Repression of GCN4 mRNA Translation by Nitrogen Starvation in Saccharomyces cerevisiae

Saccharomyces cerevisiae activates a regulatory network called “general control” that provides the cell with sufficient amounts of protein precursors during amino acid starvation. We investigated how starvation for nitrogen affects the general control regulatory system, because amino acid biosynthesis is part of nitrogen metabolism. Amino acid limitation results in the synthesis of the central transcription factor Gcn4p, which binds to specific DNA-binding motif sequences calledGcn4-protein-responsive elements (GCREs) that are present in the promoter regions of its target genes. Nitrogen starvation increases GCN4 transcription but efficiently represses expression of both a syntheticGCRE6::lacZ reporter gene and the natural amino acid biosynthetic gene ARO4. Repression of Gcn4p-regulated transcription by nitrogen starvation is independent of the ammonium sensing systems that include Mep2p and Gpa2p or Ure2p and Gln3p but depends on the four upstream open reading frames in theGCN4 mRNA leader sequence. Efficient translation ofGCN4 mRNA is completely blocked by nitrogen starvation, even when cells are simultaneously starved for amino acids and eukaryotic initiation factor-2α is fully phosphorylated by Gcn2p. Our data suggest that nitrogen starvation regulates translation ofGCN4 by a novel mechanism that involves the four upstream open reading frames but that still acts independently of eukaryotic initiation factor-2α phosphorylation by Gcn2p. Saccharomyces cerevisiae activates a regulatory network called “general control” that provides the cell with sufficient amounts of protein precursors during amino acid starvation. We investigated how starvation for nitrogen affects the general control regulatory system, because amino acid biosynthesis is part of nitrogen metabolism. Amino acid limitation results in the synthesis of the central transcription factor Gcn4p, which binds to specific DNA-binding motif sequences calledGcn4-protein-responsive elements (GCREs) that are present in the promoter regions of its target genes. Nitrogen starvation increases GCN4 transcription but efficiently represses expression of both a syntheticGCRE6::lacZ reporter gene and the natural amino acid biosynthetic gene ARO4. Repression of Gcn4p-regulated transcription by nitrogen starvation is independent of the ammonium sensing systems that include Mep2p and Gpa2p or Ure2p and Gln3p but depends on the four upstream open reading frames in theGCN4 mRNA leader sequence. Efficient translation ofGCN4 mRNA is completely blocked by nitrogen starvation, even when cells are simultaneously starved for amino acids and eukaryotic initiation factor-2α is fully phosphorylated by Gcn2p. Our data suggest that nitrogen starvation regulates translation ofGCN4 by a novel mechanism that involves the four upstream open reading frames but that still acts independently of eukaryotic initiation factor-2α phosphorylation by Gcn2p. Gcn4-protein-responsive elements upstream open reading frames eukaryotic initiation factor 2 3-aminotriazole In bakers' yeast, Saccharomyces cerevisiae, starvation for a single amino acid induces the transcription of more than 50 genes encoding enzymes involved in several amino acid biosynthetic pathways, amino acid tRNA synthetases (1Meussdoerffer F. Fink G.R. J. Biol. Chem. 1983; 258: 6293-6299Abstract Full Text PDF PubMed Google Scholar, 2Mirande M. Waller J.P. J. Biol. Chem. 1988; 263: 18443-18451Abstract Full Text PDF PubMed Google Scholar), or enzymes of purine biosynthesis (3Mösch H.