Dataset Information

Identification of Genes in Saccharomyces cerevisiae that Are Haploinsufficient for Overcoming Amino Acid Starvation.

ABSTRACT: The yeast Saccharomyces cerevisiae responds to amino acid deprivation by activating a pathway conserved in eukaryotes to overcome the starvation stress. We have screened the entire yeast heterozygous deletion collection to identify strains haploinsufficient for growth in the presence of sulfometuron methyl, which causes starvation for isoleucine and valine. We have discovered that cells devoid of MET15 are sensitive to sulfometuron methyl, and loss of heterozygosity at the MET15 locus can complicate screening the heterozygous deletion collection. We identified 138 cases of loss of heterozygosity in this screen. After eliminating the issues of the MET15 loss of heterozygosity, strains isolated from the collection were retested on sulfometuron methyl. To determine the general effect of the mutations for a starvation response, SMM-sensitive strains were tested for the ability to grow in the presence of canavanine, which induces arginine starvation, and strains that were MET15 were also tested for growth in the presence of ethionine, which causes methionine starvation. Many of the genes identified in our study were not previously identified as starvation-responsive genes, including a number of essential genes that are not easily screened in a systematic way. The genes identified span a broad range of biological functions, including many involved in some level of gene expression. Several unnamed proteins have also been identified, giving a clue as to possible functions of the encoded proteins.

SUBMITTER: Bae NS

PROVIDER: S-EPMC5386856 | biostudies-literature | 2017 Apr

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Identification of Genes in Saccharomyces cerevisiae that Are Haploinsufficient for Overcoming Amino Acid Starvation.

Bae Nancy S NS Seberg Andrew P AP Carroll Leslie P LP Swanson Mark J MJ

G3 (Bethesda, Md.) 20170403 4

The yeast Saccharomyces cerevisiae responds to amino acid deprivation by activating a pathway conserved in eukaryotes to overcome the starvation stress. We have screened the entire yeast heterozygous deletion collection to identify strains haploinsufficient for growth in the presence of sulfometuron methyl, which causes starvation for isoleucine and valine. We have discovered that cells devoid of MET15 are sensitive to sulfometuron methyl, and loss of heterozygosity at the MET15 ...[more]

PMID: 28209762

Similar Datasets

Project description:BackgroundAcetic acid is a byproduct of Saccharomyces cerevisiae alcoholic fermentation. Together with high concentrations of ethanol and other toxic metabolites, acetic acid may contribute to fermentation arrest and reduced ethanol productivity. This weak acid is also a present in lignocellulosic hydrolysates, a highly interesting non-feedstock substrate in industrial biotechnology. Therefore, the better understanding of the molecular mechanisms underlying S. cerevisiae tolerance to acetic acid is essential for the rational selection of optimal fermentation conditions and the engineering of more robust industrial strains to be used in processes in which yeast is explored as cell factory.ResultsThe yeast genes conferring protection against acetic acid were identified in this study at a genome-wide scale, based on the screening of the EUROSCARF haploid mutant collection for susceptibility phenotypes to this weak acid (concentrations in the range 70-110 mM, at pH 4.5). Approximately 650 determinants of tolerance to acetic acid were identified. Clustering of these acetic acid-resistance genes based on their biological function indicated an enrichment of genes involved in transcription, internal pH homeostasis, carbohydrate metabolism, cell wall assembly, biogenesis of mitochondria, ribosome and vacuole, and in the sensing, signalling and uptake of various nutrients in particular iron, potassium, glucose and amino acids. A correlation between increased resistance to acetic acid and the level of potassium in the growth medium was found. The activation of the Snf1p signalling pathway, involved in yeast response to glucose starvation, is demonstrated to occur in response to acetic acid stress but no evidence was obtained supporting the acetic acid-induced inhibition of glucose uptake.ConclusionsApproximately 490 of the 650 determinants of tolerance to acetic acid identified in this work are implicated, for the first time, in tolerance to this weak acid. These are novel candidate genes for genetic engineering to obtain more robust yeast strains against acetic acid toxicity. Among these genes there are number of transcription factors that are documented regulators of a large percentage of the genes found to exert protection against acetic acid thus being considered interesting targets for subsequent genetic engineering. The increase of potassium concentration in the growth medium was found to improve the expression of maximal tolerance to acetic acid, consistent with the idea that the adequate manipulation of nutrient concentration of industrial growth medium can be an interesting strategy to surpass the deleterious effects of this weak acid in yeast cells.

