scholarly journals Induction of protein aggregation and starvation response by tRNA modification defects

2020 ◽  
Vol 66 (6) ◽  
pp. 1053-1057
Author(s):  
Roland Klassen ◽  
Alexander Bruch ◽  
Raffael Schaffrath
Keyword(s):  
Cellular Response ◽  
Anticodon Loop ◽  
Protein Aggregate ◽  
Starvation Response ◽  
Codon Recognition ◽  
Growth Defects ◽  
Yeast Strains ◽  
Transcriptional Changes ◽  
Decoding Efficiency ◽  
Synthetic Growth

Abstract Posttranscriptional modifications of anticodon loops contribute to the decoding efficiency of tRNAs by supporting codon recognition and loop stability. Consistently, strong synthetic growth defects are observed in yeast strains simultaneously lacking distinct anticodon loop modifications. These phenotypes are accompanied by translational inefficiency of certain mRNAs and disturbed protein homeostasis resulting in accumulation of protein aggregates. Different combinations of anticodon loop modification defects were shown to affect distinct tRNAs but provoke common transcriptional changes that are reminiscent of the cellular response to nutrient starvation. Multiple mechanisms may be involved in mediating inadequate starvation response upon loss of critical tRNA modifications. Recent evidence suggests protein aggregate induction to represent one such trigger.

scholarly journals Deletion of Autophagy gene ATG1 and Glyoxylate cycle Regulator encoding gene UCC1 together leads to Synthetic Growth Defects and Sensitivity to Genotoxic agents in Saccharomyces cerevisiae

2020 ◽  
Author(s):  
Monika Pandita ◽  
Vishali Sharma ◽  
Meenu Sharma ◽  
Heena Shoket ◽  
Sadia Parvez ◽  
...  
Keyword(s):  
Cellular Response ◽  
Genetic Interaction ◽  
Glyoxylate Cycle ◽  
Genotoxic Stress ◽  
Ubiquitin Proteasome System ◽  
Growth Defects ◽  
Ubiquitin Proteasome ◽  
Encoding Gene ◽  
And Function ◽  
Synthetic Growth

AbstractAtg1 of S. cerevisiae is a key component of autophagy encoded by ATG1 gene, involved in the process of degradation of cytosolic components through autophagy. UCC1, an F-box encoding gene is involved in the negative regulation of glyoxylate pathway via degradation of Cit2 enzyme by ubiquitin proteasome system. We investigated the genetic interaction between ATG1 and UCC1 using the gene deletion approach. The atg1Δucc1Δ cells showed the synthetic growth defects with abnormal budding and sensitivity to genotoxic and oxidative stress agents. Based on the observations, we report that ATG1 and UCC1 interact genetically to regulate the cell growth fitness and function in parallel pathway in cellular response to the genotoxic stress agents. The present investigation also revealed the cross talks among autophagy, ubiquitin proteasome system, and glyoxylate pathways.

scholarly journals Systematic Humanization of the Yeast Cytoskeleton Discerns Functionally Replaceable from Divergent Human Genes

Genetics ◽  
2020 ◽  
Vol 215 (4) ◽  
pp. 1153-1169 ◽  
Author(s):  
Riddhiman K. Garge ◽  
Jon M. Laurent ◽  
Aashiq H. Kachroo ◽  
Edward M. Marcotte
Keyword(s):  
Gene Families ◽  
Growth Defect ◽  
Gene Duplications ◽  
Yeast Gene ◽  
Macromolecular Transport ◽  
Cellular Processes ◽  
Growth Defects ◽  
Yeast Strains ◽  
Human Genes ◽  
Interaction Partners