-U. Scheier B. Lahti R. Mantsala P. Braus G.H. J. Biol. Chem. 1991; 266: 20453-20460Abstract Full Text PDF PubMed Google Scholar). This genetic system is called “general control of amino acid biosynthesis” (4Hinnebusch A.G. Jones E.W. Pringle J.R. Broach J.R. General and Pathway-specific Regulatory Mechanisms Controlling the Synthesis of Amino Acid Biosynthetic Enzymes in Saccharomyces cerevisiae: The Molecular and Cellular Biology of the Yeast Saccharomyces. Cold Spring Harbor Laboratory, Cold Spring Harbor, New York1992: 319-414Google Scholar). Amino acid limitation results in the synthesis of the transcription factor Gcn4p (5Ellenberger T.E. Brandl C.J. Struhl K. Harrison S.C. Cell. 1992; 71: 1223-1237Abstract Full Text PDF PubMed Scopus (813) Google Scholar), which binds to specificGcn4-protein-responsive elements (GCREs)1 present in the promoter regions of its target genes. Gcn4p stimulates transcription of its target genes by a factor of 2–10 (6Hope I.A. Struhl K. Cell. 1986; 46: 885-894Abstract Full Text PDF PubMed Scopus (547) Google Scholar). Whereas some target genes are regulated via several GCREs in their promoter regions likeHIS3 or TRP4, for example (7Braus G. Mösch H.-U. Vogel K. Hinnen A. Hutter R. EMBO J. 1989; 8: 939-945Crossref PubMed Scopus (46) Google Scholar, 8Hill D.E. Hope I.A. Macke J.P. Struhl K. Science. 1986; 234: 451-457Crossref PubMed Scopus (211) Google Scholar, 9Hope I.A. Struhl K. Cell. 1985; 43: 177-188Abstract Full Text PDF PubMed Scopus (265) Google Scholar, 10Mösch H.-U. Graf R. Schmidheini T. Braus G. EMBO J. 1990; 9: 2951-2957Crossref PubMed Scopus (17) Google Scholar), others contain only a single GCRE site that is both essential and sufficient for regulation by Gcn4p as found for the ARO4 gene (11Künzler M. Paravicini G. Egli C.M. Irniger S. Braus G.H. Gene ( Amst. ). 1992; 113: 67-74Crossref PubMed Scopus (43) Google Scholar).The best understood regulatory mechanism of Gcn4p expression is translational control of its mRNA, which requires four smallupstream open readingframes (uORFs) present in the GCN45′-untranslated region (12Hinnebusch A.G. Proc. Natl. Acad. Sci. U. S. A. 1984; 81: 6442-6446Crossref PubMed Scopus (276) Google Scholar, 13Thireos G. Penn M.D. Greer H. Proc. Natl. Acad. Sci. U. S. A. 1984; 81: 5096-5100Crossref PubMed Scopus (109) Google Scholar). When cells are growing under non-starvation conditions, ribosomes translate the first uORF, reinitiate at uORFs 2–4, and are unable to recognize theGCN4 start codon (14Hinnebusch A.G. J. Biol. Chem. 1997; 272: 21661-21664Abstract Full Text Full Text PDF PubMed Scopus (431) Google Scholar). In cells starved for amino acids, ribosomes that have translated the first uORF reinitiate at theGCN4 translational start site leading to high expression of Gcn4p. Inactivation of the four uORFs in the GCN4 leader by either deletion or mutation of the four ATG start codons uncouples translational control of GCN4 from the general control system, leading to high expression of Gcn4p already under non-starvation conditions (15Mueller P.P. Hinnebusch A.G. Cell. 1986; 45: 201-207Abstract Full Text PDF PubMed Scopus (355) Google Scholar). Translational regulation ofGCN4 depends on the sensor kinase Gcn2p, which includes an N-terminal protein kinase domain and a C-terminal aminoacyl tRNA synthetase-like domain (16Roussou I. Thireos G. Hauge B.M. Mol. Cell. Biol. 1988; 8: 2132-2139Crossref PubMed Scopus (59) Google Scholar, 17Wek R.C. Jackson B.M. Hinnebusch A.G. Proc. Natl. Acad. Sci. U. S. A. 1989; 86: 4579-4583Crossref PubMed Scopus (202) Google Scholar). Under amino acid starvation conditions, Gcn2p detects uncharged tRNAs and in response phosphorylates the α-subunit of eukaryotic initiation factor 2 (eIF-2). Phosphorylated eIF-2 inhibits translation of most mRNAs mediated by the inhibition of the guanine nucleotide exchange activity of eIF-2B. This allows ribosomes to scan past the remaining uORFs in the GCN4 5′-untranslated region and translate the mRNA of the GCN4 gene (18Dever T.E. Feng L. Wek R.C. Cigan A.M. Donahue T.F. Hinnebusch A.G. Cell. 1992; 68: 585-596Abstract Full Text PDF PubMed Scopus (560) Google Scholar, 19Wek S.A. Zhu S. Wek R.C. Mol. Cell. Biol. 1995; 15: 4497-4506Crossref PubMed Google Scholar).Amino acid biosynthesis is part of the general nitrogen metabolism of yeast cells. Ammonium is among the inorganic nitrogen sources in nature that support optimal growth of yeast cells (20Wiame J.M. Grenson M. Arst Jr., H.N. Adv. Microb. Physiol. 