Project description:Transcriptome profiling of cdc14-3 and cdc14-3_snf1 mutants at non-permissive temp (35C) for 90min or 2hr followed by 30min NaCl stress. The experiment showed the aberrant cell cycle induction in cdc14-3 mutants upon NaCl response is abrogated with snf1 deletion in cdc14-3 background.
Project description:Alkaline pH stress invokes in S. cerevisiae a potent and fast transcriptional response that includes many genes repressed by glucose. Certain mutants in the glucose-sensing and response pathways, such as those lacking the Snf1 kinase, are sensitive to alkalinization. We show that addition of glucose to the medium improves growth of wild type cells at high pH, fully abolish the snf1 alkali-sensitive phenotype and attenuates high pH-induced Snf1 phosphorylation at Thr210. The elm1 mutant, lacking one of the three upstream Snf1 kinases (tos3, elm1 and sak1), is markedly alkali sensitive, whereas the phenotype of the tos3 elm1 sak1 strain is even stronger than that of snf1 cells and it is not fully rescued by glucose supplementation. DNA microarray analysis reveals that about 75% of genes induced at short term by high pH are also induced by glucose scarcity. Snf1 mediates, in full or in part, the activation of a significant subset (38%) of short-term alkali-induced genes, including those coding high-affinity hexose transporters and phosphorylating enzymes. Induction of genes encoding enzymes involved in glycogen (but not trehalose) metabolism is largely dependent of the presence of Snf1. Therefore, the function of Snf1 in adaptation to glucose scarcity appears crucial for alkaline pH tolerance. Incorporation of micromolar amounts of iron and copper to a glucose-supplemented medium result in an additive effect and allows near normal growth at high pH, thus indicating that these three nutrients are key limiting factors for growth in an alkaline environment.
Project description:Snf1 and TORC1 are two global regulators that sense the nutrient availability and regulate the cell growth in yeast Saccharomyces cerevisiae. Here we undertook a systems biology approach to study the effect of deletion of these genes and investigate the interaction between Snf1 and TORC1 in regulation of gene expression and cell metabolism.
Project description:CDC14 phosphatases are critical components of the cell cycle machinery that drives exit from mitosis in yeast. However, the two mammalian paralogs, CDC14A and CDC14B, are dispensable for cell cycle progression or exit, and their function remains unclear. By generating a double Cdc14a; Cdc14b-null mouse model, we report here that CDC14 phosphatases control cell differentiation in pluripotent cells and their absence results in deficient development of the neural system. Lack of CDC14 impairs neural differentiation from embryonic stem cells (ESCs) accompanied by deficient induction of genes controlled by bivalent promoters. During ESC differentiation, CDC14 directly dephosphorylates and destabilizes Undifferentiated embryonic Transcription Factor 1 (UTF1), a critical regulator of stemness. In the absence of CDC14, increased UTF1 levels prevent the firing of bivalent promoters, resulting in defective induction of the transcriptional programs required for differentiation. These results suggest that mammalian CDC14 phosphatases function during the terminal exit from the cell cycle by modulating the transition from the pluripotent to the differentiated chromatin state, at least partially by controlling chromatin dynamics and transcription in a UTF1-dependent manner.
