Structural and functional conservation of human and yeast HCP1 genes which can suppress the growth defect of the Saccharomyces cerevisiae ire15 mutant

Abstract The DOM34 gene of Saccharomyces cerevisiae is similar togenes found in diverse eukaryotes and archaebacteria. Analysis of dom34 strains shows that progression through the G1 phase of the cell cycle is delayed, mutant cells enter meiosis aberrantly, and their ability to form pseudohyphae is significantly diminished. RPS30A, which encodes ribosomal protein S30, was identified in a screen for high-copy suppressors of the dom34Δ growth defect. dom34Δ mutants display an altered polyribosome profile that is rescued by expression of RPS30A. Taken together, these data indicate that Dom34p functions in protein translation to promote G1 progression and differentiation. A Drosophila homolog of Dom34p, pelota, is required for the proper coordination of meiosis and spermatogenesis. Heterologous expression of pelota in dom34Δ mutants restores wild-type growth and differentiation, suggesting conservation of function between the eukaryotic members of the gene family.

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

Molecular and Cellular Biology ◽

10.1128/mcb.16.8.4387 ◽

1996 ◽

Vol 16 (8) ◽

pp. 4387-4395 ◽

Cited By ~ 29

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.

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Subtelomeric Repeat Amplification Is Associated With Growth at Elevated Temperature in yku70 Mutants of Saccharomyces cerevisiae

Genetics ◽

10.1093/genetics/154.3.1039 ◽

2000 ◽

Vol 154 (3) ◽

pp. 1039-1051

Author(s):

Barbara Fellerhoff ◽

Friederike Eckardt-Schupp ◽

Anna A Friedl

Keyword(s):

Saccharomyces Cerevisiae ◽

Elevated Temperature ◽

Alternative Pathway ◽

Dna Amplification ◽

Strand Break ◽

Significant Loss ◽

Growth Defect ◽

Slight Reduction ◽

Telomeric Sequences ◽

Repair Defect

Abstract Inactivation of the Saccharomyces cerevisiae gene YKU70 (HDF1), which encodes one subunit of the Ku heterodimer, confers a DNA double-strand break repair defect, shortening of and structural alterations in the telomeres, and a severe growth defect at 37°. To elucidate the basis of the temperature sensitivity, we analyzed subclones derived from rare yku70 mutant cells that formed a colony when plated at elevated temperature. In all these temperature-resistant subclones, but not in cell populations shifted to 37°, we observed substantial amplification and redistribution of subtelomeric Y′ element DNA. Amplification of Y′ elements and adjacent telomeric sequences has been described as an alternative pathway for chromosome end stabilization that is used by postsenescence survivors of mutants deficient for the telomerase pathway. Our data suggest that the combination of Ku deficiency and elevated temperature induces a potentially lethal alteration of telomere structure or function. Both in yku70 mutants and in wild type, incubation at 37° results in a slight reduction of the mean length of terminal restriction fragments, but not in a significant loss of telomeric (C1-3A/TG1-3)n sequences. We propose that the absence of Ku, which is known to bind to telomeres, affects the telomeric chromatin so that its chromosome end-defining function is lost at 37°.

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Interactions of GMP with Human Glrx3 and with Saccharomyces cerevisiae Grx3 and Grx4 Converge in the Regulation of the Gcn2 Pathway

Applied and Environmental Microbiology ◽

10.1128/aem.00221-20 ◽

2020 ◽

Vol 86 (14) ◽

Author(s):

Mónica A. Mechoud ◽

Nuria Pujol-Carrion ◽

Sandra Montella-Manuel ◽

Maria Angeles de la Torre-Ruiz

Keyword(s):

