scholarly journals Identification of SSF1, CNS1, and HCH1 as multicopy suppressors of a Saccharomyces cerevisiae Hsp90 loss-of-function mutation

1999 ◽  
Vol 96 (4) ◽  
pp. 1409-1414 ◽  
Author(s):  
D. F. Nathan ◽  
M. H. Vos ◽  
S. Lindquist
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Loss Of Function ◽  
Multicopy Suppressors

scholarly journals Identification of the bud emergence gene BEM4 and its interactions with rho-type GTPases in Saccharomyces cerevisiae.

1996 ◽  
Vol 16 (8) ◽  
pp. 4387-4395 ◽  
Author(s):  
D Mack ◽  
K Nishimura ◽  
B K Dennehey ◽  
T Arbogast ◽  
J Parkinson ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Synthetic Lethality ◽  
Cell Polarization ◽  
Growth Defect ◽  
Nucleotide Exchange ◽  
Loss Of Function ◽  
Multicopy Suppressor ◽  
Bud Emergence ◽  
Multicopy Suppressors ◽  
Multiple Copies

The Rho-type GTPase Cdc42p is required for cell polarization and bud emergence in Saccharomyces cerevisiae. To identify genes whose functions are linked to CDC42, we screened for (i) multicopy suppressors of a Ts- cdc42 mutant, (ii) mutants that require multiple copies of CDC42 for survival, and (iii) mutations that display synthetic lethality with a partial-loss-of-function allele of CDC24, which encodes a guanine nucleotide exchange factor for Cdc42p. In all three screens, we identified a new gene, BEM4. Cells from which BEM4 was deleted were inviable at 37 degrees C. These cells became unbudded, large, and round, consistent with a model in which Bem4p acts together with Cdc42p in polarity establishment and bud emergence. In some strains, the ability of CDC42 to serve as a multicopy suppressor of the Ts- growth defect of deltabem4 cells required co-overexpression of Rho1p, which is an essential Rho-type GTPase necessary for cell wall integrity. This finding suggests that Bem4p also affects Rho1p function. Bem4p displayed two-hybrid interactions with Cdc42p, Rho1p, and two of the three other known yeast Rho-type GTPases, suggesting that Bem4p can interact with multiple Rho-type GTPases. Models for the role of Bem4p include that it serves as a chaperone or modulates the interaction of these GTPases with one or more of their targets or regulators.

scholarly journals Responsiveness to exogenous cAMP of a Saccharomyces cerevisiae strain conferred by naturally occurring alleles of PDE1 and PDE2.

Genetics ◽  
1993 ◽  
Vol 135 (2) ◽  
pp. 321-326 ◽  
Author(s):  
H Mitsuzawa
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Slow Growth ◽  
Loss Of Function ◽  
Wild Type ◽  
Triple Mutant ◽  
Apparent Absence ◽  
Camp Pathway ◽  
Naturally Occurring ◽  
Elevated Activity ◽  
Growth Phenotype

Abstract The Saccharomyces cerevisiae strain P-28-24C, from which cAMP requiring mutants derived, responded to exogenously added cAMP. Upon the addition of cAMP, this strain showed phenotypes shared by mutants with elevated activity of the cAMP pathway. Genetic analysis involving serial crosses of this strain to a strain with another genetic background revealed that the responsiveness to cAMP results from naturally occurring loss-of-function alleles of PDE1 and PDE2, which encode low and high affinity cAMP phosphodiesterases, respectively. In addition, P-28-24C was found to carry a mutation conferring slow growth that lies in CYR1, which encodes adenylate cyclase, and the slow growth phenotype caused by the cyr1 mutation was suppressed by the pde2 mutation. Therefore P-28-24C is fortuitously a pde1 pde2 cyr1 triple mutant. Responsiveness to cAMP conferred by pde mutations suggests that S. cerevisiae cells are permeable to cAMP to some extent and that the apparent absence of effect of exogenously added cAMP on wild-type cells is due to immediate degradation by cAMP phosphodiesterases.

scholarly journals GSP1 and GSP2, genetic suppressors of the prp20-1 mutant in Saccharomyces cerevisiae: GTP-binding proteins involved in the maintenance of nuclear organization.