Project description:Alkaline pH stress invokes in S. cerevisiae a potent and fast transcriptional response that includes many genes repressed by glucose. Certain mutants in the glucose-sensing and response pathways, such as those lacking the Snf1 kinase, are sensitive to alkalinization. We show that addition of glucose to the medium improves growth of wild type cells at high pH, fully abolish the snf1 alkali-sensitive phenotype and attenuates high pH-induced Snf1 phosphorylation at Thr210. The elm1 mutant, lacking one of the three upstream Snf1 kinases (tos3, elm1 and sak1), is markedly alkali sensitive, whereas the phenotype of the tos3 elm1 sak1 strain is even stronger than that of snf1 cells and it is not fully rescued by glucose supplementation. DNA microarray analysis reveals that about 75% of genes induced at short term by high pH are also induced by glucose scarcity. Snf1 mediates, in full or in part, the activation of a significant subset (38%) of short-term alkali-induced genes, including those coding high-affinity hexose transporters and phosphorylating enzymes. Induction of genes encoding enzymes involved in glycogen (but not trehalose) metabolism is largely dependent of the presence of Snf1. Therefore, the function of Snf1 in adaptation to glucose scarcity appears crucial for alkaline pH tolerance. Incorporation of micromolar amounts of iron and copper to a glucose-supplemented medium result in an additive effect and allows near normal growth at high pH, thus indicating that these three nutrients are key limiting factors for growth in an alkaline environment. We identified the changes in the expression profiles caused by alkalinization of the medium (pH8 vs. pH5.5 for 10 min) in several strains: wild type cells (4 chips), snf1 mutant cells (4 chips) We also identified the transcriptomic changes that occur after glucose deprivation (0.05% vs 2% for 15 min) in: wild type cells (2 chips) snf1 mutant cells (2 chips) Total: 12 chips
Project description:CDC14 phosphatases are critical components of the cell cycle machinery that drives exit from mitosis in yeast. However, the two mammalian paralogs, CDC14A and CDC14B, are dispensable for cell cycle progression or exit, and their function remains unclear. By generating a double Cdc14a; Cdc14b -null mouse model, we report here that CDC14 phosphatases control cell differentiation in pluripotent cells, and their absence results in deficient development of the neural system. Lack of CDC14 impairs neural differentiation from embryonic stem cells (ESCs) accompanied by deficient induction of genes controlled by bivalent promoters. CDC14 directly dephosphorylates and destabilizes Undifferentiated embryonic Transcription Factor 1 (UTF1) during the exit from stemness. Multiomic single-cell analysis of differentiating ESCs suggest that increased UTF1 levels in the absence of CDC14 prevent the firing of bivalent promoters required for differentiation. These results, along recent data suggesting a critical role for cell cycle kinases in pluripotency, suggest that cell cycle kinase-phosphatase modules such as CDK-CDC14 are critical for linking cell cycle regulation and self-renewal, with a specific function for CDC14 phosphatases modulating key epigenetic regulators during the terminal exit from pluripotency.
Project description:CDC14 phosphatases are critical components of the cell cycle machinery that drives exit from mitosis in yeast. However, the two mammalian paralogs, CDC14A and CDC14B, are dispensable for cell cycle progression or exit, and their function remains unclear. By generating a double Cdc14a; Cdc14b -null mouse model, we report here that CDC14 phosphatases control cell differentiation in pluripotent cells, and their absence results in deficient development of the neural system. Lack of CDC14 impairs neural differentiation from embryonic stem cells (ESCs) accompanied by deficient induction of genes controlled by bivalent promoters. CDC14 directly dephosphorylates and destabilizes Undifferentiated embryonic Transcription Factor 1 (UTF1) during the exit from stemness. Multiomic single-cell analysis of differentiating ESCs suggest that increased UTF1 levels in the absence of CDC14 prevent the firing of bivalent promoters required for differentiation. These results, along recent data suggesting a critical role for cell cycle kinases in pluripotency, suggest that cell cycle kinase-phosphatase modules such as CDK-CDC14 are critical for linking cell cycle regulation and self-renewal, with a specific function for CDC14 phosphatases modulating key epigenetic regulators during the terminal exit from pluripotency.
Project description:Snf1 and TORC1 are two global regulators that sense the nutrient availability and regulate the cell growth in yeast Saccharomyces cerevisiae. Here we undertook a systems biology approach to study the effect of deletion of these genes and investigate the interaction between Snf1 and TORC1 in regulation of gene expression and cell metabolism. 3 mutant strains (snf1?, tor1?, snf1?tor1?) together with 1 reference strain grown under both glucose-limited or amonia-limited defined media with three biological replicates for each strain
Project description:The energy metabolism pathways are significantly influenced by the available carbon sources. In Saccharomyces cerevisiae, energy is primarily produced through aerobic respiration during glycerol cultivation, a process believed to depend on the yeast AMPK (AMP-activated protein kinase) homolog, Snf1. It has been reported that Snf1 increases the expression of respiratory genes by phosphorylating transcriptional activators in environments where glucose is unavailable. We discovered that Tda1, activated by Snf1, phosphorylates Hxk2. Hxk2 has been reported to function as a transcriptional repressor. Therefore, we analyzed how Tda1 affects the expression of respiratory genes by RNA-seq analysis of glycerol-cultured cells.