The Saccharomyces cerevisiae genome contains functional and nonfunctional copies of transposon Ty1

Saccharomyces cerevisiae Ty elements are transposons closely related to retroviruses. The DNA sequence of a functional Ty element (TyH3) is presented. The long terminal repeat sequences are different, suggesting that TyH3 is a recombinant Ty element. A chromosomal Ty element near the LYS2 gene, Ty173, was found to be nonfunctional, even though it has no detectable insertions or deletions. The defect in Ty173 transposition is caused by a missense mutation giving rise to a Leu-to-Ile substitution in the TYB (pol) open reading frame. Several chromosomal Ty elements carry this lesion in their DNA, indicating that nonfunctional Ty elements are common in the yeast genome.

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Primary structure of the Saccharomyces cerevisiae GAL4 gene.

Molecular and Cellular Biology ◽

10.1128/mcb.4.2.260 ◽

1984 ◽

Vol 4 (2) ◽

pp. 260-267 ◽

Cited By ~ 133

Author(s):

A Laughon ◽

R F Gesteland

Keyword(s):

Saccharomyces Cerevisiae ◽

Amino Acid ◽

Amino Acid Sequence ◽

Dna Sequence ◽

Primary Structure ◽

Open Reading Frame ◽

Binding Motif ◽

Reading Frame ◽

Activator Proteins ◽

Inducible Genes

The GAL4 gene encodes a positive regulator of the galactose-inducible genes in Saccharomyces cerevisiae. Recently, GAL4 has been cloned and its 2.8-kilobase mRNA has been identified. We report here the DNA sequence of GAL4 and the mapping of the 5' and 3' ends of its transcripts. The region sequenced contains a single open reading frame, 881 codons long, which could encode a 99,350-dalton protein. The 5' ends of the GAL4 transcripts fall into two clusters. Transcripts which begin at the upstream cluster would encode the 99,350-dalton protein, whereas those starting at the downstream cluster may result in the synthesis of a shorter, 91,600-dalton protein. The putative GAL4 proteins contain an amino acid sequence near their amino termini which resembles a DNA-binding motif found in bacterial and phage repressors and gene activator proteins.

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Primary structure of the Saccharomyces cerevisiae GAL4 gene

Molecular and Cellular Biology ◽

10.1128/mcb.4.2.260-267.1984 ◽

1984 ◽

Vol 4 (2) ◽

pp. 260-267

Author(s):

A Laughon ◽

R F Gesteland

Keyword(s):

Saccharomyces Cerevisiae ◽

Amino Acid ◽

Amino Acid Sequence ◽

Dna Sequence ◽

Primary Structure ◽

Open Reading Frame ◽

Binding Motif ◽

Reading Frame ◽

Activator Proteins ◽

Inducible Genes

The GAL4 gene encodes a positive regulator of the galactose-inducible genes in Saccharomyces cerevisiae. Recently, GAL4 has been cloned and its 2.8-kilobase mRNA has been identified. We report here the DNA sequence of GAL4 and the mapping of the 5' and 3' ends of its transcripts. The region sequenced contains a single open reading frame, 881 codons long, which could encode a 99,350-dalton protein. The 5' ends of the GAL4 transcripts fall into two clusters. Transcripts which begin at the upstream cluster would encode the 99,350-dalton protein, whereas those starting at the downstream cluster may result in the synthesis of a shorter, 91,600-dalton protein. The putative GAL4 proteins contain an amino acid sequence near their amino termini which resembles a DNA-binding motif found in bacterial and phage repressors and gene activator proteins.

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Tempo and Mode of Ty Element Evolution in Saccharomyces cerevisiae

Genetics ◽

10.1093/genetics/151.4.1341 ◽

1999 ◽

Vol 151 (4) ◽

pp. 1341-1351 ◽

Cited By ~ 2

Author(s):

I King Jordan ◽

John F McDonald

Keyword(s):

