Putative GTP-binding protein, Gtr1, associated with the function of the Pho84 inorganic phosphate transporter in Saccharomyces cerevisiae

1992 ◽  
Vol 12 (7) ◽  
pp. 2958-2966
Author(s):  
M Bun-Ya ◽  
S Harashima ◽  
Y Oshima
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Amino Acid Residues ◽  
Transport Activity ◽  
Reading Frame ◽  
Gtp Binding ◽  
Pi Transport ◽  
Recombination System ◽  
Opposite Strand ◽  
Constitutive Synthesis ◽  
Site Specific Recombination System

We have found an open reading frame which is 1.1 kb upstream of PHO84 (which encodes a Pi transporter) and is transcribed from the opposite strand. In Saccharomyces cerevisiae, this gene is distal to the TUB3 locus on the left arm of chromosome XIII and is named GTR1. GTR1 encodes a protein consisting of 310 amino acid residues containing, in its N-terminal region, the characteristic tripartite consensus elements for binding GTP conserved in GTP-binding proteins, except for histidine in place of a widely conserved aspargine residue in element III. Disruption of the GTR1 gene resulted in slow growth at 30 degrees C and no growth at 15 degrees C; other phenotypes resembled those of pho84 mutants and included constitutive synthesis of repressible acid phosphatase, reduced Pi transport activity, and resistance to arsenate. The latter phenotypes were shown to be due to a defect in Pi uptake, and the Gtr1 protein was found to be functionally associated with the Pho84 Pi transporter. Recombination between chromosome V (at the URA3 locus) and chromosome XIII (in the GTR1-PHO84-TUB3 region) by using a plasmid-encoded site-specific recombination system indicated that the order of these genes was telomere-TUB3-PHO84-GTR1-CENXIII.

scholarly journals Putative GTP-binding protein, Gtr1, associated with the function of the Pho84 inorganic phosphate transporter in Saccharomyces cerevisiae.

1992 ◽  
Vol 12 (7) ◽  
pp. 2958-2966 ◽  
Author(s):  
M Bun-Ya ◽  
S Harashima ◽  
Y Oshima
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Amino Acid Residues ◽  
Transport Activity ◽  
Reading Frame ◽  
Gtp Binding ◽  
Pi Transport ◽  
Recombination System ◽  
Opposite Strand ◽  
Constitutive Synthesis ◽  
Site Specific Recombination System

We have found an open reading frame which is 1.1 kb upstream of PHO84 (which encodes a Pi transporter) and is transcribed from the opposite strand. In Saccharomyces cerevisiae, this gene is distal to the TUB3 locus on the left arm of chromosome XIII and is named GTR1. GTR1 encodes a protein consisting of 310 amino acid residues containing, in its N-terminal region, the characteristic tripartite consensus elements for binding GTP conserved in GTP-binding proteins, except for histidine in place of a widely conserved aspargine residue in element III. Disruption of the GTR1 gene resulted in slow growth at 30 degrees C and no growth at 15 degrees C; other phenotypes resembled those of pho84 mutants and included constitutive synthesis of repressible acid phosphatase, reduced Pi transport activity, and resistance to arsenate. The latter phenotypes were shown to be due to a defect in Pi uptake, and the Gtr1 protein was found to be functionally associated with the Pho84 Pi transporter. Recombination between chromosome V (at the URA3 locus) and chromosome XIII (in the GTR1-PHO84-TUB3 region) by using a plasmid-encoded site-specific recombination system indicated that the order of these genes was telomere-TUB3-PHO84-GTR1-CENXIII.

The PHO84 gene of Saccharomyces cerevisiae encodes an inorganic phosphate transporter

1991 ◽  
Vol 11 (6) ◽  
pp. 3229-3238
Author(s):  
M Bun-Ya ◽  
M Nishimura ◽  
S Harashima ◽  
Y Oshima
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Fusion Gene ◽  
Amino Acid Residues ◽  
Sugar Transporters ◽  
Pi Transport ◽  
Constitutive Synthesis ◽  
A Site ◽  
Mutant Cells ◽  
Hydropathy Analysis ◽  
Beta Galactosidase

The PHO84 gene specifies Pi-transport in Saccharomyces cerevisiae. A DNA fragment bearing the PHO84 gene was cloned by its ability to complement constitutive synthesis of repressible acid phosphatase of pho84 mutant cells. Its nucleotide sequence predicted a protein of 596 amino acids with a sequence homologous to that of a superfamily of sugar transporters. Hydropathy analysis suggested that the secondary structure of the PHO84 protein consists of two blocks of six transmembrane domains separated by 74 amino acid residues. The cloned PH084 DNA restored the Pi transport activity of pho84 mutant cells. The PHO84 transcription was regulated by Pi like those of the PHO5, PHO8, and PHO81 genes. A PHO84-lacZ fusion gene produced beta-galactosidase activity under the regulation of Pi, and the activity was suggested to be bound to a membrane fraction. Gene disruption of PHO84 was not lethal. By comparison of nucleotide sequences and by tetrad analysis with GAL80 as a standard, the PHO84 locus was mapped at a site beside the TUB3 locus on the left arm of chromosome XIII.

scholarly journals The PHO84 gene of Saccharomyces cerevisiae encodes an inorganic phosphate transporter.

