scholarly journals Thickening of the cell wall increases the resistance of S. cerevisiae to commercial formulations of glyphosate

2019 ◽  
Author(s):  
Apoorva Ravishankar ◽  
Amaury Pupo ◽  
Jennifer E.G. Gallagher
Keyword(s):  
Gene Expression ◽  
Saccharomyces Cerevisiae ◽  
Cell Wall ◽  
Adverse Effects ◽  
Mapk Signaling ◽  
Fragile Sites ◽  
Yeast Cells ◽  
Global Changes ◽  
Wide Range ◽  
A Cell

AbstractThe use of glyphosate-based herbicides is widespread and despite its extensive use, its effects are yet to be deciphered completely. The additives in commercial formulations of glyphosate, though labeled as inert when used individually, have adverse effects when used in combination with other additives and the active ingredient. As a species, Saccharomyces cerevisiae has a wide range of resistance to glyphosate-based herbicides. To investigate the underlying genetic differences between sensitive and resistant strains, global changes in gene expression were measured when yeast were exposed to a commercial formulation of glyphosate (CFG). Changes in gene expression involved in numerous pathways such as DNA replication, MAPK signaling, meiosis, and cell wall synthesis. Because so many diverse pathways were affected, these strains were then subjected to in-lab-evolutions (ILE) to select mutations that confer increased resistance. Common fragile sites were found to play a role in adaptation mechanisms used by cells to attain resistance with long-term exposure to CFG. The cell wall structure acts as a protective barrier in alleviating the stress caused by exposure to CFG. The thicker the cell wall, the more resistant the cell is against CFG. Hence, a detailed study of the changes occurring at the genome and transcriptome level is essential to better understand the possible effects of CFG on the cell as a whole.Author SummaryWe are exposed to various chemicals in the environment on a daily basis. Some of these chemicals are herbicides that come in direct contact with the food we consume. This makes the thorough investigation of these chemicals crucial. Some of the most commonly used herbicides around the world are glyphosate-based. Their mode of action effects a biosynthetic pathway that is absent in mammals and insects and so it is deemed safe for consumption. However, there are many additives to these herbicides that increase its effects. Thorough testing of these commercially available herbicides is essential to decipher all the potentially adverse effects that it could have on a cell. Saccharomyces cerevisiae has a wide range of genetic diversity, making it is suitable to test different chemicals and identify any harmful effects. In this study, we exposed yeast cells to some glyphosate-based herbicides available in the market, to understand what effects it could have on a cell. We found that the additives in the herbicides have an effect on the cell wall and the mode of entry of glyphosate into the cell.

scholarly journals Resistance Mechanisms of Saccharomyces cerevisiae to Commercial Formulations of Glyphosate Involve DNA Damage Repair, the Cell Cycle, and the Cell Wall Structure

2020 ◽  
Vol 10 (6) ◽  
pp. 2043-2056
Author(s):  
Apoorva Ravishankar ◽  
Amaury Pupo ◽  
Jennifer E. G. Gallagher
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Wall ◽  
Resistance Mechanisms ◽  
Mapk Signaling ◽  
Fragile Sites ◽  
Cell Wall Structure ◽  
Wall Structure ◽  
Global Changes ◽  
Expression Of Genes ◽  
Wide Range

The use of glyphosate-based herbicides is widespread and despite their extensive use, their effects are yet to be deciphered completely. The additives in commercial formulations of glyphosate, though labeled inert when used individually, have adverse effects when used in combination with other additives along with the active ingredient. As a species, Saccharomyces cerevisiae has a wide range of resistance to glyphosate-based herbicides. To investigate the underlying genetic differences between sensitive and resistant strains, global changes in gene expression were measured, when yeast were exposed to a glyphosate-based herbicide (GBH). Expression of genes involved in numerous pathways crucial to the cell’s functioning, such as DNA replication, MAPK signaling, meiosis, and cell wall synthesis changed. Because so many diverse pathways were affected, these strains were then subjected to in-lab-evolutions (ILE) to select mutations that confer increased resistance. Common fragile sites were found to play a role in adaptation to resistance to long-term exposure of GBHs. Copy number increased in approximately 100 genes associated with cell wall proteins, mitochondria, and sterol transport. Taking ILE and transcriptomic data into account it is evident that GBHs affect multiple biological processes in the cell. One such component is the cell wall structure which acts as a protective barrier in alleviating the stress caused by exposure to inert additives in GBHs. Sed1, a GPI-cell wall protein, plays an important role in tolerance of a GBH. Hence, a detailed study of the changes occurring at the genome and transcriptome levels is essential to better understand the effects of an environmental stressor such as a GBH, on the cell as a whole.

scholarly journals Yeast Upf Proteins Required for RNA Surveillance Affect Global Expression of the Yeast Transcriptome

1999 ◽  
Vol 19 (10) ◽  
pp. 6710-6719 ◽  
Author(s):  
Michael J. Lelivelt ◽  
Michael R. Culbertson
Keyword(s):  
Gene Expression ◽  
Saccharomyces Cerevisiae ◽  
Protein Sequences ◽  
Average Level ◽  
Global Changes ◽  
Oligonucleotide Arrays ◽  
Rna Surveillance ◽  
Wide Range ◽  
Information Center ◽  
Nonsense Mediated Mrna Decay

ABSTRACT mRNAs are monitored for errors in gene expression by RNA surveillance, in which mRNAs that cannot be fully translated are degraded by the nonsense-mediated mRNA decay pathway (NMD). RNA surveillance ensures that potentially deleterious truncated proteins are seldom made. NMD pathways that promote surveillance have been found in a wide range of eukaryotes. In Saccharomyces cerevisiae, the proteins encoded by the UPF1, UPF2, andUPF3 genes catalyze steps in NMD and are required for RNA surveillance. In this report, we show that the Upf proteins are also required to control the total accumulation of a large number of mRNAs in addition to their role in RNA surveillance. High-density oligonucleotide arrays were used to monitor global changes in the yeast transcriptome caused by loss of UPF gene function. Null mutations in the UPF genes caused altered accumulation of hundreds of mRNAs. The majority were increased in abundance, but some were decreased. The same mRNAs were affected regardless of which of the three UPF gene was inactivated. The proteins encoded byUPF-dependent mRNAs were broadly distributed by function but were underrepresented in two MIPS (Munich Information Center for Protein Sequences) categories: protein synthesis and protein destination. In a UPF + strain, the average level of expression of UPF-dependent mRNAs was threefold lower than the average level of expression of all mRNAs in the transcriptome, suggesting that highly abundant mRNAs were underrepresented. We suggest a model for how the abundance of hundreds of mRNAs might be controlled by the Upf proteins.

scholarly journals The Pep4p vacuolar proteinase contributes to the turnover of oxidized proteins but PEP4 overexpression is not sufficient to increase chronological lifespan in Saccharomyces cerevisiae

Microbiology ◽  
2006 ◽  
Vol 152 (12) ◽  
pp. 3595-3605 ◽  
Author(s):  
Marta Marques ◽  
Dominik Mojzita ◽  
Maria A. Amorim ◽  
Teresa Almeida ◽  
Stefan Hohmann ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Gene Expression ◽  
Saccharomyces Cerevisiae ◽  
Protein Turnover ◽  
Yeast Cells ◽  
Global Changes ◽  
20S Proteasome ◽  
Oxidized Proteins ◽  
Genes Encoding ◽  
Induced Genes

Turnover of damaged molecules is considered to play a key role in housekeeping of cells exposed to oxidative stress, and during the progress of ageing. In this work, global changes in the transcriptome were analysed during recovery of yeast cells after H2O2 stress. Regarding induced genes, those associated with protein fate were the most significantly over-represented. In addition to genes encoding subunits of the 20S proteasome, genes related to vacuolar proteolysis (PEP4 and LAP4), protein sorting into the vacuole, and vacuolar fusion were found to be induced. The upregulation of PEP4 gene expression was associated with an increase in Pep4p activity. The induction of genes related to proteolysis was correlated with an increased protein turnover after H2O2-induced oxidation. Furthermore, protein degradation and the removal of oxidized proteins decreased in Pep4p-deficient cells. Pep4p activity also increased during chronological ageing, and cells lacking Pep4p displayed a shortened lifespan associated with higher levels of carbonylated proteins. PEP4 overexpression prevented the accumulation of oxidized proteins, but did not increase lifespan. These results indicate that Pep4p is important for protein turnover after oxidative damage; however, increased removal of oxidized proteins is not sufficient to enhance lifespan.

scholarly journals Screening and Genetic Network Analysis of Genes Involved in Freezing and Thawing Resistance in DaMDHAR—Expressing Saccharomyces cerevisiae Using Gene Expression Profiling

Genes ◽  
2021 ◽  
Vol 12 (2) ◽  
pp. 219
Author(s):  
Il-Sup Kim ◽  
Woong Choi ◽  
Jonghyeon Son ◽  
Jun Hyuck Lee ◽  
Hyoungseok Lee ◽  
...  
Keyword(s):  
Gene Expression ◽  
Saccharomyces Cerevisiae ◽  
Transcription Factors ◽  
Stress Tolerance ◽  
Genetic Network ◽  
Yeast Cells ◽  
Enzymatic Antioxidants ◽  
Freezing And Thawing ◽  
Metal Ion Homeostasis ◽  
Transgenic Yeast

The cryoprotection of cell activity is a key determinant in frozen-dough technology. Although several factors that contribute to freezing tolerance have been reported, the mechanism underlying the manner in which yeast cells respond to freezing and thawing (FT) stress is not well established. Therefore, the present study demonstrated the relationship between DaMDHAR encoding monodehydroascorbate reductase from Antarctic hairgrass Deschampsia antarctica and stress tolerance to repeated FT cycles (FT2) in transgenic yeast Saccharomyces cerevisiae. DaMDHAR-expressing yeast (DM) cells identified by immunoblotting analysis showed high tolerance to FT stress conditions, thereby causing lower damage for yeast cells than wild-type (WT) cells with empty vector alone. To detect FT2 tolerance-associated genes, 3′-quant RNA sequencing was employed using mRNA isolated from DM and WT cells exposed to FT (FT2) conditions. Approximately 332 genes showed ≥2-fold changes in DM cells and were classified into various groups according to their gene expression. The expressions of the changed genes were further confirmed using western blot analysis and biochemical assay. The upregulated expression of 197 genes was associated with pentose phosphate pathway, NADP metabolic process, metal ion homeostasis, sulfate assimilation, β-alanine metabolism, glycerol synthesis, and integral component of mitochondrial and plasma membrane (PM) in DM cells under FT2 stress, whereas the expression of the remaining 135 genes was partially related to protein processing, selenocompound metabolism, cell cycle arrest, oxidative phosphorylation, and α-glucoside transport under the same condition. With regard to transcription factors in DM cells, MSN4 and CIN5 were activated, but MSN2 and MGA1 were not. Regarding antioxidant systems and protein kinases in DM cells under FT stress, CTT1, GTO, GEX1, and YOL024W were upregulated, whereas AIF1, COX2, and TRX3 were not. Gene activation represented by transcription factors and enzymatic antioxidants appears to be associated with FT2-stress tolerance in transgenic yeast cells. RCK1, MET14, and SIP18, but not YPK2, have been known to be involved in the protein kinase-mediated signalling pathway and glycogen synthesis. Moreover, SPI18 and HSP12 encoding hydrophilin in the PM were detected. Therefore, it was concluded that the genetic network via the change of gene expression levels of multiple genes contributing to the stabilization and functionality of the mitochondria and PM, not of a single gene, might be the crucial determinant for FT tolerance in DaMDAHR-expressing transgenic yeast. These findings provide a foundation for elucidating the DaMDHAR-dependent molecular mechanism of the complex functional resistance in the cellular response to FT stress.

Codon replacement in the PGK1 gene of Saccharomyces cerevisiae: experimental approach to study the role of biased codon usage in gene expression

1987 ◽  
Vol 7 (8) ◽  
pp. 2914-2924
Author(s):  
A Hoekema ◽  
R A Kastelein ◽  
M Vasser ◽  
H A de Boer
Keyword(s):  
Gene Expression ◽  
Saccharomyces Cerevisiae ◽  
Steady State ◽  
Codon Usage ◽  
Experimental Approach ◽  
Mrna Translation ◽  
Yeast Cells ◽  
Expression Level ◽  
Start Codon ◽  
Mrna Levels