Many gene families have been expanded by gene duplications along the human lineage, relative to ancestral opisthokonts, but the extent to which the duplicated genes function similarly is understudied. Here, we focused on structural cytoskeletal genes involved in critical cellular processes, including chromosome segregation, macromolecular transport, and cell shape maintenance. To determine functional redundancy and divergence of duplicated human genes, we systematically humanized the yeast actin, myosin, tubulin, and septin genes, testing ∼81% of human cytoskeletal genes across seven gene families for their ability to complement a growth defect induced by inactivation or deletion of the corresponding yeast ortholog. In five of seven families—all but α-tubulin and light myosin, we found at least one human gene capable of complementing loss of the yeast gene. Despite rescuing growth defects, we observed differential abilities of human genes to rescue cell morphology, meiosis, and mating defects. By comparing phenotypes of humanized strains with deletion phenotypes of their interaction partners, we identify instances of human genes in the actin and septin families capable of carrying out essential functions, but failing to fully complement the cytoskeletal roles of their yeast orthologs, thus leading to abnormal cell morphologies. Overall, we show that duplicated human cytoskeletal genes appear to have diverged such that only a few human genes within each family are capable of replacing the essential roles of their yeast orthologs. The resulting yeast strains with humanized cytoskeletal components now provide surrogate platforms to characterize human genes in simplified eukaryotic contexts.

scholarly journals Genetic Interactions WithCLF1Identify Additional Pre-mRNA Splicing Factors and a Link Between Activators of Yeast Vesicular Transport and Splicing

Genetics ◽  
2003 ◽  
Vol 164 (3) ◽  
pp. 895-907 ◽  
Author(s):  
Kevin Vincent ◽  
Qiang Wang ◽  
Steven Jay ◽  
Kathryn Hobbs ◽  
Brian C Rymond
Keyword(s):  
Mrna Splicing ◽  
Splicing Factors ◽  
Sequence Information ◽  
Specific Sequence ◽  
Synthetic Lethal ◽  
Growth Defects ◽  
Mrna Maturation ◽  
Assembly Factor ◽  
Gtpase Activation ◽  
Synthetic Growth

AbstractClf1 is a conserved spliceosome assembly factor composed predominately of TPR repeats. Here we show that the TPR elements are not functionally equivalent, with the amino terminus of Clf1 being especially sensitive to change. Deletion and add-back experiments reveal that the splicing defect associated with TPR removal results from the loss of TPR-specific sequence information. Twelve mutants were found that show synthetic growth defects when combined with an allele that lacks TPR2 (i.e., clf1Δ2). The identified genes encode the Mud2, Ntc20, Prp16, Prp17, Prp19, Prp22, and Syf2 splicing factors and four proteins without established contribution to splicing (Bud13, Cet1, Cwc2, and Rds3). Each synthetic lethal with clf1Δ2 (slc) mutant is splicing defective in a wild-type CLF1 background. In addition to the splicing factors, SSD1, BTS1, and BET4 were identified as dosage suppressors of clf1Δ2 or selected slc mutants. These results support Clf1 function through multiple stages of the spliceosome cycle, identify additional genes that promote cellular mRNA maturation, and reveal a link between Rab/Ras GTPase activation and the process of pre-mRNA splicing.

scholarly journals Yeasts as Complementary Model Systems for the Study of the Pathological Repercussions of Enhanced Synphilin-1 Glycation and Oxidation

2021 ◽  
Vol 22 (4) ◽  
pp. 1677
Author(s):  
David Seynnaeve ◽  
Daniel P. Mulvihill ◽  
Joris Winderickx ◽  
Vanessa Franssens
Keyword(s):  
Lewy Bodies ◽  
Inhibitory Effect ◽  
Model Systems ◽  
Wild Type ◽  
Glyoxalase System ◽  
Inclusion Formation ◽  
Growth Defects ◽  
Yeast Strains ◽  
Autophagic Degradation ◽  
Detrimental Impact