Project description:Ticks are obligate hematophagous arthropods and must tolerate starvation during off-host periods. Macroautophagy (hereafter autophagy) is a well-conserved self-eating mechanism of cell survival and is essential for recycling cellular contents during periods of starvation, stress, and injury in organisms. Although the genome sequence of Ixodes scapularis (Say) is available, the characteristics and functions of autophagy-related gene families remain largely unknown. To advance our understanding of autophagy in I. scapularis, we used comprehensive genomic approaches to identify Atg genes. Homologues of 14 Atg genes were identified, and their protein motif compositions were predicted. Phylogenetic analysis indicated that ATGs in I. scapularis were evolutionarily closely related to their homologues in Haemaphysalis longicornis and Rhipicephalus microplus ticks. Expression patterns of Atg genes differed across tick developmental stages. Immunofluorescence results by monodansylcadaverine (MDC) staining indicated that autophagy was activated after amino acid starvation treatments in I. scapularis embryo-derived cell lines ISE6 and IDE8. Subsequently, the expression of key Atg genes involved in autophagy pathway in both cell lines were examined. In ISE6 cells, the expression levels of three Atg genes (Atg4B, Atg6 and Atg8A) increased significantly after amino acid starvation; similarly, four Atg genes (Atg4A, Atg4B, Atg6 and Atg8B) were upregulated in IDE8 cells in response to starvation. In parallel, the MDC and lysotracker staining results indicated that autophagy was triggered after amino acid starvation treatments in R. microplus embryo-derived cell line BME26. Our observations showed that Atg family genes are highly conserved in ticks and function in autophagy pathway induced by amino acid starvation. These results also provide valuable insight for further autophagy-related research as a new strategy for blocking the transmission of tick-borne pathogens.

Project description:The isoleucine and valine biosynthetic enzyme acetolactate synthase (Ilv2p) is an attractive antifungal drug target, since the isoleucine and valine biosynthetic pathway is not present in mammals, Saccharomyces cerevisiae ilv2Delta mutants do not survive in vivo, Cryptococcus neoformans ilv2 mutants are avirulent, and both S. cerevisiae and Cr. neoformans ilv2 mutants die upon isoleucine and valine starvation. To further explore the potential of Ilv2p as an antifungal drug target, we disrupted Candida albicans ILV2, and demonstrated that Ca. albicans ilv2Delta mutants were significantly attenuated in virulence, and were also profoundly starvation-cidal, with a greater than 100-fold reduction in viability after only 4 h of isoleucine and valine starvation. As fungicidal starvation would be advantageous for drug design, we explored the basis of the starvation-cidal phenotype in both S. cerevisiae and Ca. albicans ilv2Delta mutants. Since the mutation of ILV1, required for the first step of isoleucine biosynthesis, did not suppress the ilv2Delta starvation-cidal defects in either species, the cidal phenotype was not due to alpha-ketobutyrate accumulation. We found that starvation for isoleucine alone was more deleterious in Ca. albicans than in S. cerevisiae, and starvation for valine was more deleterious than for isoleucine in both species. Interestingly, while the target of rapamycin (TOR) pathway inhibitor rapamycin further reduced S. cerevisiae ilv2Delta starvation viability, it increased Ca. albicans ilv1Delta and ilv2Delta viability. Furthermore, the recovery from starvation was dependent on the carbon source present during recovery for S. cerevisiae ilv2Delta mutants, reminiscent of isoleucine and valine starvation inducing a viable but non-culturable-like state in this species, while Ca. albicans ilv1Delta and ilv2 Delta viability was influenced by the carbon source present during starvation, supporting a role for glucose wasting in the Ca. albicans cidal phenotype.

Dataset Information

Identification of Genes in Saccharomyces cerevisiae that Are Haploinsufficient for Overcoming Amino Acid Starvation.

Publications

Identification of Genes in <i>Saccharomyces cerevisiae</i> that Are Haploinsufficient for Overcoming Amino Acid Starvation.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets