scholarly journals Doubling Ty1 element copy number in Saccharomyces cerevisiae: host genome stability and phenotypic effects.

Genetics ◽  
1991 ◽  
Vol 129 (4) ◽  
pp. 1043-1052
Author(s):  
J D Boeke ◽  
D J Eichinger ◽  
G Natsoulis
Keyword(s):  
Copy Number ◽  
Genome Stability ◽  
Genome Structure ◽  
Yeast Cells ◽  
Wild Type ◽  
Opposite Mating Type ◽  
Spore Viability ◽  
Normal Number ◽  
Yeast Strains ◽  
Type Strains

Abstract Haploid yeast strains bearing approximately double the normal number of Ty1 elements have been constructed using marked GAL/Ty1 fusion plasmids. The strains maintain their high transposon copy number and overall genome structure in the absence of selection. The strains bearing extra Ty1 copies are surprisingly similar phenotypically to the parental strain. The results suggest that the limit to transposon copy number, if any, has not been reached. When these strains are crossed by wild-type strains (i.e., bearing the normal complement of Ty1 elements) or by strains of opposite mating type also bearing excess Ty1 elements, normal to very slightly reduced spore viability is observed, indicating that increasing the extent of transposon homology scattered around the genome does not result in significant increases in frequency of ectopic reciprocal recombination. The results suggest that yeast cells have evolved mechanisms for coping with excess transposon copies in the genome.

scholarly journals RSR1, a ras-like gene homologous to Krev-1 (smg21A/rap1A): role in the development of cell polarity and interactions with the Ras pathway in Saccharomyces cerevisiae.

1992 ◽  
Vol 12 (2) ◽  
pp. 758-766 ◽  
Author(s):  
R Ruggieri ◽  
A Bender ◽  
Y Matsui ◽  
S Powers ◽  
Y Takai ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Polarity ◽  
Copy Number ◽  
Mammalian Cells ◽  
Direct Interaction ◽  
Dominant Negative ◽  
Yeast Cells ◽  
Wild Type ◽  
Ras Gene ◽  
Ras Pathway

The Saccharomyces cerevisiae ras-like gene RSR1 is particularly closely related to the mammalian gene Krev-1 (also known as smg21A and rap1A). RSR1 was originally isolated as a multicopy suppressor of a cdc24 mutation, which causes an inability to bud or establish cell polarity. Deletion of RSR1 itself does not affect growth but causes a randomization of bud position. We have now constructed mutant alleles of RSR1 encoding proteins with substitutions of Val for Gly at position 12 (analogous to constitutively activated Ras proteins) or Asn for Lys at position 16 (analogous to a dominant-negative Ras protein). rsr1Val-12 could not restore a normal budding pattern to an rsr1 deletion strain but could suppress a cdc24 mutation when overexpressed. rsr1Asn-16 could randomize the budding pattern of a wild-type strain even in low copy number but was not lethal even in high copy number. These and other results suggest that Rsr1p functions only in bud site selection and not in subsequent events of polarity establishment and bud formation, that this function involves a cycling between GTP-bound and GDP-bound forms of the protein, and that the suppression of cdc24 involves direct interaction between Rsr1p[GTP] and Cdc24p. Functional homology between Rsr1p and Krev-1 p21 was suggested by the observations that expression of the latter protein in yeast cells could both suppress a cdc24 mutation and randomize the budding pattern of wild-type cells. As Krev-1 overexpression can suppress ras-induced transformation of mammalian cells, we looked for effects of RSR1 on the S. cerevisiae Ras pathway. Although no suppression of the activated RAS2Val-19 allele was observed, overexpression of rsr1Val-12 suppressed the lethality of strains lacking RAS gene function, apparently through a direct activation of adenyl cyclase. This interaction of Rsr1p with the effector of Ras in S. cerevisiae suggests that Krev-1 may revert ras-induced transformation of mammalian cells by affecting the interaction of ras p21 with its effector.

scholarly journals Molecular analysis of the yeast VPS3 gene and the role of its product in vacuolar protein sorting and vacuolar segregation during the cell cycle.