Synphilin-1 has previously been identified as an interaction partner of α-Synuclein (αSyn), a primary constituent of neurodegenerative disease-linked Lewy bodies. In this study, the repercussions of a disrupted glyoxalase system and aldose reductase function on Synphilin-1 inclusion formation characteristics and cell growth were investigated. To this end, either fluorescent dsRed-tagged or non-tagged human SNCAIP, which encodes the Synphilin-1 protein, was expressed in Saccharomyces cerevisiae and Schizosaccharomyces pombe yeast strains devoid of enzymes Glo1, Glo2, and Gre3. Presented data shows that lack of Glo2 and Gre3 activity in S. cerevisiae increases the formation of large Synphilin-1 inclusions. This correlates with enhanced oxidative stress levels and an inhibitory effect on exponential growth, which is most likely caused by deregulation of autophagic degradation capacity, due to excessive Synphilin-1 aggresome build-up. These findings illustrate the detrimental impact of increased oxidation and glycation on Synphilin-1 inclusion formation. Similarly, polar-localised inclusions were observed in wild-type S. pombe cells and strains deleted for either glo1+ or glo2+. Contrary to S. cerevisiae, however, no growth defects were observed upon expression of SNCAIP. Altogether, our findings show the relevance of yeasts, especially S. cerevisiae, as complementary models to unravel mechanisms contributing to Synphilin-1 pathology in the context of neurodegenerative diseases.

scholarly journals Ups1p and Ups2p antagonistically regulate cardiolipin metabolism in mitochondria

2009 ◽  
Vol 185 (6) ◽  
pp. 1029-1045 ◽  
Author(s):  
Yasushi Tamura ◽  
Toshiya Endo ◽  
Miho Iijima ◽  
Hiromi Sesaki
Keyword(s):  
Fatty Acid ◽  
Molecular Mechanism ◽  
Protein Import ◽  
Protein Complexes ◽  
Mitochondrial Protein ◽  
Intermembrane Space ◽  
Mitochondrial Protein Import ◽  
Growth Defects ◽  
Mitochondrial Inner Membrane ◽  
Synthetic Growth

Cardiolipin, a unique phospholipid composed of four fatty acid chains, is located mainly in the mitochondrial inner membrane (IM). Cardiolipin is required for the integrity of several protein complexes in the IM, including the TIM23 translocase, a dynamic complex which mediates protein import into the mitochondria through interactions with the import motor presequence translocase–associated motor (PAM). In this study, we report that two homologous intermembrane space proteins, Ups1p and Ups2p, control cardiolipin metabolism and affect the assembly state of TIM23 and its association with PAM in an opposing manner. In ups1Δ mitochondria, cardiolipin levels were decreased, and the TIM23 translocase showed altered conformation and decreased association with PAM, leading to defects in mitochondrial protein import. Strikingly, loss of Ups2p restored normal cardiolipin levels and rescued TIM23 defects in ups1Δ mitochondria. Furthermore, we observed synthetic growth defects in ups mutants in combination with loss of Pam17p, which controls the integrity of PAM. Our findings provide a novel molecular mechanism for the regulation of cardiolipin metabolism.

scholarly journals Systematic humanization of the yeast cytoskeleton discerns functionally replaceable from divergent human genes

2019 ◽  
Author(s):  
Riddhiman K. Garge ◽  
Jon M. Laurent ◽  
Aashiq H. Kachroo ◽  
Edward M. Marcotte
Keyword(s):  
Gene Families ◽  
Growth Defect ◽  
Gene Duplications ◽  
Yeast Gene ◽  
Macromolecular Transport ◽  
Cellular Processes ◽  
Growth Defects ◽  
Yeast Strains ◽  
Human Genes ◽  
Interaction Partners

