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 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 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.

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.

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.

An Essential Function of a Phosphoinositide-Specific Phospholipase C Is Relieved by Inhibition of a Cyclin-Dependent Protein Kinase in the Yeast Saccharomyces cerevisiae

Genetics ◽  
1998 ◽  
Vol 148 (1) ◽  
pp. 33-47 ◽  
Author(s):  
Jeffrey S Flick ◽  
Jeremy Thorner
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Protein Kinase ◽  
Phospholipase C ◽  
Growth Defect ◽  
Temperature Sensitive ◽  
Complete Medium ◽  
Restrictive Temperature ◽  
Dependent Protein Kinase ◽  
Yeast Saccharomyces Cerevisiae ◽  
Dependent Protein

Abstract The PLC1 gene product of Saccharomyces cerevisiae is a homolog of the δ isoform of mammalian phosphoinositide-specific phospholipase C (PI-PLC). We found that two genes (SPL1 and SPL2), when overexpressed, can bypass the temperature-sensitive growth defect of a plc1Δ cell. SPL1 is identical to the PHO81 gene, which encodes an inhibitor of a cyclin (Pho80p)-dependent protein kinase (Pho85p) complex (Cdk). In addition to overproduction of Pho81p, two other conditions that inactivate this Cdk, a cyclin (pho80Δ) mutation and growth on low-phosphate medium, also permitted growth of plc1Δ cells at the restrictive temperature. Suppression of the temperature sensitivity of plc1Δ cells by pho80Δ does not depend upon the Pho4p transcriptional regulator, the only known substrate of the Pho80p/Pho85p Cdk. The second suppressor, SPL2, encodes a small (17-kD) protein that bears similarity to the ankyrin repeat regions present in Pho81p and in other known Cdk inhibitors. Both pho81Δ and spl2Δ show a synthetic phenotype in combination with plc1Δ. Unlike single mutants, plc1Δ pho81Δ and plc1Δ spl2Δ double mutants were unable to grow on synthetic complete medium, but were able to grow on rich medium.

scholarly journals Nonlethal sec71-1 and sec72-1 mutations eliminate proteins associated with the Sec63p-BiP complex from S. cerevisiae.

1994 ◽  
Vol 5 (9) ◽  
pp. 933-942 ◽  
Author(s):  
H Fang ◽  
N Green
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Water Soluble ◽  
Growth Defect ◽  
Sequence Analyses ◽  
Gene Encoding ◽  
Yeast Saccharomyces Cerevisiae ◽  
Reading Frame ◽  
Transmembrane Glycoprotein ◽  
Genetic Assay ◽  
Er Membrane

The sec71-1 and sec72-1 mutations were identified by a genetic assay that monitored membrane protein integration into the endoplasmic reticulum (ER) membrane of the yeast Saccharomyces cerevisiae. The mutations inhibited integration of various chimeric membrane proteins and translocation of a subset of water soluble proteins. In this paper we show that SEC71 encodes the 31.5-kDa transmembrane glycoprotein (p31.5) and SEC72 encodes the 23-kDa protein (p23) of the Sec63p-BiP complex. SEC71 is therefore identical to SEC66 (HSS1), which was previously shown to encode p31.5. DNA sequence analyses reveal that sec71-1 cells contain a nonsense mutation that removes approximately two-thirds of the cytoplasmic C-terminal domain of p31.5. The sec72-1 mutation shifts the reading frame of the gene encoding p23. Unexpectedly, the sec71-1 mutant lacks p31.5 and p23. Neither mutation is lethal, although sec71-1 cells exhibit a growth defect at 37 degrees C. These results show that p31.5 and p23 are important for the trafficking of a subset of proteins to the ER membrane.

scholarly journals Secretion of Quinolinic Acid, an Intermediate in the Kynurenine Pathway, for Utilization in NAD + Biosynthesis in the Yeast Saccharomyces cerevisiae

2013 ◽  
Vol 12 (5) ◽  
pp. 648-653 ◽  
Author(s):  
Kazuto Ohashi ◽  
Shigeyuki Kawai ◽  
Kousaku Murata
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Nicotinic Acid ◽  
Quinolinic Acid ◽  
De Novo ◽  
Kynurenine Pathway ◽  
Salvage Pathway ◽  
Yeast Saccharomyces Cerevisiae ◽  
Content Type ◽  
High Affinity ◽  
Transcriptional Induction

ABSTRACT NAD + is synthesized from tryptophan either via the kynurenine ( de novo ) pathway or via the salvage pathway by reutilizing intermediates such as nicotinic acid or nicotinamide ribose. Quinolinic acid is an intermediate in the kynurenine pathway. We have discovered that the budding yeast Saccharomyces cerevisiae secretes quinolinic acid into the medium and also utilizes extracellular quinolinic acid as a novel NAD + precursor. We provide evidence that extracellular quinolinic acid enters the cell via Tna1, a high-affinity nicotinic acid permease, and thereby helps to increase the intracellular concentration of NAD + . Transcription of genes involved in the kynurenine pathway and Tna1 was increased, responding to a low intracellular NAD + concentration, in cells bearing mutations of these genes; this transcriptional induction was suppressed by supplementation with quinolinic acid or nicotinic acid. Our data thus shed new light on the significance of quinolinic acid, which had previously been recognized only as an intermediate in the kynurenine pathway.

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