1990 ◽  
Vol 111 (3) ◽  
pp. 877-892 ◽  
Author(s):  
C K Raymond ◽  
P J O'Hara ◽  
G Eichinger ◽  
J H Rothman ◽  
T H Stevens
Keyword(s):  
Selection Procedure ◽  
Temperature Shift ◽  
Yeast Cells ◽  
Carboxypeptidase Y ◽  
Wild Type ◽  
Yeast Saccharomyces Cerevisiae ◽  
Yeast Strains ◽  
Type Pattern ◽  
Mother Cells ◽  
Vacuolar Protein

vps3 mutants of the yeast Saccharomyces cerevisiae are impaired in the sorting of newly synthesized soluble vacuolar proteins and in the acidification of the vacuole (Rothman, J. H., and T. H. Stevens. Cell. 47:1041-1051; Rothman, J. H., C. T. Yamashiro, C. K. Raymond, P. M. Kane, and T. H. Stevens. 1989. J. Cell Biol. 109:93-100). The VPS3 gene, which was cloned using a novel selection procedure, encodes a low abundance, hydrophilic protein of 117 kD that most likely resides in the cytoplasm. Yeast strains bearing a deletion of the VPS3 gene (vps3-delta 1) are viable, yet their growth rate is significantly reduced relative to wild-type cells. Temperature shift experiments with strains carrying a temperature conditional vps3 allele demonstrate that cells rapidly lose the capacity to sort the vacuolar protein carboxypeptidase Y upon loss of VPS3 function. Vacuolar morphology was examined in wild-type and vps3-delta 1 yeast strains by fluorescence microscopy. The vacuoles in wild-type yeast cells are morphologically complex, and they appear to be actively partitioned between mother cells and buds during an early phase of bud growth. Vacuolar morphology in vps3-delta 1 mutants is significantly altered from the wild-type pattern, and the vacuolar segregation process seen in wild-type strains is defective in these mutants. With the exception of a vacuolar acidification defect, the phenotypes of vps3-delta 1 strains are significantly different from those of mutants lacking the vacuolar proton-translocating ATPase. These data demonstrate that the acidification defect in vps3-delta 1 cells is not the primary cause of the pleiotropic defects in vacuolar function observed in these mutants.

Multistress resistance of Saccharomyces cerevisiae is generated by insertion of retrotransposon Ty into the 5' coding region of the adenylate cyclase gene

1988 ◽  
Vol 8 (12) ◽  
pp. 5555-5560
Author(s):  
H Iida
Keyword(s):  
Heat Treatment ◽  
Saccharomyces Cerevisiae ◽  
Heat Shock ◽  
Adenylate Cyclase ◽  
Uv Light ◽  
Yeast Cells ◽  
Wild Type ◽  
Coding Region ◽  
Resistant Mutants ◽  
Type Strains

Heat shock-resistant mutants, which were isolated by their ability to withstand lethal heat treatment, were characterized. Resistance was demonstrated to be a consequence of insertion of retrotransposon Ty into either the 5' coding or noncoding region, close to the putative initiation codon of the adenylate cyclase gene CYR1 (or CDC35). These heat shock-resistant mutants contained about threefold lower adenylate cyclase activity than wild-type strains. The mutants were also observed to be resistant to other stresses such as UV light and ethanol. These results demonstrate that multistress resistance, which may confer a survival advantage to yeast cells, can be generated by transposition of a Ty element into CYR1.

scholarly journals Involvement of the Checkpoint Protein Mec1p in Silencing of Gene Expression at Telomeres in Saccharomyces cerevisiae

2000 ◽  
Vol 20 (7) ◽  
pp. 2378-2384 ◽  
Author(s):  
Rolf J. Craven ◽  
Thomas D. Petes
Keyword(s):  
Gene Expression ◽  
Yeast Cells ◽  
Wild Type ◽  
Checkpoint Response ◽  
Dna Damaging Agents ◽  
Yeast Strains ◽  
Telomeric Silencing ◽  
Checkpoint Pathway ◽  
Wild Type Yeast ◽  
Silence Gene