AbstractMany gene families have been expanded by gene duplications along the human lineage, relative to ancestral opisthokonts, but the extent to which the duplicated genes function similarly is understudied. Here, we focused on structural cytoskeletal genes involved in critical cellular processes including chromosome segregation, macromolecular transport, and cell shape maintenance. To determine functional redundancy and divergence of duplicated human genes, we systematically humanized the yeast actin, myosin, tubulin, and septin genes, testing ∼85% of human cytoskeletal genes across 7 gene families for their ability to complement a growth defect induced by deletion of the corresponding yeast ortholog. In 5 of 7 families—all but α-tubulin and light myosin, we found at least one human gene capable of complementing loss of the yeast gene. Despite rescuing growth defects, we observed differential abilities of human genes to rescue cell morphology, meiosis, and mating defects. By comparing phenotypes of humanized strains with deletion phenotypes of their interaction partners, we identify instances of human genes in the actin and septin families capable of carrying out essential functions, but apparently failing to interact with components of the yeast cytoskeleton, thus leading to abnormal cell morphologies. Overall, we show that duplicated human cytoskeletal genes appear to have diverged such that only a few human genes within each family are capable of replacing the essential roles of their yeast orthologs. The resulting yeast strains with humanized cytoskeletal components now provide surrogate platforms to characterize human genes in simplified eukaryotic contexts.

scholarly journals Modifications Modulate Anticodon Loop Dynamics and Codon Recognition in E. coli tRNA Arg1,2

2015 ◽  
Vol 29 (S1) ◽  
Author(s):  
Kathryn Sarachan ◽  
William Cantara ◽  
Jessica Spears ◽  
Hasan DeMirci ◽  
Frank Murphy ◽  
...  
Keyword(s):  
Anticodon Loop ◽  
E Coli ◽  
Codon Recognition ◽  
Loop Dynamics

scholarly journals Sky1 regulates the expression of sulfur metabolism genes in response to cisplatin

Microbiology ◽  
2014 ◽  
Vol 160 (7) ◽  
pp. 1357-1368 ◽  
Author(s):  
Silvia Rodríguez-Lombardero ◽  
Ángel Vizoso-Vázquez ◽  
Luis J. Lombardía ◽  
Manuel Becerra ◽  
M. Isabel González-Siso ◽  
...  
Keyword(s):  
Amino Acids ◽  
Cellular Response ◽  
Dna Microarrays ◽  
Sulfur Metabolism ◽  
Yeast Cells ◽  
Genes Encoding ◽  
Cisplatin Sensitivity ◽  
Transcriptional Changes ◽  
Upregulated Genes ◽  
Sulfur Containing

Cisplatin is commonly used in cancer therapy and yeast cells are also sensitive to this compound. We present a transcriptome analysis discriminating between RNA changes induced by cisplatin treatment, which are dependent on or independent of SKY1 function – a gene whose deletion increases resistance to the drug. Gene expression changes produced by addition of cisplatin to W303 and W303-Δsky1 cells were recorded using DNA microarrays. The data, validated by quantitative PCR, revealed 122 differentially expressed genes: 69 upregulated and 53 downregulated. Among the upregulated genes, those related to sulfur metabolism were over-represented and partially dependent on Sky1. Deletions of MET4 or other genes encoding co-regulators of the expression of sulfur-metabolism-related genes, with the exception of MET28, did not modify the cisplatin sensitivity of yeast cells. One of the genes with the highest cisplatin-induced upregulation was SEO1, encoding a putative permease of sulfur compounds. We also measured the platinum, sulfur and glutathione content in W303, W303-Δsky1 and W303-Δseo1 cells after cisplatin treatment, and integration of the data suggested that these transcriptional changes might represent a cellular response that allowed chelation of cisplatin with sulfur-containing amino acids and also helped DNA repair by stimulating purine biosynthesis. The transcription pattern of stimulation of sulfur-containing amino acids and purine synthesis decreased, or even disappeared, in the W303-Δsky1 strain.

scholarly journals Synthetic Lethal Screens Identify Gene Silencing Processes in Yeast and Implicate the Acetylated Amino Terminus of Sir3 in Recognition of the Nucleosome Core

2008 ◽  
Vol 28 (11) ◽  
pp. 3861-3872 ◽  
Author(s):  
Tibor van Welsem ◽  
Floor Frederiks ◽  
Kitty F. Verzijlbergen ◽  
Alex W. Faber ◽  
Zara W. Nelson ◽  
...  
Keyword(s):  
Nonspecific Binding ◽  
Synthetic Lethal ◽  
Nucleosome Core ◽  
Growth Defects ◽  
Sir Proteins ◽  
Epistasis Analysis ◽  
Genome Wide ◽  
Bah Domain ◽  
H3k79 Methylation ◽  
Synthetic Growth