Saccharomyces Cerevisiae ◽

Negative Selection ◽

Sequence Variation ◽

Size Variation ◽

Sequence Comparisons ◽

Ty Elements ◽

Ty Element ◽

Amino Acid Sequence Variation ◽

Element Evolution ◽

High Level

Abstract The Saccharomyces cerevisiae genome contains five families of long terminal repeat (LTR) retrotransposons, Ty1–Ty5. The sequencing of the S. cerevisiae genome provides an unprecedented opportunity to examine the patterns of molecular variation existing among the entire genomic complement of Ty retrotransposons. We report the results of an analysis of the nucleotide and amino acid sequence variation within and between the five Ty element families of the S. cerevisiae genome. Our results indicate that individual Ty element families tend to be highly homogenous in both sequence and size variation. Comparisons of within-element 5′ and 3′ LTR sequences indicate that the vast majority of Ty elements have recently transposed. Furthermore, intrafamily Ty sequence comparisons reveal the action of negative selection on Ty element coding sequences. These results taken together suggest that there is a high level of genomic turnover of S. cerevisiae Ty elements, which is presumably in response to selective pressure to escape host-mediated repression and elimination mechanisms.

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GIT1, a Gene Encoding a Novel Transporter for Glycerophosphoinositol in Saccharomyces cerevisiae

Genetics ◽

10.1093/genetics/149.4.1707 ◽

1998 ◽

Vol 149 (4) ◽

pp. 1707-1715 ◽

Cited By ~ 2

Author(s):

J L Patton-Vogt ◽

S A Henry

Keyword(s):

Saccharomyces Cerevisiae ◽

Regulatory Gene ◽

Sugar Transport ◽

Open Reading Frame ◽

Gene Encoding ◽

Reading Frame ◽

Membrane Spanning ◽

Membrane Spanning Domains ◽

Uptake And Metabolism ◽

Cells Cultured

Abstract Phosphatidylinositol catabolism in Saccharomyces cerevisiae cells cultured in media containing inositol results in the release of glycerophosphoinositol (GroPIns) into the medium. As the extracellular concentration of inositol decreases with growth, the released GroPIns is transported back into the cell. Exploiting the ability of the inositol auxotroph, ino1, to use exogenous GroPIns as an inositol source, we have isolated mutants (Git−) defective in the uptake and metabolism of GroPIns. One mutant was found to be affected in the gene encoding the transcription factor, SPT7. Mutants of the positive regulatory gene INO2, but not of its partner, INO4, also have the Git− phenotype. Another mutant was complemented by a single open reading frame (ORF) termed GIT1 (glycerophosphoinositol). This ORF consists of 1556 bp predicted to encode a polypeptide of 518 amino acids and 57.3 kD. The predicted Git1p has similarity to a variety of S. cerevisiae transporters, including a phosphate transporter (Pho84p), and both inositol transporters (Itr1p and Itr2p). Furthermore, Git1p contains a sugar transport motif and 12 potential membrane-spanning domains. Transport assays performed on a git1 mutant together with the above evidence indicate that the GIT1 gene encodes a permease involved in the uptake of GroPIns.

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Sequence and expression of the dCMP deaminase gene (DCD1) of Saccharomyces cerevisiae

Molecular and Cellular Biology ◽

10.1128/mcb.6.5.1711-1721.1986 ◽

1986 ◽

Vol 6 (5) ◽

pp. 1711-1721

Author(s):

E M McIntosh ◽

R H Haynes

Keyword(s):

Saccharomyces Cerevisiae ◽

Amino Acids ◽

Amino Acid ◽

Amino Acid Sequences ◽

Open Reading Frame ◽

Northern Analysis ◽

Hindiii Site ◽

Reading Frame ◽

Hindiii Restriction Site ◽

Cycle Dependent

The dCMP deaminase gene (DCD1) of Saccharomyces cerevisiae has been isolated by screening a Sau3A clone bank for complementation of the dUMP auxotrophy exhibited by dcd1 dmp1 haploids. Plasmid pDC3, containing a 7-kilobase (kb) Sau3A insert, restores dCMP deaminase activity to dcd1 mutants and leads to an average 17.5-fold overproduction of the enzyme in wild-type cells. The complementing activity of the plasmid was localized to a 4.2-kb PvuII restriction fragment within the Sau3A insert. Subcloning experiments demonstrated that a single HindIII restriction site within this fragment lies within the DCD1 gene. Subsequent DNA sequence analysis revealed a 936-nucleotide open reading frame encompassing this HindIII site. Disruption of the open reading frame by integrative transformation led to a loss of enzyme activity and confirmed that this region constitutes the dCMP deaminase gene. Northern analysis indicated that the DCD1 mRNA is a 1.15-kb poly(A)+ transcript. The 5' end of the transcript was mapped by primer extension and appears to exhibit heterogeneous termini. Comparison of the amino acid sequence of the T2 bacteriophage dCMP deaminase with that deduced for the yeast enzyme revealed a limited degree of homology which extends over the entire length of the phage polypeptide (188 amino acids) but is confined to the carboxy-terminal half of the yeast protein (312 amino acids). A potential dTTP-binding site in the yeast and phage enzymes was identified by comparison of homologous regions with the amino acid sequences of a variety of other dTTP-binding enzymes. Despite the role of dCMP deaminase in dTTP biosynthesis, Northern analysis revealed that the DCD1 gene is not subject to the same cell cycle-dependent pattern of transcription recently found for the yeast thymidylate synthetase gene (TMP1).