Saccharomyces Cerevisiae ◽

Heterologous Expression ◽

Mammalian Cells ◽

Iron Homeostasis ◽

Life Extension ◽

Iron Sulfur Clusters ◽

Content Type ◽

Functional Conservation ◽

Iron Sulfur ◽

Cell Life

ABSTRACT The human monothiol glutaredoxin Glrx3 (PICOT) is ubiquitously distributed in cytoplasm and nuclei in mammalian cells. Its overexpression has been associated with the development of several types of tumors, whereas its deficiency might cause retardation in embryogenesis. Its exact biological role has not been well resolved, although a function as a chaperone distributing iron/sulfur clusters is currently accepted. Yeast humanization and the use of a mouse library have allowed us to find a new partner for PICOT: the human GMP synthase (hGMPs). Both proteins carry out collaborative functions regarding the downregulation of the Saccharomyces cerevisiae Gcn2 pathway under conditions of nutritional stress. Glrx3/hGMPs interact through conserved residues that bridge iron/sulfur clusters and glutathione. This mechanism is also conserved in budding yeast, whose proteins Grx3/Grx4, along with GUA1 (S. cerevisiae GMPs), also downregulate the integrated stress response (ISR) pathway. The heterologous expression of Glrx3/hGMPs efficiently complements Grx3/Grx4. Moreover, the heterologous expression of Glrx3 efficiently complements the novel participation in chronological life span that has been characterized for both Grx3 and Grx4. Our results underscore that the Glrx3/Grx3/Grx4 family presents an evolutionary and functional conservation in signaling events that is partly related to GMP function and contributes to cell life extension. IMPORTANCE Saccharomyces cerevisiae is an optimal eukaryotic microbial model to study biological processes in higher organisms despite the divergence in evolution. The molecular function of yeast glutaredoxins Grx3 and Grx4 is enormously interesting, since both proteins are required to maintain correct iron homeostasis and an efficient response to oxidative stress. The human orthologous Glrx3 (PICOT) is involved in a number of human diseases, including cancer. Our research expanded its utility to human cells. Yeast has allowed the characterization of GMP synthase as a new interacting partner for Glrx3 and also for yeast Grx3 and Grx4, the complex monothiol glutaredoxins/GMPs that participate in the downregulation of the activity of the Gcn2 stress pathway. This mechanism is conserved in yeast and humans. Here, we also show that this family of glutaredoxins, Grx3/Grx4/Glrx3, also has a function related to life extension.

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Overlapping function of Hrd1 and Ste24 in translocon quality control provides robust channel surveillance

Journal of Biological Chemistry ◽

10.1074/jbc.ac120.016191 ◽

2020 ◽

Vol 295 (47) ◽

pp. 16113-16120

Author(s):

Avery M. Runnebohm ◽

Kyle A. Richards ◽

Courtney Broshar Irelan ◽

Samantha M. Turk ◽

Halie E. Vitali ◽

...

Keyword(s):

Saccharomyces Cerevisiae ◽

Quality Control ◽

Endoplasmic Reticulum ◽

Ubiquitin Ligase ◽

Biological Membranes ◽

The Other ◽

Growth Defect ◽

Zinc Metalloprotease

Translocation of proteins across biological membranes is essential for life. Proteins that clog the endoplasmic reticulum (ER) translocon prevent the movement of other proteins into the ER. Eukaryotes have multiple translocon quality control (TQC) mechanisms to detect and destroy proteins that persistently engage the translocon. TQC mechanisms have been defined using a limited panel of substrates that aberrantly occupy the channel. The extent of substrate overlap among TQC pathways is unknown. In this study, we found that two TQC enzymes, the ER-associated degradation ubiquitin ligase Hrd1 and zinc metalloprotease Ste24, promote degradation of characterized translocon-associated substrates of the other enzyme in Saccharomyces cerevisiae. Although both enzymes contribute to substrate turnover, our results suggest a prominent role for Hrd1 in TQC. Yeast lacking both Hrd1 and Ste24 exhibit a profound growth defect, consistent with overlapping function. Remarkably, two mutations that mildly perturb post-translational translocation and reduce the extent of aberrant translocon engagement by a model substrate diminish cellular dependence on TQC enzymes. Our data reveal previously unappreciated mechanistic complexity in TQC substrate detection and suggest that a robust translocon surveillance infrastructure maintains functional and efficient translocation machinery.

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Cns1 Is an Essential Protein Associated with the Hsp90 Chaperone Complex in Saccharomyces cerevisiae That Can Restore Cyclophilin 40-Dependent Functions in cpr7ΔCells

Molecular and Cellular Biology ◽

10.1128/mcb.18.12.7353 ◽

1998 ◽

Vol 18 (12) ◽

pp. 7353-7359 ◽

Cited By ~ 63

Author(s):

James A. Marsh ◽

Helen M. Kalton ◽

Richard F. Gaber

Keyword(s):