1985; 26: 1-88Crossref PubMed Scopus (237) Google Scholar). Three ammonium permeases, Mep1p, Mep2p, and Mep3p, are known that control uptake of ammonium into the cytoplasm (21Marini A.M. Soussi-Boudekou S. Vissers S. Andre B. Mol. Cell. Biol. 1997; 17: 4282-4293Crossref PubMed Scopus (444) Google Scholar). Deletion of all three MEPgenes renders yeast cells inviable on media containing less than 5 mm ammonium sulfate as the sole nitrogen source. In addition to its functions as ammonium permease, Mep2p is thought to act as an ammonium sensor protein in a signaling system that controls pseudohyphal development of diploid Σ1278bstrains, because diploid mep2Δ mutant strains are unable to form pseudohyphae in response to nitrogen starvation. Pseudohyphal development of mep2Δ mutants can be restored by expression of dominant activated forms of Gpa2p or Ras2p, GTP-binding proteins that both regulate intracellular cAMP levels (22Kübler E. Mösch H.-U. Rupp S. Lisanti M.P. J. Biol. Chem. 1997; 272: 20321-20323Abstract Full Text Full Text PDF PubMed Scopus (166) Google Scholar, 23Lorenz M.C. Heitman J. EMBO J. 1997; 16: 7008-7018Crossref PubMed Google Scholar). These studies have led to the model that Mep2p is a central ammonium sensor protein that activates Gpa2p and cAMP-dependent protein kinase in response to nitrogen starvation. The proteins Ure2p and Gln3p are two additional important regulators in the ammonium utilization pathway and are required under all conditions (24ter Schure E.G. van Riel N.A. Verrips C.T. FEMS Microbiol. Rev. 2000; 24: 67-83Crossref PubMed Scopus (205) Google Scholar).As amino acid biosynthesis is part of the general nitrogen metabolism in yeast, we investigated how starvation for nitrogen affects the general control system of amino acids. We found that deprivation of a suitable nitrogen source efficiently counteracts the activation of Gcn4p-controlled gene expression. Expression of a syntheticGCRE6::lacZ reporter gene and natural amino acid biosynthetic genes such as ARO4 is suppressed when cells are starved for starvation for only on Gcn4p-controlled gene expression. We the mRNA ofGCN4 that nitrogen starvation translation of The four uORFs in the GCN4 mRNA leader are for the the ammonium sensing system that includes Mep2p or and the proteins Ure2p or Gln3p are cells to their metabolism for optimal growth and Nitrogen starvation requires the expression of genes for enzymes to cells by to a growth In we investigated how starvation for nitrogen affects the “general control” regulatory network that induces amino acid biosynthetic gene expression when yeast cells are starved for amino acids. amino acid biosynthesis requires at network is an important part of the general nitrogen metabolism. Yeast cells by into and (24ter Schure E.G. van Riel N.A. Verrips C.T. FEMS Microbiol. Rev. 2000; 24: 67-83Crossref PubMed Scopus (205) Google Scholar). This to the general of includes the more specific of amino and activates the general control system and its Gcn4p. In we found the that a general of nitrogen source and efficiently inhibits gene of that inhibition of general gene expression by nitrogen starvation at the of GCN4 mRNA We that protein levels in the of levels of Gcn4p levels or expression can only when translated from GCN4 mRNA the uORFs in the leader sequence. Nitrogen starvation completely amino acid expression and Gcn4p on the of the phosphorylation of by Gcn2p is blocked by nitrogen starvation. the we that nitrogen starvation to an even in phosphorylation than amino acid starvation. starvation for nitrogen completely translation of GCN4 mRNA, even under amino acid starvation This that nitrogen starvation GCN4 mRNA translation by an additional nitrogen starvation Gcn2p or protein levels the of a of the general control system that acts of be in or expression. This an additional translational regulation mechanism the that nitrogen starvation, from translation of GCN4 mRNA, the protein on a This is on two Whereas in the of expression is by nitrogen starvation, expression of reporter is still to a When uORFs are in theGCN4 expression translation is by nitrogen starvation, but both protein levels and expression of reporter are be that nitrogen starvation protein Whereas amino acid starvation inhibits Gcn4p by the system B. Fink G.R. EMBO J. PubMed Scopus Google Scholar), nitrogen starvation have and of nitrogen sensing and signaling system to translational of two are known that are regulated by the of pseudohyphal and signaling system that of Mep2p, and the cAMP-dependent protein kinase is thought to regulate pseudohyphal development when cells are starved for nitrogen (22Kübler E. Mösch H.