1993 ◽  
Vol 13 (4) ◽  
pp. 2152-2161 ◽  
Author(s):  
P Belhumeur ◽  
A Lee ◽  
R Tam ◽  
T DiPaolo ◽  
N Fortin ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Dna Replication ◽  
Essential Gene ◽  
Negative Control ◽  
Temperature Sensitive ◽  
Vitro Assay ◽  
Gtp Binding ◽  
Genetic Suppressors ◽  
Multicopy Suppressors

The temperature-sensitive mutation prp20-1 of Saccharomyces cerevisiae exhibits a pleiotropic phenotype associated with a general failure to maintain a proper organization of the nucleus. Its mammalian homolog, RCC1, is not only reported to be involved in the negative control of chromosome condensation but is also believed to assist in the coupling of DNA replication to the entry into mitosis. Recent studies on Xenopus RCC1 have strongly suggested a further role for this protein in the formation or maintenance of the DNA replication machinery. To elucidate the nature of the various components required for this PRP20 control pathway in S. cerevisiae, we undertook a search for multicopy suppressors of a prp20 thermosensitive mutant. Two genes, GSP1 and GSP2, were identified that encode almost identical polypeptides of 219 and 220 amino acids. Sequence analyses of these proteins show them to contain the ras consensus domains involved in GTP binding and metabolism. The levels of the GSP1 transcript are about 10-fold those of GSP2. As for S. cerevisiae RAS2, GSP2 expression exhibits carbon source dependency, while GSP1 expression does not. GSP1 is an essential gene, and GSP2 is not required for cell viability. We show that GSP1p is nuclear, that it can bind GTP in an in vitro assay, and finally, that a mutation in GSP1p which activates small ras-like proteins by increasing the stability of the GTP-bound form causes a dominant lethal phenotype. We believe that these two gene products may serve in regulating the activities of the multicomponent PRP20 complex.

scholarly journals Translational Regulation of Nuclear Gene COX4 Expression by Mitochondrial Content of Phosphatidylglycerol and Cardiolipin in Saccharomyces cerevisiae

2006 ◽  
Vol 26 (3) ◽  
pp. 743-753 ◽  
Author(s):  
Xuefeng Su ◽  
William Dowhan
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Translational Regulation ◽  
Nuclear Gene ◽  
Green Fluorescence Protein ◽  
Untranslated Regions ◽  
Loss Of Function ◽  
Green Fluorescence ◽  
Recessive Mutations ◽  
Protein Factor ◽  
Fluorescence Protein

ABSTRACT Previous results indicated that translation of four mitochondrion-encoded genes and one nucleus-encoded gene (COX4) is repressed in mutants (pgs1Δ) of Saccharomyces cerevisiae lacking phosphatidylglycerol and cardiolipin. COX4 translation was studied here using a mitochondrially targeted green fluorescence protein (mtGFP) fused to the COX4 promoter and its 5′ and 3′ untranslated regions (UTRs). Lack of mtGFP expression independent of carbon source and strain background was established to be at the translational level. The translational defect was not due to deficiency of mitochondrial respiratory function but was rather caused directly by the lack of phosphatidylglycerol and cardiolipin in mitochondrial membranes. Reintroduction of a functional PGS1 gene under control of the ADH1 promoter restored phosphatidylglycerol synthesis and expression of mtGFP. Deletion analysis of the 5′ UTR COX4 revealed the presence of a 50-nucleotide fragment with two stem-loops as a cis-element inhibiting COX4 translation. Binding of a protein factor(s) specifically to this sequence was observed with cytoplasm from pgs1Δ but not PGS1 cells. Using HIS3 and lacZ as reporters, extragenic spontaneous recessive mutations that allowed expression of His3p and β-galactosidase were isolated, which appeared to be loss-of-function mutations, suggesting that the genes mutated may encode the trans factors that bind to the cis element in pgs1Δ cells.

scholarly journals The Methionine 549 and Leucine 552 Residues of Friedelin Synthase from Maytenus ilicifolia Are Important for Substrate Binding Specificity