Saccharomyces Cerevisiae ◽

Molecular Chaperone ◽

Single Gene ◽

Normal Growth ◽

Maximal Activity ◽

Negative Regulation ◽

Growth Defect ◽

Chaperone Complex ◽

Cyclophilin 40

ABSTRACT Saccharomyces cerevisiae harbors two cyclophilin 40-type enzymes, Cpr6 and Cpr7, which are components of the Hsp90 molecular chaperone machinery. Cpr7 is required for normal growth and is required for maximal activity of heterologous Hsp90-dependent substrates, including glucocorticoid receptor (GR) and the oncogenic tyrosine kinase pp60v-src . In addition, it has recently been shown that Cpr7 plays a major role in negative regulation of the S. cerevisiae heat shock transcription factor (HSF). To better understand functions associated with Cpr7, a search was undertaken for multicopy suppressors of the cpr7Δ slow-growth phenotype. The screen identified a single gene, designatedCNS1 (for cyclophilin seven suppressor), capable of suppressing the cpr7Δ growth defect. Overexpression ofCNS1 in cpr7Δ cells also largely restored GR activity and negative regulation of HSF. In vitro protein retention experiments in which Hsp90 heterocomplexes were precipitated resulted in coprecipitation of Cns1. Interaction between Cns1 and the carboxy terminus of Hsp90 was also shown by two-hybrid analysis. The functional consequences of CNS1 overexpression and its physical association with the Hsp90 machinery indicate that Cns1 is a previously unidentified component of molecular chaperone complexes. Thus far, Cns1 is the only tetratricopeptide repeat-containing component of Hsp90 heterocomplexes found to be essential for cell viability under all conditions tested.

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Manganese Suppresses the Haploinsufficiency of Heterozygous trpy1Δ/TRPY1 Saccharomyces cerevisiae Cells and Stimulates the TRPY1-Dependent Release of Vacuolar Ca2+ under H2O2 Stress

Cells ◽

10.3390/cells8020079 ◽

2019 ◽

Vol 8 (2) ◽

pp. 79 ◽

Cited By ~ 4

Author(s):

Lavinia Ruta ◽

Ioana Nicolau ◽

Claudia Popa ◽

Ileana Farcasanu

Keyword(s):

Oxidative Stress ◽

Saccharomyces Cerevisiae ◽

Cellular Response ◽

Trp Channels ◽

Homozygous Deletion ◽

Growth Defect ◽

Mechanical Stimuli ◽

Hyperosmotic Shock ◽

Diploid Cells ◽

In Cells

Transient potential receptor (TRP) channels are conserved cation channels found in most eukaryotes, known to sense a variety of chemical, thermal or mechanical stimuli. The Saccharomyces cerevisiae TRPY1 is a TRP channel with vacuolar localization involved in the cellular response to hyperosmotic shock and oxidative stress. In this study, we found that S. cerevisiae diploid cells with heterozygous deletion in TRPY1 gene are haploinsufficient when grown in synthetic media deficient in essential metal ions and that this growth defect is alleviated by non-toxic Mn2+ surplus. Using cells expressing the Ca2+-sensitive photoprotein aequorin we found that Mn2+ augmented the Ca2+ flux into the cytosol under oxidative stress, but not under hyperosmotic shock, a trait that was absent in the diploid cells with homozygous deletion of TRPY1 gene. TRPY1 activation under oxidative stress was diminished in cells devoid of Smf1 (the Mn2+-high-affinity plasma membrane transporter) but it was clearly augmented in cells lacking Pmr1 (the endoplasmic reticulum (ER)/Golgi located ATPase responsible for Mn2+ detoxification via excretory pathway). Taken together, these observations lead to the conclusion that increased levels of intracytosolic Mn2+ activate TRPY1 in the response to oxidative stress.

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PTA1, an essential gene of Saccharomyces cerevisiae affecting pre-tRNA processing.

Molecular and Cellular Biology ◽

10.1128/mcb.12.9.3843 ◽

1992 ◽

Vol 12 (9) ◽

pp. 3843-3856 ◽

Cited By ~ 20

Author(s):

J P O'Connor ◽

C L Peebles

Keyword(s):

Saccharomyces Cerevisiae ◽

Gene Product ◽

Genomic Library ◽

Essential Gene ◽

Growth Defect ◽

Temperature Sensitive ◽

Putative Open Reading Frame ◽

Reading Frame ◽

Trna Processing ◽

Chromosome I

We have identified an essential Saccharomyces cerevisiae gene, PTA1, that affects pre-tRNA processing. PTA1 was initially defined by a UV-induced mutation, pta1-1, that causes the accumulation of all 10 end-trimmed, intron-containing pre-tRNAs and temperature-sensitive but osmotic-remedial growth. pta1-1 does not appear to be an allele of any other known gene affecting pre-tRNA processing. Extracts prepared from pta1-1 strains had normal pre-tRNA splicing endonuclease activity. pta1-1 was suppressed by the ochre suppressor tRNA gene SUP11, indicating that the pta1-1 mutation creates a termination codon within a protein reading frame. The PTA1 gene was isolated from a genomic library by complementation of the pta1-1 growth defect. Episome-borne PTA1 directs recombination to the pta1-1 locus. PTA1 has been mapped to the left arm of chromosome I near CDC24; the gene was sequenced and could encode a protein of 785 amino acids with a molecular weight of 88,417. No other protein sequences similar to that of the predicted PTA1 gene product have been identified within the EMBL or GenBank data base. Disruption of PTA1 near the carboxy terminus of the putative open reading frame was lethal. Possible functions of the PTA1 gene product are discussed.