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Isolation and characterization of PEP5, a gene essential for vacuolar biogenesis in Saccharomyces cerevisiae.

Genetics ◽

10.1093/genetics/125.4.739 ◽

1990 ◽

Vol 125 (4) ◽

pp. 739-752 ◽

Cited By ~ 6

Author(s):

C A Woolford ◽

C K Dixon ◽

M F Manolson ◽

R Wright ◽

E W Jones

Keyword(s):

Saccharomyces Cerevisiae ◽

Molecular Mass ◽

Open Reading Frame ◽

Relative Molecular Mass ◽

Cell Fractionation ◽

Calculated Molecular Mass ◽

Reading Frame ◽

Dna Library ◽

Isolation And Characterization ◽

Genomic Dna Library

Abstract pep5 mutants of Saccharomyces cerevisiae accumulate inactive precursors to the vacuolar hydrolases. The PEP5 gene was isolated from a genomic DNA library by complementation of the pep5-8 mutation. Deletion analysis localized the complementing activity to a 3.3-kb DNA fragment. DNA sequence analysis of the PEP5 gene revealed an open reading frame of 1029 codons with a calculated molecular mass for the encoded protein of 117,403 D. Deletion/disruption of the PEP5 gene did not kill the cells. The resulting strains grow very slowly at 37 degrees. The disruption mutant showed greatly decreased activities of all vacuolar hydrolases examined, including PrA, PrB, CpY, and the repressible alkaline phosphatase. Apparently normal precursors forms of the proteases accumulated in pep5 mutants, as did novel forms of PrB antigen. Antibodies raised to a fusion protein that contained almost half of the PEP5 open reading frame allowed detection by immunoblot of a protein of relative molecular mass 107 kD in extracts prepared from wild-type cells. Cell fractionation showed the PEP5 gene product is enriched in the vacuolar fraction and appears to be a peripheral vacuolar membrane protein.

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Molecular and genetic analysis of the gene encoding the Saccharomyces cerevisiae strand exchange protein Sep1

Molecular and Cellular Biology ◽

10.1128/mcb.11.5.2593-2608.1991 ◽

1991 ◽

Vol 11 (5) ◽

pp. 2593-2608 ◽

Cited By ~ 1

Author(s):

D X Tishkoff ◽

A W Johnson ◽

R D Kolodner

Keyword(s):

Saccharomyces Cerevisiae ◽

Open Reading Frame ◽

Strand Exchange ◽

Wild Type ◽

Homologous Pairing ◽

Gene Encoding ◽

Reading Frame ◽

Cloned Gene ◽

Diploid Cells ◽

Exchange Protein

Vegetatively grown Saccharomyces cerevisiae cells contain an activity that promotes a number of homologous pairing reactions. A major portion of this activity is due to strand exchange protein 1 (Sep1), which was originally purified as a 132,000-Mr species (R. Kolodner, D. H. Evans, and P. T. Morrison, Proc. Natl. Acad. Sci. USA 84:5560-5564, 1987). The gene encoding Sep1 was cloned, and analysis of the cloned gene revealed a 4,587-bp open reading frame capable of encoding a 175,000-Mr protein. The protein encoded by this open reading frame was overproduced and purified and had a relative molecular weight of approximately 160,000. The 160,000-Mr protein was at least as active in promoting homologous pairing as the original 132,000-Mr species, which has been shown to be a fragment of the intact 160,000-Mr Sep1 protein. The SEP1 gene mapped to chromosome VII within 20 kbp of RAD54. Three Tn10LUK insertion mutations in the SEP1 gene were characterized. sep1 mutants grew more slowly than wild-type cells, showed a two- to fivefold decrease in the rate of spontaneous mitotic recombination between his4 heteroalleles, and were delayed in their ability to return to growth after UV or gamma irradiation. Sporulation of sep1/sep1 diploids was defective, as indicated by both a 10- to 40-fold reduction in spore formation and reduced spore viability of approximately 50%. The majority of sep1/sep1 diploid cells arrested in meiosis after commitment to recombination but prior to the meiosis I cell division. Return-to-growth experiments showed that sep1/sep1 his4X/his4B diploids exhibited a five- to sixfold greater meiotic induction of His+ recombinants than did isogenic SEP1/SEP1 strains. sep1/sep1 mutants also showed an increased frequency of exchange between HIS4, LEU2, and MAT and a lack of positive interference between these markers compared with wild-type controls. The interaction between sep1, rad50, and spo13 mutations suggested that SEP1 acts in meiosis in a pathway that is parallel to the RAD50 pathway.