1991 ◽  
Vol 11 (6) ◽  
pp. 3229-3238 ◽  
Author(s):  
M Bun-Ya ◽  
M Nishimura ◽  
S Harashima ◽  
Y Oshima
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Fusion Gene ◽  
Amino Acid Residues ◽  
Sugar Transporters ◽  
Pi Transport ◽  
Constitutive Synthesis ◽  
A Site ◽  
Mutant Cells ◽  
Hydropathy Analysis ◽  
Beta Galactosidase

The PHO84 gene specifies Pi-transport in Saccharomyces cerevisiae. A DNA fragment bearing the PHO84 gene was cloned by its ability to complement constitutive synthesis of repressible acid phosphatase of pho84 mutant cells. Its nucleotide sequence predicted a protein of 596 amino acids with a sequence homologous to that of a superfamily of sugar transporters. Hydropathy analysis suggested that the secondary structure of the PHO84 protein consists of two blocks of six transmembrane domains separated by 74 amino acid residues. The cloned PH084 DNA restored the Pi transport activity of pho84 mutant cells. The PHO84 transcription was regulated by Pi like those of the PHO5, PHO8, and PHO81 genes. A PHO84-lacZ fusion gene produced beta-galactosidase activity under the regulation of Pi, and the activity was suggested to be bound to a membrane fraction. Gene disruption of PHO84 was not lethal. By comparison of nucleotide sequences and by tetrad analysis with GAL80 as a standard, the PHO84 locus was mapped at a site beside the TUB3 locus on the left arm of chromosome XIII.

scholarly journals Temperature-sensitive cdc7 mutations of Saccharomyces cerevisiae are suppressed by the DBF4 gene, which is required for the G1/S cell cycle transition.

Genetics ◽  
1992 ◽  
Vol 131 (1) ◽  
pp. 21-29 ◽  
Author(s):  
K Kitada ◽  
L H Johnston ◽  
T Sugino ◽  
A Sugino
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
Genomic Library ◽  
Temperature Sensitive ◽  
Multicopy Plasmid ◽  
Amino Acid Residues ◽  
Reading Frame ◽  
Allele Specific ◽  
Cell Cycle Transition ◽  
Mutant Cells

Abstract When present on a multicopy plasmid, a gene from a Saccharomyces cerevisiae genomic library suppresses the temperature-sensitive cdc7-1 mutation. The gene was identified as DBF4, which was previously isolated by complementation in dbf4-1 mutant cells and is required for the G1----S phase progression of the cell cycle. DBF4 has an open reading frame encoding 695 amino acid residues and the predicted molecular mass of the gene product is 80 kD. The suppression is allele-specific because a CDC7 deletion is not suppressed by DBF4. Suppression is mitosis-specific and the sporulation defect of cdc7 mutations is not suppressed by DBF4. Conversely, CDC7 on a multicopy plasmid suppresses the dbf4-1, -2, -3 and -4 mutations but not dbf4-5 and DBF4 deletion mutations. Furthermore, cdc7 mutations are incompatible with the temperature-sensitive dbf4 mutations. These results suggest that the CDC7 and DBF4 polypeptides interact directly or indirectly to permit initiation of yeast chromosome replication.

Functional expression of the cre-lox site-specific recombination system in the yeast Saccharomyces cerevisiae

1987 ◽  
Vol 7 (6) ◽  
pp. 2087-2096
Author(s):  
B Sauer
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Functional Expression ◽  
Yeast Saccharomyces Cerevisiae ◽  
Efficient Manner ◽  
Site Specific ◽  
Site Specific Recombination ◽  
Leu2 Gene ◽  
Recombination System ◽  
A Site ◽  
Site Specific Recombination System

The procaryotic cre-lox site-specific recombination system of coliphage P1 was shown to function in an efficient manner in a eucaryote, the yeast Saccharomyces cerevisiae. The cre gene, which codes for a site-specific recombinase, was placed under control of the yeast GALI promoter. lox sites flanking the LEU2 gene were integrated into two different chromosomes in both orientations. Excisive recombination at the lox sites (as measured by loss of the LEU2 gene) was promoted efficiently and accurately by the Cre protein and was dependent upon induction by galactose. These results demonstrate that a procaryotic recombinase can enter a eucaryotic nucleus and, moreover, that the ability of the Cre recombinase to perform precise recombination events on the chromosomes of S. cerevisiae is unimpaired by chromatin structure.

scholarly journals Functional expression of the cre-lox site-specific recombination system in the yeast Saccharomyces cerevisiae.