-U. Rupp S. Lisanti M.P. J. Biol. Chem. 1997; 272: 20321-20323Abstract Full Text Full Text PDF PubMed Scopus (166) Google Scholar, M.C. Heitman J. Google Scholar). Mep2p Gpa2p to be involved in translational control of In the of yeast the utilization of nitrogen such as and This is as nitrogen J. Mol. PubMed Scopus Google Scholar). The genetic system involves the of the transcription factor Gln3p that is regulated by the protein Ure2p in the of strains which that proteins are involved in GCN4 under nitrogen starvation. The of the intracellular ammonium by the ammonium led to a of Gcn4p This the of an additional sensor system for the intracellular ammonium which is to a for of theGCN4 mRNA studies that yeast cells amino acid biosynthesis only when precursors are Nitrogen starvation represses activity of the general control system but the expression of amino acid biosynthetic genes. This that amino acid biosynthesis is completely even under starvation The of amino acid biosynthesis under conditions is by the regulatory mechanism that we found in yeast cells of amino acid biosynthesis when nitrogen sources are mRNA levels for the Gcn4p are at a in to translation of when nitrogen In bakers' yeast, Saccharomyces cerevisiae, starvation for a single amino acid induces the transcription of more than 50 genes encoding enzymes involved in several amino acid biosynthetic pathways, amino acid tRNA synthetases (1Meussdoerffer F. Fink G.R. J. Biol. Chem. 1983; 258: 6293-6299Abstract Full Text PDF PubMed Google Scholar, 2Mirande M. Waller J.P. J. Biol. Chem. 1988; 263: 18443-18451Abstract Full Text PDF PubMed Google Scholar), or enzymes of purine biosynthesis (3Mösch H.-U. Scheier B. Lahti R. Mantsala P. Braus G.H. J. Biol. Chem. 1991; 266: 20453-20460Abstract Full Text PDF PubMed Google Scholar). This genetic system is called “general control of amino acid biosynthesis” (4Hinnebusch A.G. Jones E.W. Pringle J.R. Broach J.R. General and Pathway-specific Regulatory Mechanisms Controlling the Synthesis of Amino Acid Biosynthetic Enzymes in Saccharomyces cerevisiae: The Molecular and Cellular Biology of the Yeast Saccharomyces. Cold Spring Harbor Laboratory, Cold Spring Harbor, New York1992: 319-414Google Scholar). Amino acid limitation results in the synthesis of the transcription factor Gcn4p (5Ellenberger T.E. Brandl C.J. Struhl K. Harrison S.C. Cell. 1992; 71: 1223-1237Abstract Full Text PDF PubMed Scopus (813) Google Scholar), which binds to specificGcn4-protein-responsive elements (GCREs)1 present in the promoter regions of its target genes. Gcn4p stimulates transcription of its target genes by a factor of 2–10 (6Hope I.A. Struhl K. Cell. 1986; 46: 885-894Abstract Full Text PDF PubMed Scopus (547) Google Scholar). Whereas some target genes are regulated via several GCREs in their promoter regions likeHIS3 or TRP4, for example (7Braus G. Mösch H.-U. Vogel K. Hinnen A. Hutter R. EMBO J. 1989; 8: 939-945Crossref PubMed Scopus (46) Google Scholar, 8Hill D.E. Hope I.A. Macke J.P. Struhl K. Science. 1986; 234: 451-457Crossref PubMed Scopus (211) Google Scholar, 9Hope I.A. Struhl K. Cell. 1985; 43: 177-188Abstract Full Text PDF PubMed Scopus (265) Google Scholar, 10Mösch H.-U. Graf R. Schmidheini T. Braus G. EMBO J. 1990; 9: 2951-2957Crossref PubMed Scopus (17) Google Scholar), others contain only a single GCRE site that is both essential and sufficient for regulation by Gcn4p as found for the ARO4 gene (11Künzler M. Paravicini G. Egli C.M. Irniger S. Braus G.H. Gene ( Amst. ). 