The coding sequences of genes in the yeast Saccharomyces cerevisiae show a preference for 25 of the 61 possible coding triplets. The degree of this biased codon usage in each gene is positively correlated to its expression level. Highly expressed genes use these 25 major codons almost exclusively. As an experimental approach to studying biased codon usage and its possible role in modulating gene expression, systematic codon replacements were carried out in the highly expressed PGK1 gene. The expression of phosphoglycerate kinase (PGK) was studied both on a high-copy-number plasmid and as a single copy gene integrated into the chromosome. Replacing an increasing number (up to 39% of all codons) of major codons with synonymous minor ones at the 5' end of the coding sequence caused a dramatic decline of the expression level. The PGK protein levels dropped 10-fold. The steady-state mRNA levels also declined, but to a lesser extent (threefold). Our data indicate that this reduction in mRNA levels was due to destabilization caused by impaired translation elongation at the minor codons. By preventing translation of the PGK mRNAs by the introduction of a stop codon 3' and adjacent to the start codon, the steady-state mRNA levels decreased dramatically. We conclude that efficient mRNA translation is required for maintaining mRNA stability in S. cerevisiae. These findings have important implications for the study of the expression of heterologous genes in yeast cells.

scholarly journals Nutritionally variant streptococci.

1991 ◽  
Vol 4 (2) ◽  
pp. 184-190 ◽  
Author(s):  
K L Ruoff
Keyword(s):  
Cell Wall ◽  
Oral Cavity ◽  
Successful Treatment ◽  
Distinct Species ◽  
Growth Conditions ◽  
Clinical Specimens ◽  
Wide Range ◽  
Streptococcal Species ◽  
A Cell

Streptococci requiring either pyridoxal or L-cysteine for growth were first observed 30 years ago as organisms forming satellite colonies adjacent to colonies of "helper" bacteria. Although they were previously considered nutritional mutants of viridans streptococcal species, the nutritionally variant streptococci (NVS) are currently thought to belong to distinct species of the genus Streptococcus. NVS strains may display pleomorphic cellular morphologies, depending on their growth conditions, and are distinguished from most other streptococci by enzymatic and serological characteristics and the presence of a cell wall chromophore. NVS are found as normal inhabitants of the oral cavity, and in addition to their participation in endocarditis, they have been isolated from a wide range of clinical specimens. Endocarditis caused by NVS is often difficult to eradicate; combinations of penicillin and an aminoglycoside are recommended for treatment. The unique physiological features of the NVS contribute to the difficulties encountered in their recovery from clinical specimens and may play a role in the problems associated with successful treatment of NVS endocarditis.

scholarly journals Physiological function of Flo11p domains and the particular role of amyloid core sequences of this adhesin in Saccharomyces cerevisiae

2021 ◽  
Author(s):  
Clara Bouyx ◽  
Marion Schiavone ◽  
Marie-Ange Teste ◽  
Etienne Dague ◽  
Nathalie Sieczkowski ◽  
...  
Keyword(s):  
Cell Wall ◽  
Amyloid Β ◽  
Surface Hydrophobicity ◽  
Yeast Cells ◽  
Invasive Growth ◽  
Specific Domain ◽  
A Genome ◽  
A Cell ◽  
Cellular Aggregates

Flocculins are a family of glycosylated proteins that provide yeast cells with several properties such as biofilm formation, flocculation, invasive growth or formation of velum. These proteins are similarly organised with a N-terminal (adhesion) domain, a stalk-like central B-domain with several repeats and a C-terminal sequence carrying a cell wall anchor site. They also contain amyloid β-aggregation-prone sequences whose functional role is still unclear. In this work, we show that Flo11p differs from other flocculins by the presence of unique amyloid-forming sequences, whose the number is critical in the formation of adhesion nanodomains under a physical shear force. Using a genome editing approach to identify the function of domains in Flo11p phenotypes, we show that the formation of cellular aggregates whose density increases with the number of amyloid sequences cannot be attributed to a specific domain of Flo11p. The same is true for plastic adhesion and surface hydrophobicity the intensity of which depends mainly on the abundance of Flo11p on the cell surface. In contrast, the N and C domains of Flo11p are essential for invasive growth in agar, whereas a reduction in the number of repeats of the B domain weakens this phenotype. However, expression of FLO11 alone is not sufficient to trigger this invasion phenotype. Finally, we show that this flocculin contributes to the integrity of the cell wall.

scholarly journals Remodeling of the Histoplasma Capsulatum Membrane Induced by Monoclonal Antibodies

Vaccines ◽  
2020 ◽  
Vol 8 (2) ◽  
pp. 269 ◽  
Author(s):  
Meagan C. Burnet ◽  
Daniel Zamith-Miranda ◽  
Heino M. Heyman ◽  
Karl K. Weitz ◽  
Erin L. Bredeweg ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Fatty Acid ◽  
Fatty Acid Metabolism ◽  
Acid Metabolism ◽  
Histoplasma Capsulatum ◽  
Cellular Responses ◽  
Mapk Signaling ◽  
Yeast Cells ◽  
Global Changes ◽  
Sterol Metabolism