ABSTRACT Yeast strains with a mutation in the MEC1 gene are deficient in the cellular checkpoint response to DNA-damaging agents and have short telomeres (K. B. Ritchie, J. C. Mallory, and T. D. Petes, Mol. Cell. Biol. 19:6065–6075, 1999; T. A. Weinert, G. L. Kiser, and L. H. Hartwell, Genes Dev. 8:652–665, 1994). In wild-type yeast cells, genes inserted near the telomeres are transcriptionally silenced (D. E. Gottschling, O. M. Aparichio, B. L. Billington, and V. A. Zakian, Cell 63:751–762, 1990). We show that mec1strains have reduced ability to silence gene expression near the telomere. This deficiency was alleviated by the sml1mutation. Overexpression of Mec1p also resulted in a silencing defect, although this overexpression did not affect the checkpoint function of Mec1p. Telomeric silencing was not affected by mutations in several other genes in the Mec1p checkpoint pathway (null mutations inRAD9 and CHK1 or in several hypomorphicrad53 alleles) but was reduced by a null mutation ofDUN1. In addition, the loss of telomeric silencing inmec1 strains was not a consequence of the slightly shortened telomeres observed in these strains.

scholarly journals Deletion of Yeast p24 Genes Activates the Unfolded Protein Response

2001 ◽  
Vol 12 (4) ◽  
pp. 957-969 ◽  
Author(s):  
William J. Belden ◽  
Charles Barlowe
Keyword(s):  
Unfolded Protein Response ◽  
Yeast Cells ◽  
Secretory Proteins ◽  
Wild Type ◽  
Vitro Assay ◽  
Unfolded Protein ◽  
Extracellular Secretion ◽  
The Unfolded Protein Response ◽  
Protein Response ◽  
Type Strains

Yeast cells lacking a functional p24 complex accumulate a subset of secretory proteins in the endoplasmic reticulum (ER) and increase the extracellular secretion of HDEL-containing ER residents such as Kar2p/BiP. We report that a loss of p24 function causes activation of the unfolded protein response (UPR) and leads to increasedKAR2 expression. The HDEL receptor (Erd2p) is functional and traffics in p24 deletion strains as in wild-type strains, however the capacity of the retrieval pathway is exceeded. Other conditions that activate the UPR and elevate KAR2 expression also lead to extracellular secretion of Kar2p. Using an in vitro assay that reconstitutes budding from the ER, we detect elevated levels of Kar2p in ER-derived vesicles from p24 deletion strains and from wild-type strains with an activated UPR. Silencing the UPR byIRE1 deletion diminished Kar2p secretion under these conditions. We suggest that activation of the UPR plays a major role in extracellular secretion of Kar2p.

scholarly journals Multistress resistance of Saccharomyces cerevisiae is generated by insertion of retrotransposon Ty into the 5' coding region of the adenylate cyclase gene.

1988 ◽  
Vol 8 (12) ◽  
pp. 5555-5560 ◽  
Author(s):  
H Iida
Keyword(s):  
Heat Treatment ◽  
Saccharomyces Cerevisiae ◽  
Heat Shock ◽  
Adenylate Cyclase ◽  
Uv Light ◽  
Yeast Cells ◽  
Wild Type ◽  
Coding Region ◽  
Resistant Mutants ◽  
Type Strains

Heat shock-resistant mutants, which were isolated by their ability to withstand lethal heat treatment, were characterized. Resistance was demonstrated to be a consequence of insertion of retrotransposon Ty into either the 5' coding or noncoding region, close to the putative initiation codon of the adenylate cyclase gene CYR1 (or CDC35). These heat shock-resistant mutants contained about threefold lower adenylate cyclase activity than wild-type strains. The mutants were also observed to be resistant to other stresses such as UV light and ethanol. These results demonstrate that multistress resistance, which may confer a survival advantage to yeast cells, can be generated by transposition of a Ty element into CYR1.

scholarly journals Diversity of Genome Structure in Salmonella enterica Serovar Typhi Populations