1992; 113: 67-74Crossref PubMed Scopus (43) Google Scholar). The best understood regulatory mechanism of Gcn4p expression is translational control of its mRNA, which requires four smallupstream open readingframes (uORFs) present in the GCN45′-untranslated region (12Hinnebusch A.G. Proc. Natl. Acad. Sci. U. S. A. 1984; 81: 6442-6446Crossref PubMed Scopus (276) Google Scholar, 13Thireos G. Penn M.D. Greer H. Proc. Natl. Acad. Sci. U. S. A. 1984; 81: 5096-5100Crossref PubMed Scopus (109) Google Scholar). When cells are growing under non-starvation conditions, ribosomes translate the first uORF, reinitiate at uORFs 2–4, and are unable to recognize theGCN4 start codon (14Hinnebusch A.G. J. Biol. Chem. 1997; 272: 21661-21664Abstract Full Text Full Text PDF PubMed Scopus (431) Google Scholar). In cells starved for amino acids, ribosomes that have translated the first uORF reinitiate at theGCN4 translational start site leading to high expression of Gcn4p. Inactivation of the four uORFs in the GCN4 leader by either deletion or mutation of the four ATG start codons uncouples translational control of GCN4 from the general control system, leading to high expression of Gcn4p already under non-starvation conditions (15Mueller P.P. Hinnebusch A.G. Cell. 1986; 45: 201-207Abstract Full Text PDF PubMed Scopus (355) Google Scholar). Translational regulation ofGCN4 depends on the sensor kinase Gcn2p, which includes an N-terminal protein kinase domain and a C-terminal aminoacyl tRNA synthetase-like domain (16Roussou I. Thireos G. Hauge B.M. Mol. Cell. Biol. 1988; 8: 2132-2139Crossref PubMed Scopus (59) Google Scholar, 17Wek R.C. Jackson B.M. Hinnebusch A.G. Proc. Natl. Acad. Sci. U. S. A. 1989; 86: 4579-4583Crossref PubMed Scopus (202) Google Scholar). Under amino acid starvation conditions, Gcn2p detects uncharged tRNAs and in response phosphorylates the α-subunit of eukaryotic initiation factor 2 (eIF-2). Phosphorylated eIF-2 inhibits translation of most mRNAs mediated by the inhibition of the guanine nucleotide exchange activity of eIF-2B. This allows ribosomes to scan past the remaining uORFs in the GCN4 5′-untranslated region and translate the mRNA of the GCN4 gene (18Dever T.E. Feng L. Wek R.C. Cigan A.M. Donahue T.F. Hinnebusch A.G. Cell. 1992; 68: 585-596Abstract Full Text PDF PubMed Scopus (560) Google Scholar, 19Wek S.A. Zhu S. Wek R.C. Mol. Cell. Biol. 1995; 15: 4497-4506Crossref PubMed Google Scholar). Amino acid biosynthesis is part of the general nitrogen metabolism of yeast cells. Ammonium is among the inorganic nitrogen sources in nature that support optimal growth of yeast cells (20Wiame J.M. Grenson M. Arst Jr., H.N. Adv. Microb. Physiol. 1985; 26: 1-88Crossref PubMed Scopus (237) Google Scholar). Three ammonium permeases, Mep1p, Mep2p, and Mep3p, are known that control uptake of ammonium into the cytoplasm (21Marini A.M. Soussi-Boudekou S. Vissers S. Andre B. Mol. Cell. Biol. 1997; 17: 4282-4293Crossref PubMed Scopus (444) Google Scholar). Deletion of all three MEPgenes renders yeast cells inviable on media containing less than 5 mm ammonium sulfate as the sole nitrogen source. In addition to its functions as ammonium permease, Mep2p is thought to act as an ammonium sensor protein in a signaling system that controls pseudohyphal development of diploid Σ1278bstrains, because diploid mep2Δ mutant strains are unable to form pseudohyphae in response to nitrogen starvation. Pseudohyphal development of mep2Δ mutants can be restored by expression of dominant activated forms of Gpa2p or Ras2p, GTP-binding proteins that both regulate intracellular cAMP levels (22Kübler E. Mösch H.