Antibodies play a central role in host immunity by directly inactivating or recognizing an invading pathogen to enhance different immune responses to combat the invader. However, the cellular responses of pathogens to the presence of antibodies are not well-characterized. Here, we used different mass spectrometry techniques to study the cellular responses of the pathogenic fungus Histoplasma capsulatum to monoclonal antibodies (mAb) against HSP60, the surface protein involved in infection. A proteomic analysis of H. capsulatum yeast cells revealed that mAb binding regulates a variety of metabolic and signaling pathways, including fatty acid metabolism, sterol metabolism, MAPK signaling and ubiquitin-mediated proteolysis. The regulation of the fatty acid metabolism was accompanied by increases in the level of polyunsaturated fatty acids, which further augmented the degree of unsaturated lipids in H. capsulatum’s membranes and energy storage lipids, such as triacylglycerols, phosphatidylcholines, phosphatidylethanolamines and phosphatidylinositols. MAb treatment also regulated sterol metabolism by increasing the levels of cholesterol and ergosterol in the cells. We also showed that global changes in the lipid profiles resulted in an increased susceptibility of H. capsulatum to the ergosterol-targeting drug amphotericin B. Overall, our data showed that mAb induction of global changes in the composition of H. capsulatum membranes can potentially impact antifungal treatment during histoplasmosis.

scholarly journals Determination of the Global Pattern of Gene Expression in Yeast Cells by Intracellular Levels of Guanine Nucleotides

mBio ◽  
2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Andy Hesketh ◽  
Marta Vergnano ◽  
Stephen G. Oliver
Keyword(s):  
Gene Expression ◽  
Saccharomyces Cerevisiae ◽  
Carbon Source ◽  
Gene Transcription ◽  
Adenine Nucleotide ◽  
High Energy ◽  
Yeast Cells ◽  
Purine Nucleotides ◽  
Global Pattern ◽  
Content Type

ABSTRACT Correlations between gene transcription and the abundance of high-energy purine nucleotides in Saccharomyces cerevisiae have often been noted. However, there has been no systematic investigation of this phenomenon in the absence of confounding factors such as nutrient status and growth rate, and there is little hard evidence for a causal relationship. Whether transcription is fundamentally responsive to prevailing cellular energetic conditions via sensing of intracellular purine nucleotides, independently of specific nutrition, remains an important question. The controlled nutritional environment of chemostat culture revealed a strong correlation between ATP and GTP abundance and the transcription of genes required for growth. Short pathways for the inducible and futile consumption of ATP or GTP were engineered into S. cerevisiae, permitting analysis of the transcriptional effect of an increased demand for these nucleotides. During steady-state growth using the fermentable carbon source glucose, the futile consumption of ATP led to a decrease in intracellular ATP concentration but an increase in GTP and the guanylate energy charge (GEC). Expression of transcripts encoding proteins involved in ribosome biogenesis, and those controlled by promoters subject to SWI/SNF-dependent chromatin remodelling, was correlated with these nucleotide pool changes. Similar nucleotide abundance changes were observed using a nonfermentable carbon source, but an effect on the growth-associated transcriptional programme was absent. Induction of the GTP-cycling pathway had only marginal effects on nucleotide abundance and gene transcription. The transcriptional response of respiring cells to glucose was dampened in chemostats induced for ATP cycling, but not GTP cycling, and this was primarily associated with altered adenine nucleotide levels. IMPORTANCE This paper investigates whether, independently of the supply of any specific nutrient, gene transcription responds to the energy status of the cell by monitoring ATP and GTP levels. Short pathways for the inducible and futile consumption of ATP or GTP were engineered into the yeast Saccharomyces cerevisiae, and the effect of an increased demand for these purine nucleotides on gene transcription was analyzed. The resulting changes in transcription were most consistently associated with changes in GTP and GEC levels, although the reprogramming in gene expression during glucose repression is sensitive to adenine nucleotide levels. The results show that GTP levels play a central role in determining how genes act to respond to changes in energy supply and that any comprehensive understanding of the control of eukaryotic gene expression requires the elucidation of how changes in guanine nucleotide abundance are sensed and transduced to alter the global pattern of transcription.

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