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Meiotic recombination between repeated transposable elements in Saccharomyces cerevisiae

Molecular and Cellular Biology ◽

10.1128/mcb.8.7.2942-2954.1988 ◽

1988 ◽

Vol 8 (7) ◽

pp. 2942-2954

Author(s):

M Kupiec ◽

T D Petes

Keyword(s):

Saccharomyces Cerevisiae ◽

Transposable Elements ◽

Gene Conversion ◽

Meiotic Recombination ◽

Small Region ◽

Ty Elements ◽

Ty Element ◽

Reciprocal Exchange ◽

Gene Conversions ◽

Conversion Tract

We have measured the frequency of meiotic recombination between marked Ty elements in the Saccharomyces cerevisiae genome. These recombination events were usually nonreciprocal (gene conversions) and sometimes involved nonhomologous chromosomes. The frequency of ectopic gene conversion among Ty elements appeared lower than expected on the basis of previous studies of recombination between artificially constructed repeats. The conversion events involved either a subset of the total Ty elements in the genome or the conversion tract was restricted to a small region of the Ty element. In addition, the observed conversion events were very infrequently associated with reciprocal exchange.

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Comparative Biology of Two Natural Variants of the IncQ-2 Family Plasmids, pRAS3.1 and pRAS3.2

Journal of Bacteriology ◽

10.1128/jb.00864-09 ◽

2009 ◽

Vol 191 (20) ◽

pp. 6436-6446 ◽

Cited By ~ 6

Author(s):

Wesley Loftie-Eaton ◽

Douglas E. Rawlings

Keyword(s):

Copy Number ◽

Promoter Region ◽

Plasmid Stability ◽

Inducible Promoter ◽

Open Reading Frame ◽

Comparative Biology ◽

Incompatibility Group ◽

Reading Frame ◽

Repeat Sequences ◽

Natural Variants

ABSTRACT Plasmids pRAS3.1 and pRAS3.2 are two closely related, natural variants of the IncQ-2 plasmid family that have identical plasmid backbones except for two differences. Plasmid pRAS3.1 has five 6-bp repeat sequences in the promoter region of the mobB gene and four 22-bp iterons in its oriV region, whereas pRAS3.2 has only four 6-bp repeats and three 22-bp iterons. Plasmid pRAS3.1 was found to have a higher copy number than pRAS3.2, and we show that the extra 6-bp repeat results in an increase in mobB and downstream mobA/repB expression. Placement of repB (primase) behind an arabinose-inducible promoter in trans resulted in an increase in repB expression and an approximately twofold increase in the copy number of plasmids with identical numbers of 22-bp iterons. The pRAS3 plasmids were shown to have a previously unrecognized toxin-antitoxin plasmid stability module within their replicons. The ability of the pRAS3 plasmids to mobilize the oriT regions of two other plasmids of the IncQ-2 family, pTF-FC2 and pTC-F14, suggested that the mobilization proteins pRAS3 are relaxed and can mobilize oriT regions with substantially different sequences. Plasmids pRAS3.1 and pRAS3.2 were highly incompatible with plasmids pTF-FC2 and pTC-F14, and this incompatibility was removed on inactivation of an open reading frame situated downstream of the mobCDE mobilization genes rather than being due to the 22-bp oriV-associated iterons. We propose that the pRAS3 plasmids represent a third, γ incompatibility group within the IncQ-2 family plasmids.

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