-U. Rupp S. Lisanti M.P. J. Biol. Chem. 1997; 272: 20321-20323Abstract Full Text Full Text PDF PubMed Scopus (166) Google Scholar, 23Lorenz M.C. Heitman J. EMBO J. 1997; 16: 7008-7018Crossref PubMed Google Scholar). These studies have led to the model that Mep2p is a central ammonium sensor protein that activates Gpa2p and cAMP-dependent protein kinase in response to nitrogen starvation. The proteins Ure2p and Gln3p are two additional important regulators in the ammonium utilization pathway and are required under all conditions (24ter Schure E.G. van Riel N.A. Verrips C.T. FEMS Microbiol. Rev. 2000; 24: 67-83Crossref PubMed Scopus (205) Google Scholar). amino acid biosynthesis is part of the general nitrogen metabolism in yeast, we investigated how starvation for nitrogen affects the general control system of amino acids. We found that deprivation of a suitable nitrogen source efficiently counteracts the activation of Gcn4p-controlled gene expression. Expression of a syntheticGCRE6::lacZ reporter gene and natural amino acid biosynthetic genes such as ARO4 is suppressed when cells are starved for starvation for only on Gcn4p-controlled gene expression. We the mRNA ofGCN4 that nitrogen starvation translation of The four uORFs in the GCN4 mRNA leader are for the the ammonium sensing system that includes Mep2p or and the proteins Ure2p or Gln3p are cells to their metabolism for optimal growth and Nitrogen starvation requires the expression of genes for enzymes to cells by to a growth In we investigated how starvation for nitrogen affects the “general control” regulatory network that induces amino acid biosynthetic gene expression when yeast cells are starved for amino acids. amino acid biosynthesis requires at network is an important part of the general nitrogen metabolism. Yeast cells by into and (24ter Schure E.G. van Riel N.A. Verrips C.T. FEMS Microbiol. Rev. 2000; 24: 67-83Crossref PubMed Scopus (205) Google Scholar). This to the general of includes the more specific of amino and activates the general control system and its Gcn4p. In we found the that a general of nitrogen source and efficiently inhibits gene of that inhibition of general gene expression by nitrogen starvation at the of GCN4 mRNA We that protein levels in the of levels of Gcn4p levels or expression can only when translated from GCN4 mRNA the uORFs in the leader sequence. Nitrogen starvation completely amino acid expression and Gcn4p on the of the phosphorylation of by Gcn2p is blocked by nitrogen starvation. the we that nitrogen starvation to an even in phosphorylation than amino acid starvation. starvation for nitrogen completely translation of GCN4 mRNA, even under amino acid starvation This that nitrogen starvation GCN4 mRNA translation by an additional nitrogen starvation Gcn2p or protein levels the of a of the general control system that acts of be in or expression. This an additional translational regulation mechanism the that nitrogen starvation, from translation of GCN4 mRNA, the protein on a This is on two Whereas in the of expression is by nitrogen starvation, expression of reporter is still to a When uORFs are in theGCN4 expression translation is by nitrogen starvation, but both protein levels and expression of reporter are be that nitrogen starvation protein Whereas amino acid starvation inhibits Gcn4p by the system B. Fink G.R. EMBO J. PubMed Scopus Google Scholar), nitrogen starvation have and of nitrogen sensing and signaling system to translational of two are known that are regulated by the of pseudohyphal and signaling system that of Mep2p, and the cAMP-dependent protein kinase is thought to regulate pseudohyphal development when cells are starved for nitrogen (22Kübler E. Mösch H.-U. Rupp S. Lisanti M.P. J. Biol. Chem. 1997; 272: 20321-20323Abstract Full Text Full Text PDF PubMed Scopus (166) Google Scholar, M.C. Heitman J. Google Scholar). Mep2p Gpa2p to be involved in translational control of In the of yeast the utilization of nitrogen such as and This is as nitrogen J. Mol. PubMed Scopus Google Scholar). The genetic system involves the of the transcription factor Gln3p that is regulated by the protein Ure2p in the of strains which that proteins are involved in GCN4 under nitrogen starvation. The of the intracellular ammonium by the ammonium led to a of Gcn4p This the of an additional sensor system for the intracellular ammonium which is to a for of theGCN4 mRNA studies that yeast cells amino acid biosynthesis only when precursors are Nitrogen starvation represses activity