Molecules ◽  
2021 ◽  
Vol 26 (22) ◽  
pp. 6806
Author(s):  
Bruna F. Mazzeu ◽  
Tatiana M. Souza-Moreira ◽  
Andrew A. Oliveira ◽  
Melissa Remlinger ◽  
Lidiane G. Felippe ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Biological Activities ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis ◽  
Enzyme Specificity ◽  
Amino Acid Residues ◽  
Loss Of Function ◽  
Pentacyclic Triterpene ◽  
Oxidosqualene Cyclases ◽  
Maytenus Ilicifolia

Friedelin, a pentacyclic triterpene found in the leaves of the Celastraceae species, demonstrates numerous biological activities and is a precursor of quinonemethide triterpenes, which are promising antitumoral agents. Friedelin is biosynthesized from the cyclization of 2,3-oxidosqualene, involving a series of rearrangements to form a ketone by deprotonation of the hydroxylated intermediate, without the aid of an oxidoreductase enzyme. Mutagenesis studies among oxidosqualene cyclases (OSCs) have demonstrated the influence of amino acid residues on rearrangements during substrate cyclization: loss of catalytic activity, stabilization, rearrangement control or specificity changing. In the present study, friedelin synthase from Maytenus ilicifolia (Celastraceae) was expressed heterologously in Saccharomyces cerevisiae. Site-directed mutagenesis studies were performed by replacing phenylalanine with tryptophan at position 473 (Phe473Trp), methionine with serine at position 549 (Met549Ser) and leucine with phenylalanine at position 552 (Leu552Phe). Mutation Phe473Trp led to a total loss of function; mutants Met549Ser and Leu552Phe interfered with the enzyme specificity leading to enhanced friedelin production, in addition to α-amyrin and β-amyrin. Hence, these data showed that methionine 549 and leucine 552 are important residues for the function of this synthase.

scholarly journals Transcriptomic Changes Induced by Deletion of Transcriptional Regulator GCR2 on Pentose Sugar Metabolism in Saccharomyces cerevisiae

2021 ◽  
Vol 9 ◽  
Author(s):  
Minhye Shin ◽  
Heeyoung Park ◽  
Sooah Kim ◽  
Eun Joong Oh ◽  
Deokyeol Jeong ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Xylose Fermentation ◽  
Sugar Metabolism ◽  
Pentose Phosphate ◽  
Biofuel Production ◽  
Rna Seq ◽  
Loss Of Function ◽  
Lignocellulosic Biofuel ◽  
Pentose Sugar ◽  
Transcriptomic Changes

Being a microbial host for lignocellulosic biofuel production, Saccharomyces cerevisiae needs to be engineered to express a heterologous xylose pathway; however, it has been challenging to optimize the engineered strain for efficient and rapid fermentation of xylose. Deletion of PHO13 (Δpho13) has been reported to be a crucial genetic perturbation in improving xylose fermentation. A confirmed mechanism of the Δpho13 effect on xylose fermentation is that the Δpho13 transcriptionally activates the genes in the non-oxidative pentose phosphate pathway (PPP). In the current study, we found a couple of engineered strains, of which phenotypes were not affected by Δpho13 (Δpho13-negative), among many others we examined. Genome resequencing of the Δpho13-negative strains revealed that a loss-of-function mutation in GCR2 was responsible for the phenotype. Gcr2 is a global transcriptional factor involved in glucose metabolism. The results of RNA-seq confirmed that the deletion of GCR2 (Δgcr2) led to the upregulation of PPP genes as well as downregulation of glycolytic genes, and changes were more significant under xylose conditions than those under glucose conditions. Although there was no synergistic effect between Δpho13 and Δgcr2 in improving xylose fermentation, these results suggested that GCR2 is a novel knockout target in improving lignocellulosic ethanol production.

scholarly journals Saccharomyces cerevisiae kinesin- and dynein-related proteins required for anaphase chromosome segregation.

