Cdc1 and the Vacuole Coordinately Regulate Mn2+ Homeostasis in the Yeast Saccharomyces cerevisiae

Genetics ◽  
1998 ◽  
Vol 148 (4) ◽  
pp. 1787-1798 ◽  
Author(s):  
Madan Paidhungat ◽  
Stephen Garrett
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Membrane Protein ◽  
Double Mutant ◽  
Vacuolar Membrane ◽  
Deletion Strain ◽  
Growth Defect ◽  
Yeast Saccharomyces Cerevisiae ◽  
Suppressor Genes ◽  
Essential Protein ◽  
Essential Function

Abstract The yeast CDC1 gene encodes an essential protein that has been implicated in the regulation of cytosolic [Mn2+]. To identify factors that impinge upon Cdc1 or the Cdc1-dependent process, we isolated secondsite suppressors of the conditional cdc1-1(Ts) growth defect. Recessive suppressors define 15 COS (CdcOne Suppressor) genes. Seven of the fifteen COS genes are required for biogenesis of the vacuole, an organelle known to sequester intracellular Mn2+. An eighth gene, COS16, encodes a vacuolar membrane protein that seems to be involved in Mn2+ homeostasis. These results suggest mutations that block vacuolar Mn2+ sequestration compensate for defects in Cdc1 function. Interestingly, Cdc1 is dispensable in a cos16Δ deletion strain, and a cdc1Δ cos16Δ double mutant exhibits robust growth on medium supplemented with Mn2+. Thus, the single, essential function of Cdc1 is to regulate intracellular, probably cytosolic, Mn2+.

scholarly journals 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

2004 ◽  
Vol 3 (3) ◽  
pp. 620-631 ◽  
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.

scholarly journals Cell surface recycling in yeast: mechanisms and machineries

2016 ◽  
Vol 44 (2) ◽  
pp. 474-478 ◽  
Author(s):  
Chris MacDonald ◽  
Robert C. Piper
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Plasma Membrane ◽  
Membrane Protein ◽  
Cell Surface ◽  
Mammalian Cells ◽  
Genetic System ◽  
Integral Membrane Protein ◽  
Protein Activity ◽  
Yeast Saccharomyces Cerevisiae ◽  
Central Process

Sorting internalized proteins and lipids back to the cell surface controls the supply of molecules throughout the cell and regulates integral membrane protein activity at the surface. One central process in mammalian cells is the transit of cargo from endosomes back to the plasma membrane (PM) directly, along a route that bypasses retrograde movement to the Golgi. Despite recognition of this pathway for decades we are only beginning to understand the machinery controlling this overall process. The budding yeast Saccharomyces cerevisiae, a stalwart genetic system, has been routinely used to identify fundamental proteins and their modes of action in conserved trafficking pathways. However, the study of cell surface recycling from endosomes in yeast is hampered by difficulties that obscure visualization of the pathway. Here we briefly discuss how recycling is likely a more prevalent process in yeast than is widely appreciated and how tools might be built to better study the pathway.

scholarly journals Interaction among Btn1p, Btn2p, and Ist2p Reveals Potential Interplay among the Vacuole, Amino Acid Levels, and Ion Homeostasis in the Yeast Saccharomyces cerevisiae

2005 ◽  
Vol 4 (2) ◽  
pp. 281-288 ◽  
Author(s):  
Yoojin Kim ◽  
Subrata Chattopadhyay ◽  
Sarahjane Locke ◽  
David A. Pearce
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Amino Acid ◽  
Membrane Protein ◽  
Optimal Transport ◽  
Ion Homeostasis ◽  
Coiled Coil ◽  
Batten Disease ◽  
Yeast Saccharomyces Cerevisiae ◽  
Arginine Transport ◽  
Amino Acid Levels

ABSTRACT Btn2p, a novel cytosolic coiled-coil protein in Saccharomyces cerevisiae, was previously shown to interact with and to be necessary for the correct localization of Rhb1p, a regulator of arginine uptake, and Yif1p, a Golgi protein. We now report the biochemical and physical interactions of Btn2p with Ist2p, a plasma membrane protein that is thought to have a function in salt tolerance. A deletion in Btn2p (btn2Δ strains) results in a failure to correctly localize Ist2p, and strains lacking Btn2p and Ist2p (btn2Δ ist2Δ strains) are unable to grow in the presence of 0.5 or 1.0 M NaCl. Btn2p was originally identified as being up-regulated in a btn1Δ strain, which lacks the vacuolar-lysosomal membrane protein, Btn1p, and serves as a model for Batten disease. This up-regulation of Btn2p was shown to contribute to the maintenance of a stable vacuolar pH in the btn1Δ strain. Btn1p was subsequently shown to be required for the optimal transport of arginine into the vacuole. Interestingly, btn1Δ ist2Δ strains are also unable to grow in the presence of 0.5 or 1.0 M NaCl, and ist2Δ suppresses the vacuolar arginine transport defect in btn1Δ strains. Although further investigation is required, we speculate that altered vacuolar arginine transport in btn1Δ strains represents a mechanism for maintaining or balancing cellular ion homeostasis. Btn2p interacts with at least three proteins that are seemingly involved in different biological functions in different subcellular locations. Due to these multiple interactions, we conclude that Btn2p may play a regulatory role across the cell in response to alterations in the intracellular environment that may be caused by changes in amino acid levels or pH, a disruption in protein trafficking, or imbalances in ion homeostasis resulting from either genetic or environmental manipulation.

scholarly journals The HXT2 gene of Saccharomyces cerevisiae is required for high-affinity glucose transport.