of the general control system but the expression of amino acid biosynthetic genes. This that amino acid biosynthesis is completely even under starvation The of amino acid biosynthesis under conditions is by the regulatory mechanism that we found in yeast cells of amino acid biosynthesis when nitrogen sources are mRNA levels for the Gcn4p are at a in to translation of when nitrogen Yeast cells to their metabolism for optimal growth and Nitrogen starvation requires the expression of genes for enzymes to cells by to a growth In we investigated how starvation for nitrogen affects the “general control” regulatory network that induces amino acid biosynthetic gene expression when yeast cells are starved for amino acids. amino acid biosynthesis requires at network is an important part of the general nitrogen metabolism. Yeast cells by into and (24ter Schure E.G. van Riel N.A. Verrips C.T. FEMS Microbiol. Rev. 2000; 24: 67-83Crossref PubMed Scopus (205) Google Scholar). This to the general of includes the more specific of amino and activates the general control system and its Gcn4p. In we found the that a general of nitrogen source and efficiently inhibits gene expression. of that inhibition of general gene expression by nitrogen starvation at the of GCN4 mRNA We that protein levels in the of levels of Gcn4p levels or expression can only when translated from GCN4 mRNA the uORFs in the leader sequence. Nitrogen starvation completely amino acid expression and Gcn4p on the of the phosphorylation of by Gcn2p is blocked by nitrogen starvation. the we that nitrogen starvation to an even in phosphorylation than amino acid starvation. starvation for nitrogen completely translation of GCN4 mRNA, even under amino acid starvation This that nitrogen starvation GCN4 mRNA translation by an additional nitrogen starvation Gcn2p or protein levels the of a of the general control system that acts of be in or expression. This an additional translational regulation mechanism the Our that nitrogen starvation, from translation of GCN4 mRNA, the protein on a This is on two Whereas in the of expression is by nitrogen starvation, expression of reporter is still to a When uORFs are in theGCN4 expression translation is by nitrogen starvation, but both protein levels and expression of reporter are be that nitrogen starvation protein Whereas amino acid starvation inhibits Gcn4p by the system B. Fink G.R. EMBO J. PubMed Scopus Google Scholar), nitrogen starvation have and of Gcn4p. nitrogen sensing and signaling system to translational of two are known that are regulated by the of pseudohyphal and signaling system that of Mep2p, and the cAMP-dependent protein kinase is thought to regulate pseudohyphal development when cells are starved for nitrogen (22Kübler E. Mösch H.-U. Rupp S. Lisanti M.P. J. Biol. Chem. 1997; 272: 20321-20323Abstract Full Text Full Text PDF PubMed Scopus (166) Google Scholar, M.C. Heitman J. Google Scholar). Mep2p Gpa2p to be involved in translational control of In the of yeast the utilization of nitrogen such as and This is as nitrogen J. Mol. PubMed Scopus Google Scholar). The genetic system involves the of the transcription factor Gln3p that is regulated by the protein Ure2p in the of strains which that proteins are involved in GCN4 under nitrogen starvation. The of the intracellular ammonium by the ammonium led to a of Gcn4p This the of an additional sensor system for the intracellular ammonium which is to a for of theGCN4 mRNA In studies that yeast cells amino acid biosynthesis only when precursors are Nitrogen starvation represses activity of the general control system but the expression of amino acid biosynthetic genes. This that amino acid biosynthesis is completely even under starvation The of amino acid biosynthesis under conditions is by the regulatory mechanism that we found in yeast cells of amino acid biosynthesis when nitrogen sources are mRNA levels for the Gcn4p are at a in to translation of when nitrogen We Hinnebusch for several and and for We are to and for reading the and all of the Braus for We and for several