1995 ◽  
Vol 128 (4) ◽  
pp. 617-624 ◽  
Author(s):  
W S Saunders ◽  
D Koshland ◽  
D Eshel ◽  
I R Gibbons ◽  
M A Hoyt
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Chromosome Segregation ◽  
Motor Proteins ◽  
Cytoplasmic Dynein ◽  
Gene Products ◽  
Loss Of Function ◽  
Direct Role ◽  
Double Deletion ◽  
Different Types ◽  
Related Proteins

The Saccharomyces cerevisiae kinesin-related gene products Cin8p and Kip1p function to assemble the bipolar mitotic spindle. The cytoplasmic dynein heavy chain homologue Dyn1p (also known as Dhc1p) participates in proper cellular positioning of the spindle. In this study, the roles of these motor proteins in anaphase chromosome segregation were examined. While no single motor was essential, loss of function of all three completely halted anaphase chromatin separation. As combined motor activity was diminished by mutation, both the velocity and extent of chromatin movement were reduced, suggesting a direct role for all three motors in generating a chromosome-separating force. Redundancy for function between different types of microtubule-based motor proteins was also indicated by the observation that cin8 dyn1 double-deletion mutants are inviable. Our findings indicate that the bulk of anaphase chromosome segregation in S. cerevisiae is accomplished by the combined actions of these three motors.

scholarly journals Tolerance of DNA Replication Stress Is Promoted by Fumarate Through Modulation of Histone Demethylation and Enhancement of Replicative Intermediate Processing in Saccharomyces cerevisiae

Genetics ◽  
2019 ◽  
Vol 212 (3) ◽  
pp. 631-654 ◽  
Author(s):  
Faeze Saatchi ◽  
Ann L. Kirchmaier
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Dna Replication ◽  
Mammalian Cells ◽  
Cell Cancer ◽  
Replication Stress ◽  
Histone Demethylase ◽  
Loss Of Function ◽  
Cycle Enzyme ◽  
Link Type ◽  
Dna Replication Stress

Fumarase is a well-characterized TCA cycle enzyme that catalyzes the reversible conversion of fumarate to malate. In mammals, fumarase acts as a tumor suppressor, and loss-of-function mutations in the FH gene in hereditary leiomyomatosis and renal cell cancer result in the accumulation of intracellular fumarate—an inhibitor of α-ketoglutarate-dependent dioxygenases. Fumarase promotes DNA repair by nonhomologous end joining in mammalian cells through interaction with the histone variant H2A.Z, and inhibition of KDM2B, a H3 K36-specific histone demethylase. Here, we report that Saccharomyces cerevisiae fumarase, Fum1p, acts as a response factor during DNA replication stress, and fumarate enhances survival of yeast lacking Htz1p (H2A.Z in mammals). We observed that exposure to DNA replication stress led to upregulation as well as nuclear enrichment of Fum1p, and raising levels of fumarate in cells via deletion of FUM1 or addition of exogenous fumarate suppressed the sensitivity to DNA replication stress of htz1Δ mutants. This suppression was independent of modulating nucleotide pool levels. Rather, our results are consistent with fumarate conferring resistance to DNA replication stress in htz1Δ mutants by inhibiting the H3 K4-specific histone demethylase Jhd2p, and increasing H3 K4 methylation. Although the timing of checkpoint activation and deactivation remained largely unaffected by fumarate, sensors and mediators of the DNA replication checkpoint were required for fumarate-dependent resistance to replication stress in the htz1Δ mutants. Together, our findings imply metabolic enzymes and metabolites aid in processing replicative intermediates by affecting chromatin modification states, thereby promoting genome integrity.

scholarly journals Prohibitin Family Members Interact Genetically with Mitochondrial Inheritance Components in Saccharomyces cerevisiae

1998 ◽  
Vol 18 (7) ◽  
pp. 4043-4052 ◽  
Author(s):  
Karen H. Berger ◽  
Michael P. Yaffe
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Outer Membrane ◽  
Integral Membrane Proteins ◽  
Mitochondrial Outer Membrane ◽  
Mitochondrial Morphology ◽  
Mitochondrial Inheritance ◽  
Loss Of Function ◽  
Wild Type ◽  
Mitochondrial Inner Membrane ◽  
Synthetically Lethal

