Transcriptional Profiling Shows that Gcn4p Is a Master Regulator of Gene Expression during Amino Acid Starvation in Yeast

Abstract

Starvation for amino acids induces Gcn4p, a transcriptional activator of amino acid biosynthetic genes in Saccharomyces cerevisiae. In an effort to identify all genes regulated by Gcn4p during amino acid starvation, we performed cDNA microarray analysis. Data from 21 pairs of hybridization experiments using two different strains derived from S288c revealed that more than 1,000 genes were induced, and a similar number were repressed, by a factor of 2 or more in response to histidine starvation imposed by 3-aminotriazole (3AT). Profiling of a gcn4⌬ strain and a constitutively induced mutant showed that Gcn4p is required for the full induction by 3AT of at least 539 genes, termed Gcn4p targets. Genes in every amino acid biosynthetic pathway except cysteine and genes encoding amino acid precursors, vitamin biosynthetic enzymes, peroxisomal components, mitochondrial carrier proteins, and autophagy proteins were all identified as Gcn4p targets. Unexpectedly, genes involved in amino acid biosynthesis represent only a quarter of the Gcn4p target genes. Gcn4p also activates genes involved in glycogen homeostasis, and mutant analysis showed that Gcn4p suppresses glycogen levels in amino acid-starved cells. Numerous genes encoding protein kinases and transcription factors were identified as targets, suggesting that Gcn4p is a master regulator of gene expression. Interestingly, expression profiles for 3AT and the alkylating agent methyl methanesulfonate (MMS) overlapped extensively, and MMS induced GCN4 translation. Thus, the broad transcriptional response evoked by Gcn4p is produced by diverse stress conditions. Finally, profiling of a gcn4⌬ mutant uncovered an alternative induction pathway operating at many Gcn4p target genes in histidine-starved cells.In response to environmental perturbations, Saccharomyces cerevisiae cells elicit rapid transcriptional reprogramming involving both activation and repression of gene expression. Transcriptional activator proteins function by binding to specific promoter elements, called upstream activating sequences (UASs) in yeast cells, and recruiting the transcriptional machinery. Thus, transcriptional stimulation requires the expression and function of an activator and the appropriate UAS element in the promoters of its target genes. A plethora of mechanisms are known to regulate the activity or expression of transcriptional activators in response to specific signals. For example, in cells grown on glucose, Gal80p inhibits the ability of Gal4p to activate transcription of genes encoding galactosemetabolizing enzymes, whereas Gal3p alleviates this inhibition on galactose medium (83). The transcriptional activators Pho4p, Swi5p, and Yap1p are regulated by the coupling of their nuclear localization to the levels of inorganic phosphate, cell cycle and mother-daughter status, or oxidative stress, respectively (reviewed in reference 52). Starvation for amino acids, purines, and glucose limitation induces the synthesis of Gcn4p, a bZIP transcriptional activator of amino acid biosynthetic gene...

Membrane stress is coupled to a rapid translational control of gene expression in chlorpromazine-treated cells

“…In contrast, the addition of 3-aminotriazole (3-AT; an inhibitor of histidine biosynthesis) increased HIS4-lacZ activity in an eIF2 phosphorylation-dependent manner. This Wnding strengthens previous results showing that the expression of Gcn4p is not only dedicated to the control of amino acid biosynthesis (Natarajan et al 2001). Notably Gcn4p also regulates gene products involved in degradation of autophagosomes in a process known as autophagy (Klionsky and Emr 2000).…”

“…In this context, CPZ also induces GCN4 expression. Translational induction of GCN4 is part of the cellular response to changes in the environment such as depletion of amino acids (Natarajan et al 2001). In CPZ-treated cells, the expression of Gcn4p depends on the Gcn2p-stimulated eIF2 phosphorylation.…”

“…In addition to inhibiting translation initiation of almost all mRNAs, eIF2 -P activates the expression of Gcn4p by a unique translational control mechanism (reviewed in Hinnebusch 2005). Gcn4p is the major transcriptional activator of amino acid biosynthetic genes that is activated under amino acid starvation (Natarajan et al 2001), a cellular response named general amino acid control (GAAC).…”

Nonsense surveillance regulates expression of diverse classes of mammalian transcripts and mutes genomic noise

“…NMD inhibition was also associated with increased expression of crucial mediators of autophagy (bulk vacuolar degradation of cytosolic proteins to recycle amino acids in response to starvation) and sporulation. Multiple yeast transcripts that show increased expression after NMD inhibition (e.g., TRP1, ATG12 (also called APG12) and DAL5) have been shown experimentally to lack responsiveness to Gcn4p, a master transcriptional activator of amino acid biosynthetic genes in multiple pathways 26 , suggestive of complementarity between transcriptional and post-transcriptional responses to amino acid deprivation. Consistent with the paradigm that NMD has been functionally incorporated into homeostatic mechanisms, we also observed upregulation of the human homolog of Caenorhabditis elegans smg-5, a gene that is essential for mammalian NMD 27 , in NMD-deficient HeLa cells.…”

Life in the midst of scarcity: adaptations to nutrient availability in Saccharomyces cerevisiae

“…The central component of this pathway is Gcn4, a transcription factor important for activating transcription of genes needed for amino acid biosynthesis in response to amino acid starvation (Natarajan et al 2001). The pathway is induced by uncharged tRNAs, which presumably activate the kinase Gcn2.…”