1990 ◽  
Vol 10 (11) ◽  
pp. 5903-5913 ◽  
Author(s):  
A L Kruckeberg ◽  
L F Bisson
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Glucose Transport ◽  
Large Family ◽  
Transmembrane Domains ◽  
Growth Defect ◽  
Multicopy Plasmid ◽  
Sugar Transporter ◽  
Yeast Saccharomyces Cerevisiae ◽  
High Affinity ◽  
Transporter Family

The HXT2 gene of the yeast Saccharomyces cerevisiae was identified on the basis of its ability to complement the defect in glucose transport of a snf3 mutant when present on the multicopy plasmid pSC2. Analysis of the DNA sequence of HXT2 revealed an open reading frame of 541 codons, capable of encoding a protein of Mr 59,840. The predicted protein displayed high sequence and structural homology to a large family of procaryotic and eucaryotic sugar transporters. These proteins have 12 highly hydrophobic regions that could form transmembrane domains; the spacing of these putative transmembrane domains is also highly conserved. Several amino acid motifs characteristic of this sugar transporter family are also present in the HXT2 protein. An hxt2 null mutant strain lacked a significant component of high-affinity glucose transport when under derepressing (low-glucose) conditions. However, the hxt2 null mutation did not incur a major growth defect on glucose-containing media. Genetic and biochemical analyses suggest that wild-type levels of high-affinity glucose transport require the products of both the HXT2 and SNF3 genes; these genes are not linked. Low-stringency Southern blot analysis revealed a number of other sequences that cross-hybridize with HXT2, suggesting that S. cerevisiae possesses a large family of sugar transporter genes.

scholarly journals Yeast Glycogen Synthase Kinase-3 Activates Msn2p-dependent Transcription of Stress Responsive Genes

2003 ◽  
Vol 14 (1) ◽  
pp. 302-312 ◽  
Author(s):  
Yuzoh Hirata ◽  
Tomoko Andoh ◽  
Toshimasa Asahara ◽  
Akira Kikuchi
Keyword(s):  
Double Mutant ◽  
Glycogen Synthase ◽  
Binding Activity ◽  
Glycogen Synthase Kinase ◽  
Glycogen Synthase Kinase 3 ◽  
Growth Defect ◽  
Transcriptional Level ◽  
Null Mutant ◽  
Promoter Regions ◽  
Yeast Saccharomyces Cerevisiae

The yeast Saccharomyces cerevisiae has four genes,MCK1, MDS1 (RIM11),MRK1, and YOL128c, that encode homologues of mammalian glycogen synthase kinase 3 (GSK-3). A gsk-3null mutant in which these four genes are disrupted showed growth defects on galactose medium. We isolated several multicopy suppressors of this growth defect. Two of them encoded Msn2p and phosphoglucomutase (PGM). Msn2p is a transcription factor that binds to the stress-response element (STRE). PGM is an enzyme that interconverts glucose-1 phosphate and glucose-6 phosphate and is regulated by Msn2p at the transcriptional level. Expression of the mRNAs ofPGM2 and DDR2, whose promoter regions possess STRE sequences, on induction by heat shock or salt stress was reduced not only in an msn2 msn4 (msn2homologue) double mutant but also in the gsk-3 null mutant. STRE-dependent transcription was greatly inhibited in thegsk-3 null mutant or mck1 mds1 double mutant, and this phenotype was suppressed by the expression of Mck1p but not of a kinase-inactive form of Mck1p. Although Msn2p accumulated in the nucleus of the gsk-3 null mutant as well as in the wild-type strain under various stress conditions, its STRE-binding activity was reduced in extracts prepared from the gsk-3null mutant or mck1 mds1 double mutant. These results suggest that yeast GSK-3 promotes formation of a complex between Msn2p and DNA, which is required for the proper response to different forms of stress. Because neither Msn2p–GSK-3 complex formation nor GSK-3–dependent phosphorylation of Msn2p could be detected, the regulation of Msn2p by GSK-3 may be indirect.

scholarly journals The HXT1 gene product of Saccharomyces cerevisiae is a new member of the family of hexose transporters.