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Giant peroxisomes in oleic acid-induced Saccharomyces cerevisiae lacking the peroxisomal membrane protein Pmp27p.

The Journal of Cell Biology ◽

10.1083/jcb.128.4.509 ◽

1995 ◽

Vol 128 (4) ◽

pp. 509-523 ◽

Cited By ~ 201

Author(s):

R Erdmann ◽

G Blobel

Keyword(s):

Saccharomyces Cerevisiae ◽

Oleic Acid ◽

Growth Defect ◽

Wild Type ◽

Peroxisomal Membrane ◽

Daughter Cells ◽

Sds Page ◽

Multiple Buds ◽

Media Form ◽

Tubular Membranes

We have purified peroxisomal membranes from Saccharomyces cerevisiae after induction of peroxisomes in oleic acid-containing media. About 30 distinct proteins could be discerned among the HPLC- and SDS-PAGE-separated proteins of the high salt-extracted peroxisomal membranes. The most abundant of these, Pmp27p, was purified and the corresponding gene PMP27 was cloned and sequenced. Its primary structure is 32% identical to PMP31 and PMP32 of the yeast Candida biodinii (Moreno, M., R. Lark, K. L. Campbell, and M. J. Goodman. 1994. Yeast. 10:1447-1457). Immunoelectron microscopic localization of Pmp27p showed labeling of the peroxisomal membrane, but also of matrix-less and matrix containing tubular membranes nearby. Electronmicroscopical data suggest that some of these tubular extensions might interconnect peroxisomes to form a peroxisomal reticulum. Cells with a disrupted PMP27 gene (delta pmp27) still grew well on glucose or ethanol, but they failed to grow on oleate although peroxisomes were still induced by transfer to oleate-containing media. The induced peroxisomes of delta pmp27 cells were fewer but considerably larger than those of wild-type cells, suggesting that Pmp27p may be involved in parceling of peroxisomes into regular quanta. delta pmp27 cells cultured in oleate-containing media form multiple buds, of which virtually all are peroxisome deficient. The growth defect of delta pmp27 cells on oleic acid appears to result from the inability to segregate the giant peroxisomes to daughter cells.

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SYM1 Is the Stress-Induced Saccharomyces cerevisiae Ortholog of the Mammalian Kidney Disease Gene Mpv17 and Is Required for Ethanol Metabolism and Tolerance during Heat Shock

Eukaryotic Cell ◽

10.1128/ec.3.3.620-631.2004 ◽

2004 ◽

Vol 3 (3) ◽

pp. 620-631 ◽

Cited By ~ 35

Author(s):

Amy Trott ◽

Kevin A. Morano

Keyword(s):

Saccharomyces Cerevisiae ◽

Heat Shock ◽

Kidney Disease ◽

Membrane Protein ◽

Cellular Response ◽

Ethanol Metabolism ◽

Growth Defect ◽

Inner Mitochondrial Membrane ◽

Peroxisomal Membrane ◽

Metabolic Role

ABSTRACT Organisms rapidly adapt to severe environmental stress by inducing the expression of a wide array of heat shock proteins as part of a larger cellular response program. We have used a genomics approach to identify novel heat shock-induced genes in Saccharomyces cerevisiae. The uncharacterized open reading frame (ORF) YLR251W was found to be required for both metabolism and tolerance of ethanol during heat shock. YLR251W has significant homology to the mammalian peroxisomal membrane protein Mpv17, and Mpv17−/− mice exhibit age-onset glomerulosclerosis, deafness, hypertension, and, ultimately, death by renal failure. Expression of Mpv17 in ylr251wΔ cells complements the 37°C ethanol growth defect, suggesting that these proteins are functional orthologs. We have therefore renamed ORF YLR251W as SYM1 (for “stress-inducible yeast Mpv17”). In contrast to the peroxisomal localization of Mpv17, we find that Sym1 is an integral membrane protein of the inner mitochondrial membrane. In addition, transcriptional profiling of sym1Δ cells uncovered changes in gene expression, including dysregulation of a number of ethanol-repressed genes, exclusively at 37°C relative to wild-type results. Together, these data suggest an important metabolic role for Sym1 in mitochondrial function during heat shock. Furthermore, this study establishes Sym1 as a potential model for understanding the role of Mpv17 in kidney disease and cardiovascular biology.

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