ABSTRACT Phb2p, a homolog of the tumor suppressor protein prohibitin, was identified in a genetic screen for suppressors of the loss of Mdm12p, a mitochondrial outer membrane protein required for normal mitochondrial morphology and inheritance in Saccharomyces cerevisiae. Phb2p and its homolog, prohibitin (Phb1p), were localized to the mitochondrial inner membrane and characterized as integral membrane proteins which depend on each other for their stability. In otherwise wild-type genetic backgrounds, null mutations in PHB1 andPHB2 did not confer any obvious phenotypes. However, loss of function of either PHB1 or PHB2 in cells with mitochondrial DNA deleted led to altered mitochondrial morphology, and phb1 or phb2 mutations were synthetically lethal when combined with a mutation in any of three mitochondrial inheritance components of the mitochondrial outer membrane, Mdm12p, Mdm10p, and Mmm1p. These results provide the first evidence of a role for prohibitin in mitochondrial inheritance and in the regulation of mitochondrial morphology.

scholarly journals PP2AB55δ Responsible for the High Initial Rates of Alcoholic Fermentation in Sake Yeast Strains of Saccharomyces cerevisiae

2018 ◽  
Author(s):  
Daisuke Watanabe ◽  
Takuma Kajihara ◽  
Yukiko Sugimoto ◽  
Kenichi Takagi ◽  
Megumi Mizuno ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Protein Kinase ◽  
Alcoholic Fermentation ◽  
Yeast Cells ◽  
Loss Of Function ◽  
Fermentation Performance ◽  
Fermentation Rate ◽  
Yeast Strains ◽  
Evolutionarily Conserved ◽  
Fermentation Control

ABSTRACTSake yeast strain Kyokai no. 7 (K7) and its Saccharomyces cerevisiae relatives carry a homozygous loss-of-function mutation in the RIM15 gene, which encodes a Greatwall-family protein kinase. Disruption of RIM15 in non-sake yeast strains leads to improved alcoholic fermentation, indicating that the defect in Rim15p is associated with the enhanced fermentation performance of sake yeast cells. In order to understand how Rim15p mediates fermentation control, we here focused on target-of-rapamycin protein kinase complex 1 (TORC1) and protein phosphatase 2A with the B55Δ regulatory subunit (PP2AB55δ), complexes that are known to act upstream and downstream of Rim15p, respectively. Several lines of evidence, including our previous transcriptomic analysis data, suggested enhanced TORC1 signaling in sake yeast cells during sake fermentation. Fermentation tests of the TORC1-related mutants using a laboratory strain revealed that TORC1 signaling positively regulates the initial fermentation rate in a Rim15p-dependent manner. Deletion of the CDC55 gene encoding B55δ abolished the high fermentation performance of Rim15p-deficient laboratory yeast and sake yeast cells, indicating that PP2AB55δ mediates the fermentation control by TORC1 and Rim15p. The TORC1-Greatwall-PP2AB55δ pathway similarly affected the fermentation rate in the fission yeast Schizosaccharomyces pombe, strongly suggested that the evolutionarily conserved pathway governs alcoholic fermentation in yeasts. It is likely that elevated PP2AB55δ activity accounts for the high fermentation performance of sake yeast cells. Heterozygous loss-of-function mutations in CDC55 found in K7-related sake strains may indicate that the Rim15p-deficient phenotypes are disadvantageous to cell survival.IMPORTANCEThe biochemical processes and enzymes responsible for glycolysis and alcoholic fermentation by the yeast S. cerevisiae have long been the subject of scientific research. Nevertheless, the factors determining fermentation performance in vivo are not fully understood. As a result, the industrial breeding of yeast strains has required empirical characterization of fermentation by screening numerous mutants through laborious fermentation tests. To establish a rational and efficient breeding strategy, key regulators of alcoholic fermentation need to be identified. In the present study, we focused on how sake yeast strains of S. cerevisiae have acquired high alcoholic fermentation performance. Our findings provide a rational molecular basis to design yeast strains with optimal fermentation performance for production of alcoholic beverages and bioethanol. In addition, as the evolutionarily conserved TORC1-Greatwall-PP2AB55δ pathway plays a major role in the glycolytic control, our work may contribute to research on carbohydrate metabolism in higher eukaryotes.

Sign in / Sign up

Export Citation Format

Share Document