1991 ◽  
Vol 11 (7) ◽  
pp. 3804-3813 ◽  
Author(s):  
D A Lewis ◽  
L F Bisson
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Glucose Transport ◽  
Significant Proportion ◽  
Growth Defect ◽  
Lacz Gene ◽  
Multicopy Plasmid ◽  
Hexose Transport ◽  
Yeast Saccharomyces Cerevisiae ◽  
High Affinity ◽  
Membrane Spanning

Two novel genes affecting hexose transport in the yeast Saccharomyces cerevisiae have been identified. The gene HXT1 (hexose transport), isolated from plasmid pSC7, was sequenced and found to encode a hydrophobic protein which is highly homologous to the large family of sugar transporter proteins from eucaryotes and procaryotes. Multicopy expression of the HXT1 gene restored high-affinity glucose transport to the snf3 mutant, which is deficient in a significant proportion of high-affinity glucose transport. HXT1 was unable to complement the snf3 growth defect in low copy number. The HXT1 protein was found to contain 12 putative membrane-spanning domains with a central hydrophilic domain and hydrophilic N- and C-terminal domains. The HXT1 protein is 69% identical to GAL2 and 66% identical to HXT2, and all three proteins were found to have a putative leucine zipper motif at a consensus location in membrane-spanning domain 2. Disruption of the HXT1 gene resulted in loss of a portion of high-affinity glucose and mannose transport, and wild-type levels of transport required both the HXT1 and SNF3 genes. Unexpectedly, expression of beta-galactosidase activity by using a fusion of the lacZ gene to the HXT1 promoter in a multicopy plasmid was maximal during lag and early exponential phases of growth, decreasing approximately 100-fold upon further entry into exponential growth. Deletion analysis of pSC7 revealed the presence of another gene (called ORF2) capable of suppressing the snf3 null mutant phenotype by restoring high-affinity glucose transport and increased low-affinity transport.

scholarly journals The Toxic Effects of Ppz1 Overexpression Involve Nha1-Mediated Deregulation of K+ and H+ Homeostasis

2021 ◽  
Vol 7 (12) ◽  
pp. 1010
Author(s):  
Marcel Albacar ◽  
Lenka Sacka ◽  
Carlos Calafí ◽  
Diego Velázquez ◽  
Antonio Casamayor ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Deleterious Effect ◽  
Growth Defect ◽  
Profound Change ◽  
Yeast Saccharomyces Cerevisiae ◽  
Functional Disturbance ◽  
High Affinity ◽  
Excessive Entry ◽  
Potassium Influx ◽  
Cytosolic Acidification

The alteration of the fine-tuned balance of phospho/dephosphorylation reactions in the cell often results in functional disturbance. In the yeast Saccharomyces cerevisiae, the overexpression of Ser/Thr phosphatase Ppz1 drastically blocks cell proliferation, with a profound change in the transcriptomic and phosphoproteomic profiles. While the deleterious effect on growth likely derives from the alteration of multiple targets, the precise mechanisms are still obscure. Ppz1 is a negative effector of potassium influx. However, we show that the toxic effect of Ppz1 overexpression is unrelated to the Trk1/2 high-affinity potassium importers. Cells overexpressing Ppz1 exhibit decreased K+ content, increased cytosolic acidification, and fail to properly acidify the medium. These effects, as well as the growth defect, are counteracted by the deletion of NHA1 gene, which encodes a plasma membrane Na+, K+/H+ antiporter. The beneficial effect of a lack of Nha1 on the growth vanishes as the pH of the medium approaches neutrality, is not eliminated by the expression of two non-functional Nha1 variants (D145N or D177N), and is exacerbated by a hyperactive Nha1 version (S481A). All our results show that high levels of Ppz1 overactivate Nha1, leading to an excessive entry of H+ and efflux of K+, which is detrimental for growth.

scholarly journals Altered Na+ and Li+Homeostasis in Saccharomyces cerevisiae Cells Expressing the Bacterial Cation Antiporter NhaA

1998 ◽  
Vol 180 (12) ◽  
pp. 3131-3136 ◽  
Author(s):  
Roc Ros ◽  
Consuelo Montesinos ◽  
Abraham Rimon ◽  
Etana Padan ◽  
Ramón Serrano
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Plasma Membrane ◽  
Double Mutant ◽  
Vacuolar Membrane ◽  
Intracellular Sodium ◽  
Intracellular Pool ◽  
Sodium Accumulation ◽  
Sodium Tolerance ◽  
Yeast Mutants ◽  
In Cells

ABSTRACT The bacterial Na+(Li+)/H+antiporter NhaA has been expressed in the yeast Saccharomyces cerevisiae. NhaA was present in both the plasma membrane and internal membranes, and it conferred lithium but not sodium tolerance. In cells containing the yeast Ena1-4 (Na+, Li+) extrusion ATPase, the extra lithium tolerance conferred by NhaA was dependent on a functional vacuolar H+ ATPase and correlated with an increase of lithium in an intracellular pool which exhibited slow efflux of cations. In yeast mutants without (Na+, Li+) ATPase, lithium tolerance conferred by NhaA was not dependent on a functional vacuolar H+ ATPase and correlated with a decrease of intracellular lithium. NhaA was able to confer sodium tolerance and to decrease intracellular sodium accumulation in a double mutant devoid of both plasma membrane (Na+, Li+) ATPase and vacuolar H+ ATPase. These results indicate that the bacterial antiporter NhaA expressed in yeast is functional at both the plasma membrane and the vacuolar membrane. The phenotypes conferred by its expression depend on the functionality of plasma membrane (Na+, Li+) ATPase and vacuolar H+ ATPase.

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