2005 ◽  
Vol 187 (8) ◽  
pp. 2638-2650 ◽  
Author(s):  
Sushma Kothapalli ◽  
Satheesh Nair ◽  
Suneetha Alokam ◽  
Tikki Pang ◽  
Rasik Khakhria ◽  
...  
Keyword(s):  
Salmonella Enterica ◽  
Gene Dosage ◽  
Genome Structure ◽  
Wild Type ◽  
Salmonella Enterica Serovar Typhi ◽  
Large Deletions ◽  
Phage Types ◽  
Partial Digestion ◽  
Serovar Typhimurium ◽  
Type Strains

ABSTRACT The genomes of most strains of Salmonella and Escherichia coli are highly conserved. In contrast, all 136 wild-type strains of Salmonella enterica serovar Typhi analyzed by partial digestion with I-CeuI (an endonuclease which cuts within the rrn operons) and pulsed-field gel electrophoresis and by PCR have rearrangements due to homologous recombination between the rrn operons leading to inversions and translocations. Recombination between rrn operons in culture is known to be equally frequent in S. enterica serovar Typhi and S. enterica serovar Typhimurium; thus, the recombinants in S. enterica serovar Typhi, but not those in S. enterica serovar Typhimurium, are able to survive in nature. However, even in S. enterica serovar Typhi the need for genome balance and the need for gene dosage impose limits on rearrangements. Of 100 strains of genome types 1 to 6, 72 were only 25.5 kb off genome balance (the relative lengths of the replichores during bidirectional replication from oriC to the termination of replication [Ter]), while 28 strains were less balanced (41 kb off balance), indicating that the survival of the best-balanced strains was greater. In addition, the need for appropriate gene dosage apparently selected against rearrangements which moved genes from their accustomed distance from oriC. Although rearrangements involving the seven rrn operons are very common in S. enterica serovar Typhi, other duplicated regions, such as the 25 IS200 elements, are very rarely involved in rearrangements. Large deletions and insertions in the genome are uncommon, except for deletions of Salmonella pathogenicity island 7 (usually 134 kb) from fragment I-CeuI-G and 40-kb insertions, possibly a prophage, in fragment I-CeuI-E. The phage types were determined, and the origins of the phage types appeared to be independent of the origins of the genome types.

scholarly journals Characterization of yeast strains with conditionally expressed variants of ribosomal protein genes tcm1 and cyh2

1985 ◽  
Vol 5 (1) ◽  
pp. 99-108
Author(s):  
H M Fried ◽  
H G Nam ◽  
S Loechel ◽  
J Teem
Keyword(s):  
Ribosomal Protein ◽  
Ribosomal Proteins ◽  
Yeast Cells ◽  
Regulatory Sequence ◽  
Ribosomal Protein Genes ◽  
Wild Type ◽  
Yeast Strains ◽  
Reduced Rate ◽  
Hybrid Genes

We placed a regulatory sequence derived from the GAL10 locus of Saccharomyces cerevisiae at various distances from the start sites of transcription of two yeast ribosomal protein genes, tcm1 and cyh2. The hybrid ribosomal protein genes were transcribed at wild-type levels in the presence of galactose. In the absence of galactose, the hybrid genes were transcribed either at a reduced level or essentially not at all. Yeast cells which transcribe the ribosomal protein genes at a reduced rate continued to grow, suggesting that enhanced translation of the ribosomal protein mRNA may permit an adequate rate of synthesis of the corresponding protein. Consistent with this suggestion is the finding that preexisting mRNA decayed at a reduced rate when transcription was halted abruptly by removal of galactose. Yeast cells unable to transcribe tcm1 or cyh2 without galactose did not grow. These conditional lethal strains demonstrate that the ribosomal proteins encoded by tcm1 and cyh2 are essential; furthermore, these strains are potentially useful for isolating mutations in the tcm1 and cyh2 proteins affecting their transport, assembly, or function.

scholarly journals RSR1, a ras-like gene homologous to Krev-1 (smg21A/rap1A): role in the development of cell polarity and interactions with the Ras pathway in Saccharomyces cerevisiae

