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

Bae, Nancy S.; Seberg, Andrew P.; Carroll, Leslie P; Swanson, Mark J.

doi:10.1534/g3.116.037416

Cited by 9 publications

(8 citation statements)

References 175 publications

(249 reference statements)

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“…3E). Deletion of GCN2 completely blocks HIS4 induction in response to ILV2 depletion, consistent with canonical models of ISR signaling (25). Although HTS1 knockdown triggers strong eIF2α phosphorylation, GCN2 deletion only partially abrogated its transcriptional effects.…”

Section: Trna Insufficiency Triggers His4 Transcription Independentlysupporting

confidence: 78%

CiBER-seq dissects genetic networks by quantitative CRISPRi profiling of expression phenotypes

Muller¹,

Meacham²,

Ferguson³

et al. 2020

Preprint

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To realize the promise of CRISPR/Cas9-based genetics, approaches are needed to quantify a specific, molecular phenotype across genome-wide libraries of genetic perturbations. We address this challenge by profiling transcriptional, translational, and post-translational reporters using CRISPR interference with barcoded expression reporter sequencing (CiBER-seq). Our barcoding approach connects an entire library of guides to their individual phenotypic consequences using pooled sequencing. We show that CiBER-seq profiling fully recapitulates the integrated stress response (ISR) pathway in yeast. Genetic perturbations causing uncharged tRNA accumulation activated ISR reporter transcription. Surprisingly, tRNA insufficiency also activated the reporter, independent of the Gcn2 kinase that senses uncharged tRNAs. By uncovering alternate triggers for ISR activation, we illustrate how precise, comprehensive CiBER-seq profiling provides a powerful and broadly applicable tool for dissecting genetic networks.

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Section: Trna Insufficiency Triggers His4 Transcription Independentlysupporting

confidence: 78%

CiBER-seq dissects genetic networks by quantitative CRISPRi profiling of expression phenotypes

Muller¹,

Meacham²,

Ferguson³

et al. 2020

Preprint

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“…Yeast cells are known to respond at the level of translation to a multitude of environmental stresses, such as heat shock [11,12], nutrient deprivation [13,14,15], general amino acid balance [16,17,18,19], phosphate [20], sulfur [21,22], nitrogen [23,24] content and other environmental conditions [25,26], such as growth at low/high temperatures [27], in different oxygen levels [28,29,30] or in the presence of heavy metals, during desiccation [31,32,33], etc. Many of these conditions are studied in the context of industrial applications of yeast in bread baking and alcohol fermentation [34,35,36,37,38].…”

Section: Introductionmentioning

confidence: 99%

Control of Translation at the Initiation Phase During Glucose Starvation in Yeast

Janapala

Preiß

Shirokikh

2019

IJMS

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Glucose is one of the most important sources of carbon across all life. Glucose starvation is a key stress relevant to all eukaryotic cells. Glucose starvation responses have important implications in diseases, such as diabetes and cancer. In yeast, glucose starvation causes rapid and dramatic effects on the synthesis of proteins (mRNA translation). Response to glucose deficiency targets the initiation phase of translation by different mechanisms and with diverse dynamics. Concomitantly, translationally repressed mRNAs and components of the protein synthesis machinery may enter a variety of cytoplasmic foci, which also form with variable kinetics and may store or degrade mRNA. Much progress has been made in understanding these processes in the last decade, including with the use of high-throughput/omics methods of RNA and RNA:protein detection. This review dissects the current knowledge of yeast reactions to glucose starvation systematized by the stage of translation initiation, with the focus on rapid responses. We provide parallels to mechanisms found in higher eukaryotes, such as metazoans, for the most critical responses, and point out major remaining gaps in knowledge and possible future directions of research on translational responses to glucose starvation.

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“…The genetic screen of the BY4742 haploid knockout collection was performed via the phenotyping of serial dilution growth on solid media by adapting a previous screen technique ( Bae et al , 2017 ). In short, frozen 96-well microtiter plates of the knockout collection were thawed and inoculated via pinning into 96-well growth chambers containing YPD in biological triplicate.…”

Section: Methodsmentioning

confidence: 99%

Oxidative Stress Responses and Nutrient Starvation in MCHM Treated Saccharomyces cerevisiae

Ayers¹,

Sherman²,

Gallagher³

2020

G3 Genes|Genomes|Genetics

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In 2014, the coal cleaning chemical 4-methylcyclohexane methanol (MCHM) spilled into the water supply for 300,000 West Virginians. Initial toxicology tests showed relatively mild results, but the underlying effects on cellular biology were underexplored. Treated wildtype yeast cells grew poorly, but there was only a small decrease in cell viability. Cell cycle analysis revealed an absence of cells in S phase within thirty minutes of treatment. Cells accumulated in G1 over a six-hour time course, indicating arrest instead of death. A genetic screen of the haploid knockout collection revealed 329 high confidence genes required for optimal growth in MCHM. These genes encode three major cell processes: mitochondrial gene expression/translation, the vacuolar ATPase, and aromatic amino acid biosynthesis. The transcriptome showed an upregulation of pleiotropic drug response genes and amino acid biosynthetic genes and downregulation in ribosome biosynthesis. Analysis of these datasets pointed to environmental stress response activation upon treatment. Overlap in datasets included the aromatic amino acid genes ARO1, ARO3, and four of the five TRP genes. This implicated nutrient deprivation as the signal for stress response. Excess supplementation of nutrients and amino acids did not improve growth on MCHM, so the source of nutrient deprivation signal is still unclear. Reactive oxygen species and DNA damage were directly detected with MCHM treatment, but timepoints showed these accumulated slower than cells arrested. We propose that wildtype cells arrest from nutrient deprivation and survive, accumulating oxidative damage through the implementation of robust environmental stress responses.

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Identification of Genes inSaccharomyces cerevisiaethat Are Haploinsufficient for Overcoming Amino Acid Starvation

Cited by 9 publications

References 175 publications

CiBER-seq dissects genetic networks by quantitative CRISPRi profiling of expression phenotypes

CiBER-seq dissects genetic networks by quantitative CRISPRi profiling of expression phenotypes

Control of Translation at the Initiation Phase During Glucose Starvation in Yeast

Oxidative Stress Responses and Nutrient Starvation in MCHM Treated Saccharomyces cerevisiae

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