Regulation of nitrogen assimilation in Saccharomyces cerevisiae: roles of the URE2 and GLN3 genes

Abstract

Mutations in the GLN3 gene prevented a normal increase in the NAD-glutamate dehydrogenase and glutamine synthetase levels in glutamate-grown Saccharomyces cerevisiae cells, whereas mutations in the URE2 gene resulted in high levels of these enzymes in glutamate-and glutamine-grown cells. A ure2 gln3 double mutant had low levels of glutamate dehydrogenase and glutamine synthetase in cells grown on glutamate and glutamine; thus, gln3 mutations were epistatic to the ure2 mutations. The results suggest that the GLN3 product is capable of promoting increases in enzyme levels in the absence of a functional URE2 product and that the URE2 product antagonizes the GLN3 product. The URE2 and GLN3 genes were also found to regulate the level of arginase activity. This regulation is completely independent of the regulation of arginase by substrate induction. The activities of glutamate dehydrogenase, glutamine synthetase, and arginase were higher in cells grown on glutamate as the nitrogen source than they were in cells grown under a nitrogen-limiting condition.It had previously been shown that the levels of these enzymes can be increased by glutamine deprivation. We propose that the URE2-GLN3 system regulates enzyme synthesis, in response to glutamine and glutamate, to adjust the intracellular concentration of ammonia so as to maintain glutamine at the level required for optimal growth.Glutamate and glutamine play the role of amino donors in the flow of nitrogen into all organic compounds. In Saccharomyces cerevisiae there are three enzymes of central importance that are involved in the metabolism of glutamate and glutamine: NADPH-glutamate dehydrogenase (NADP-GDH), NAD-GDH, and glutamine synthetase. NADP-GDH functions in the biosynthesis of glutamate from ammonia, while NAD-GDH degrades glutamate to ammonia and glutamine synthetase catalyzes the synthesis of glutamine from glutamate and ammonia. The activities of NAD-GDH and glutamine synthetase vary depending on what nitrogen source is present in the growth medium (11,14,15,20). Their activities are high when glutamate is the nitrogen source and low when glutamine is the source. The activities of many catabolic enzymes also vary depending on the nitrogen source. The activities of these enzymes are also increased in response to the presence of their substrate (reviewed in references 4 and 25). These catabolic enzymes catalyze the degradation of nitrogen compounds. The final product of degradation is either ammonia or glutamate, which can give rise to ammonia by the activity of NAD-GDH. The activities of some of these enzymes are also increased when glutamate rather than glutamine or ammonia is the source of nitrogen. We examined the relationship of the regulation of glutamine synthetase with that of NAD-GDH and of arginase, a catabolic enzyme subject to nitrogen regulation.Mutants lacking glutamine synthetase activity were first isolated by Dubois and Grenson (11), and the structural gene encoding glutamine synthetase was shown to be at the GLNI locus by Mitchell and Magasa...

Gzf3p, a fourth GATA factor involved in nitrogen‐regulated transcription in Saccharomyces cerevisiae

“…Gln3p-activated expression is sensitive to nitrogen repression (Mitchell and Magasanik, 1984;Magasanik, 1992). Mutations in the URE2IGDHCR gene (Grenson, 1969;Drillien and Lacroute, 1972;Grenson et a/., 1974) at least partially relieve nitrogen repression of Gln3p (Courchesne and Magasanik, 1988). The Ure2/GdhCR protein is somewhat similar to glutathione transferases (Coschigano and Magasanik, 1991) and has the peculiar ability to generate prion analogues (Wickner, 1994).…”

The crystal structure of the nitrogen regulation fragment of the yeast prion protein Ure2p

“…Ure2p directly interacts with the DNA binding protein Gln3p, preventing its entry into the nucleus to promote transcription of an array of genes involved in assimilation of poor nitrogen sources (12)(13)(14)(15). Ure2p is itself regulated by Mks1p, mediating nitrogen catabolite repression (16), and by the TOR system, a global regulator that transmits nutritional information to many cellular components (14,17,18).…”

RTG-dependent Mitochondria-to-Nucleus Signaling Is Regulated by MKS1 and Is Linked to Formation of Yeast Prion [URE3]

“…A connection between the retrograde pathway and nitrogen metabolism has recently been uncovered by the finding that the target of rapamycin (TOR) kinase pathway in yeast also involves the RTG genes (Komeili et al, 2000;Shamji et al, 2000). The TOR kinase pathway promotes the formation of a complex between Gln3p, a transcription factor that controls the expression of genes required for the utilization of poor nitrogen sources (Mitchell and Magasanik, 1984;Courchesne and Magasanik, 1988;Coschigano and Magasanik, 1991;Blinder et al, 1996), and Ure2p, a negative regulator of Gln3p (Beck and Hall, 1999). When the TOR kinase pathway is inhibited by rapamycin, or when cells are grown on a poor nitrogen source such as urea, Rtg1p and Rtg3p translocate from the cytoplasm to the nucleus in an Rtg2p-dependent manner to activate target gene expression (Komeili et al, 2000).…”