991;Lam et al, 1994Lam et al, , 1995Chevalier et al, 1996) In contrast, at night, when the carbon-to-nitrogen ratio is relatively low, dark and relatively high nitrogen levels stimulate the expression of Asn synthetase, resulting in efficient conversion of aspartate into Asn (Lam et al, 1995). Our previous (Zhu- Shimoni et al, 1997) and present reports showed further that the conversion of aspartate into Asn or into aspartate family amino acids is also subject to a coordinated, reciprocal metabolic regulation of the expression of AK/HSD and Asn synthetase genes.…”

“…The metabolism of Gln, glutamate, aspartate, and Asn is subject to a coordinated metabolic regulation. Whereas the expression of an Arabidopsis and pea Gln synthetase was shown to be stimulated by light (Lam et al, 1994(Lam et al, , 1995, the expression of an Arabidopsis Asn synthetase was repressed by light and Suc, and stimulated by dark and nitrogen Coruzzi, 1990, 1991;Lam et al, 1995;Chevalier et al, 1996). It has been suggested that during the day plants accumulate Gln, glutamate, and aspartate, which are apparently used for the synthesis of other amino acids, whereas during the night, aspartate is converted into Asn for storage (Galili, 1995;Lam et al, 1995).…”