1992 ◽  
Vol 12 (2) ◽  
pp. 758-766
Author(s):  
R Ruggieri ◽  
A Bender ◽  
Y Matsui ◽  
S Powers ◽  
Y Takai ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Polarity ◽  
Copy Number ◽  
Mammalian Cells ◽  
Direct Interaction ◽  
Dominant Negative ◽  
Yeast Cells ◽  
Wild Type ◽  
Ras Gene ◽  
Ras Pathway

The Saccharomyces cerevisiae ras-like gene RSR1 is particularly closely related to the mammalian gene Krev-1 (also known as smg21A and rap1A). RSR1 was originally isolated as a multicopy suppressor of a cdc24 mutation, which causes an inability to bud or establish cell polarity. Deletion of RSR1 itself does not affect growth but causes a randomization of bud position. We have now constructed mutant alleles of RSR1 encoding proteins with substitutions of Val for Gly at position 12 (analogous to constitutively activated Ras proteins) or Asn for Lys at position 16 (analogous to a dominant-negative Ras protein). rsr1Val-12 could not restore a normal budding pattern to an rsr1 deletion strain but could suppress a cdc24 mutation when overexpressed. rsr1Asn-16 could randomize the budding pattern of a wild-type strain even in low copy number but was not lethal even in high copy number. These and other results suggest that Rsr1p functions only in bud site selection and not in subsequent events of polarity establishment and bud formation, that this function involves a cycling between GTP-bound and GDP-bound forms of the protein, and that the suppression of cdc24 involves direct interaction between Rsr1p[GTP] and Cdc24p. Functional homology between Rsr1p and Krev-1 p21 was suggested by the observations that expression of the latter protein in yeast cells could both suppress a cdc24 mutation and randomize the budding pattern of wild-type cells. As Krev-1 overexpression can suppress ras-induced transformation of mammalian cells, we looked for effects of RSR1 on the S. cerevisiae Ras pathway. Although no suppression of the activated RAS2Val-19 allele was observed, overexpression of rsr1Val-12 suppressed the lethality of strains lacking RAS gene function, apparently through a direct activation of adenyl cyclase. This interaction of Rsr1p with the effector of Ras in S. cerevisiae suggests that Krev-1 may revert ras-induced transformation of mammalian cells by affecting the interaction of ras p21 with its effector.

scholarly journals MUTATIONS IN THE PHO80 GENE CONFER PERMEABILITY TO 5'-MONONUCLEOTIDES IN SACCHAROMYCES CEREVISIAE

Genetics ◽  
1982 ◽  
Vol 102 (3) ◽  
pp. 341-359
Author(s):  
Linda F Bisson ◽  
Jeremy Thorner
Keyword(s):  
Inorganic Phosphate ◽  
Phosphate Transport ◽  
Culture Conditions ◽  
Yeast Cells ◽  
Wild Type ◽  
Coordinate Control ◽  
Complementation Groups ◽  
The Right ◽  
Yeast Mutants ◽  
Type Strains

ABSTRACT Yeast mutants permeable to dTMP (tup) were selected and two new complementation groups (tup5 and tup7) were identified. Assay of the levels of both acid and alkaline phosphatase in cells grown under either repressing (5 mm PO4  -3) or derepressing (0.03 mm PO4  -3) conditions indicated that, in general, tup mutations cause cells to be defective in their regulation of phosphatase synthesis. In addition, three of the tup mutations (tup1, tup4 and tup7) displayed markedly elevated rates of inorganic phosphate transport. The tup7 locus was found to be tightly centromere-linked on the right arm of chromosome XV, and was shown to be allelic with the pho80 regulatory locus on the basis of both genetic and biochemical criteria. Analysis of other mutations known to affect phosphatase levels (pho) indicated that some also conferred permeability to dTMP. Possible allelic relationships between tup genes and certain of these pho mutations are discussed. Regardless of the culture conditions, wild-type strains were not permeable to dTMP; in contrast, it was found in the course of this work that normal yeast cells were permeable to dUMP and that dUMP permeability was regulated by the concentration of inorganic phosphate present in the medium used to grow the cells. Thus, permeability to 5′-mononucleotides appears to be under coordinate control with phosphatase synthesis.

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