1987 ◽  
Vol 7 (6) ◽  
pp. 2087-2096 ◽  
Author(s):  
B Sauer
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Functional Expression ◽  
Yeast Saccharomyces Cerevisiae ◽  
Efficient Manner ◽  
Site Specific ◽  
Site Specific Recombination ◽  
Leu2 Gene ◽  
Recombination System ◽  
A Site ◽  
Site Specific Recombination System

The procaryotic cre-lox site-specific recombination system of coliphage P1 was shown to function in an efficient manner in a eucaryote, the yeast Saccharomyces cerevisiae. The cre gene, which codes for a site-specific recombinase, was placed under control of the yeast GALI promoter. lox sites flanking the LEU2 gene were integrated into two different chromosomes in both orientations. Excisive recombination at the lox sites (as measured by loss of the LEU2 gene) was promoted efficiently and accurately by the Cre protein and was dependent upon induction by galactose. These results demonstrate that a procaryotic recombinase can enter a eucaryotic nucleus and, moreover, that the ability of the Cre recombinase to perform precise recombination events on the chromosomes of S. cerevisiae is unimpaired by chromatin structure.

scholarly journals The maltose permease encoded by the MAL61 gene of Saccharomyces cerevisiae exhibits both sequence and structural homology to other sugar transporters.

Genetics ◽  
1989 ◽  
Vol 123 (3) ◽  
pp. 477-484 ◽  
Author(s):  
Q Cheng ◽  
C A Michels
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Glucose Transporter ◽  
Kluyveromyces Lactis ◽  
Lactose Permease ◽  
Structural Homology ◽  
Transport Activity ◽  
Maltose Permease ◽  
Reading Frame ◽  
Sugar Transporters ◽  
Hydropathy Analysis

Abstract The MAL61 gene of Saccharomyces cerevisiae encodes maltose permease, a protein required for the transport of maltose across the plasma membrane. Here we report the nucleotide sequence of the cloned MAL61 gene. A single 1842 bp open reading frame is present within this region encoding the 614 residue putative MAL61 protein. Hydropathy analysis suggests that the secondary structure consists of two blocks of six transmembrane domains separated by an approximately 71 residue intracellular region. The N-terminal and C-terminal domains of 100 and 67 residues in length, respectively, also appear to be intracellular. Significant sequence and structural homology is seen between the MAL61 protein and the Saccharomyces high-affinity glucose transporter encoded by the SNF3 gene, the Kluyveromyces lactis lactose permease encoded by the LAC12 gene, the human HepG2 glucose transporter and the Escherichia coli xylose and arabinose transporters encoded by the xylE and araE genes, indicating that all are members of a family of sugar transporters and are related either functionally or evolutionarily. A mechanism for glucose-induced inactivation of maltose transport activity is discussed.

scholarly journals Recombinational instability of a chimeric plasmid in Saccharomyces cerevisiae.

1984 ◽  
Vol 4 (1) ◽  
pp. 195-198 ◽  
Author(s):  
M S Whiteway ◽  
A Ahmed
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mitotic Recombination ◽  
Wild Type ◽  
Recombination System ◽  
Sequence Homologies ◽  
Selection For ◽  
Derivative Plasmid ◽  
Type Strains ◽  
Chromosomal Recombination ◽  
Site Specific Recombination System

Wild-type strains of Saccharomyces cerevisiae exhibit mitotic recombination between the chimeric plasmid TLC-1 and the endogenous 2mu circle that involves sequence homologies between the two plasmids that are not acted on by the 2mu circle site-specific recombination system. This generalized recombination can be detected because it separates the LEU2 and CAN1 markers of TLC-1 from each other through the formation of a plasmid containing only the S. cerevisiae LEU2 region and the 2mu circle. This derivative plasmid is maintained more stably during vegetative growth than TLC-1, and strains which carry it frequently lose the endogenous 2mu circle. Therefore, TLC-1 can provide a convenient selection for [cir0] cells. Formation of this new plasmid is greatly reduced, but not eliminated, in strains containing the rad52-1 mutation. This indicates that generalized mitotic recombination between plasmid sequences utilizes functions required for chromosomal recombination in S. cerevisiae.

Recombinational instability of a chimeric plasmid in Saccharomyces cerevisiae

1984 ◽  
Vol 4 (1) ◽  
pp. 195-198
Author(s):  
M S Whiteway ◽  
A Ahmed
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mitotic Recombination ◽  
Wild Type ◽  
Recombination System ◽  
Sequence Homologies ◽  
Selection For ◽  
Derivative Plasmid ◽  
Type Strains ◽  
Chromosomal Recombination ◽  
Site Specific Recombination System

Wild-type strains of Saccharomyces cerevisiae exhibit mitotic recombination between the chimeric plasmid TLC-1 and the endogenous 2mu circle that involves sequence homologies between the two plasmids that are not acted on by the 2mu circle site-specific recombination system. This generalized recombination can be detected because it separates the LEU2 and CAN1 markers of TLC-1 from each other through the formation of a plasmid containing only the S. cerevisiae LEU2 region and the 2mu circle. This derivative plasmid is maintained more stably during vegetative growth than TLC-1, and strains which carry it frequently lose the endogenous 2mu circle. Therefore, TLC-1 can provide a convenient selection for [cir0] cells. Formation of this new plasmid is greatly reduced, but not eliminated, in strains containing the rad52-1 mutation. This indicates that generalized mitotic recombination between plasmid sequences utilizes functions required for chromosomal recombination in S. cerevisiae.

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