ABSTRACT Dot1 methylates histone H3 lysine 79 (H3K79) on the nucleosome core and is involved in Sir protein-mediated silencing. Previous studies suggested that H3K79 methylation within euchromatin prevents nonspecific binding of the Sir proteins, which in turn facilitates binding of the Sir proteins in unmethylated silent chromatin. However, the mechanism by which the Sir protein binding is influenced by this modification is unclear. We performed genome-wide synthetic genetic array (SGA) analysis and identified interactions of DOT1 with SIR1 and POL32. The synthetic growth defects found by SGA analysis were attributed to the loss of mating type identity caused by a synthetic silencing defect. By using epistasis analysis, DOT1, SIR1, and POL32 could be placed in different pathways of silencing. Dot1 shared its silencing phenotypes with the NatA N-terminal acetyltransferase complex and the conserved N-terminal bromo adjacent homology (BAH) domain of Sir3 (a substrate of NatA). We classified all of these as affecting a common silencing process, and we show that mutations in this process lead to nonspecific binding of Sir3 to chromatin. Our results suggest that the BAH domain of Sir3 binds to histone H3K79 and that acetylation of the BAH domain is required for the binding specificity of Sir3 for nucleosomes unmethylated at H3K79.

scholarly journals Different Genetic Functions for the Rpd3(L) and Rpd3(S) Complexes Suggest Competition between NuA4 and Rpd3(S)

2008 ◽  
Vol 28 (14) ◽  
pp. 4445-4458 ◽  
Author(s):  
Debabrata Biswas ◽  
Shinya Takahata ◽  
David J. Stillman
Keyword(s):  
Transcriptional Activation ◽  
Chromatin Immunoprecipitation ◽  
Histone Acetyltransferase ◽  
Strand Break ◽  
Triple Mutant ◽  
Growth Defects ◽  
Mutant Phenotypes ◽  
Synthetic Growth ◽  
Synthetic Phenotype

ABSTRACT Rpd3(L) and Rpd3(S) are distinct multisubunit complexes containing the Rpd3 histone deacetylase. Disruption of the GCN5 histone acetyltransferase gene shows a strong synthetic phenotype when combined with either an sds3 mutation affecting only the Rpd3(L) complex or an rco1 mutation affecting only Rpd3(S). However, these synthetic growth defects are not seen in a gcn5 sds3 rco1 triple mutant, suggesting that the balance between Rpd3(L) and Rpd3(S) is critical in cells lacking Gcn5. Different genetic interactions are seen with mutations affecting the FACT chromatin reorganizing complex. An sds3 mutation affecting only Rpd3(L) has a synthetic defect with FACT mutants, while rco1 and eaf3 mutations affecting Rpd3(S) suppress FACT mutant phenotypes. Rpd3(L) therefore acts in concert with FACT, but Rpd3(S) opposes it. Combining FACT mutations with mutations in the Esa1 subunit of the NuA4 histone acetyltransferase results in synthetic growth defects, and these can be suppressed by an rco1 or set2 mutation. An rco1 mutation suppresses phenotypes caused by mutations in the ESA1 and ARP4 subunits of NuA4, while Rco1 overexpression exacerbates these defects. These results suggest a model in which NuA4 and Rpd3(S) compete. Chromatin immunoprecipitation experiments show that eliminating Rpd3(S) increases the amount of NuA4 binding to the ARG3 promoter during transcriptional activation and to the sites of DNA repair induced by a double-strand break. Our results suggest that the Rpd3(L) and Rpd3(S) complexes have distinct functions in vivo and that the relative amounts of the two forms alter the effectiveness of other chromatin-altering complexes, such as FACT and NuA4.

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