A suppressor of temperature-sensitive rna mutations that affect mRNA metabolism in Saccharomyces cerevisiae.

We have isolated a dominant suppressor of rna mutation (SRN1) that relieves the temperature-sensitive inhibition of mRNA synthesis of ribosomal protein genes in the yeast Saccharomyces cerevisiae. The suppressor was selected for its ability to alleviate simultaneously the temperature-sensitive growth phenotypes of rna2 and rna6. Several independently isolated suppressors appeared to be recessive lethal mutations. One suppressor, SRN1, was recovered as viable in haploid strains. SRN1 can suppress rna2, rna3, rna4, rna5, rna6, and rna8 singly or in pairs, although some combinations of rna mutations are less well suppressed than others. The suppressor allows strains with rna mutations to grow at 34 degrees C but is unable to suppress at 37 degrees C; however, SRN1 does not, by itself, prevent growth at 37 degrees C. In addition, SRN1 suppresses the rna1 mutation which affects general mRNA levels and also leads to the accumulation of precursor tRNA for those tRNAs that have intervening sequences. SRN1 can suppress the rna1 mutation as well as the rna1 rna2 double mutation at 34 degrees C. The suppressor does not affect the temperature-sensitive growth of two unrelated temperature-sensitive mutations, cdc4 and cdc7.

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Sporulation and rna2 lower ribosomal protein mRNA levels by different mechanisms in Saccharomyces cerevisiae.

Molecular and Cellular Biology ◽

10.1128/mcb.2.10.1199 ◽

1982 ◽

Vol 2 (10) ◽

pp. 1199-1204 ◽

Cited By ~ 11

Author(s):

E Kraig ◽

J E Haber ◽

M Rosbash

Keyword(s):

Saccharomyces Cerevisiae ◽

Ribosomal Protein ◽

Ribosomal Proteins ◽

Relative Rate ◽

Synthesis Rate ◽

Temperature Sensitive ◽

Restrictive Temperature ◽

Mrna Levels ◽

Ribosomal Protein Genes ◽

Mrna Metabolism

In Saccharomyces cerevisiae, the levels of ribosomal protein mRNAs are regulated coordinately. Vegetative strains carrying the temperature-sensitive rna2 mutation exhibit a dramatic decrease in the levels of most ribosomal protein mRNAs at the restrictive temperature. Similarly, in wild-type cells induced to sporulate by nitrogen starvation, there is a fivefold reduction in the relative synthesis rate of ribosomal proteins. Using Northern gel analysis and cloned ribosomal protein genes, we compared the way in which ribosomal protein mRNA is affected under these two conditions. In vegetative rna2 cells, incubation at 34 degrees C led to the disappearance of ribosomal protein mRNAs and the accumulation of higher-molecular-weight precursor RNAs. A different phenotype was observed during sporulation. Although sporulating conditions led to a significant reduction in the relative abundance of ribosomal protein mRNA, there was no detectable accumulation of precursor RNAs even in rna2/rna2 diploids at 34 degrees C. A suppressor of rna2 and of other rna mutations, SRN1, at least partially relieved the block in the splicing of the ribosomal protein 51 intron in vegetative rna2 cells but did not detectably affect the level of ribosomal protein mRNA in sporulating cells. We concluded that the rna2 mutation and sporulation conditions affected ribosomal protein mRNA metabolism in two quite different ways. In vegetative cells the mutant rna2 effected a block which occurred primarily in post-transcriptional processing, whereas in sporulating cells the ribosomal protein mRNA levels were decreased by some other mechanism, presumably a change in the relative rate of transcription or mRNA turnover. Furthermore, the data suggest that the mutation rna2 has no additional effect on ribosomal protein mRNA metabolism in sporulating cells.

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Sporulation and rna2 lower ribosomal protein mRNA levels by different mechanisms in Saccharomyces cerevisiae

Molecular and Cellular Biology ◽

10.1128/mcb.2.10.1199-1204.1982 ◽

1982 ◽

Vol 2 (10) ◽

pp. 1199-1204

Author(s):

E Kraig ◽

J E Haber ◽

M Rosbash

Keyword(s):

Saccharomyces Cerevisiae ◽

Ribosomal Protein ◽

Ribosomal Proteins ◽

Relative Rate ◽

Synthesis Rate ◽

Temperature Sensitive ◽

Restrictive Temperature ◽

Mrna Levels ◽

Ribosomal Protein Genes ◽

Mrna Metabolism

In Saccharomyces cerevisiae, the levels of ribosomal protein mRNAs are regulated coordinately. Vegetative strains carrying the temperature-sensitive rna2 mutation exhibit a dramatic decrease in the levels of most ribosomal protein mRNAs at the restrictive temperature. Similarly, in wild-type cells induced to sporulate by nitrogen starvation, there is a fivefold reduction in the relative synthesis rate of ribosomal proteins. Using Northern gel analysis and cloned ribosomal protein genes, we compared the way in which ribosomal protein mRNA is affected under these two conditions. In vegetative rna2 cells, incubation at 34 degrees C led to the disappearance of ribosomal protein mRNAs and the accumulation of higher-molecular-weight precursor RNAs. A different phenotype was observed during sporulation. Although sporulating conditions led to a significant reduction in the relative abundance of ribosomal protein mRNA, there was no detectable accumulation of precursor RNAs even in rna2/rna2 diploids at 34 degrees C. A suppressor of rna2 and of other rna mutations, SRN1, at least partially relieved the block in the splicing of the ribosomal protein 51 intron in vegetative rna2 cells but did not detectably affect the level of ribosomal protein mRNA in sporulating cells. We concluded that the rna2 mutation and sporulation conditions affected ribosomal protein mRNA metabolism in two quite different ways. In vegetative cells the mutant rna2 effected a block which occurred primarily in post-transcriptional processing, whereas in sporulating cells the ribosomal protein mRNA levels were decreased by some other mechanism, presumably a change in the relative rate of transcription or mRNA turnover. Furthermore, the data suggest that the mutation rna2 has no additional effect on ribosomal protein mRNA metabolism in sporulating cells.

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SEN1, a positive effector of tRNA-splicing endonuclease in Saccharomyces cerevisiae.

Molecular and Cellular Biology ◽

10.1128/mcb.12.5.2154 ◽

1992 ◽

Vol 12 (5) ◽

pp. 2154-2164 ◽

Cited By ~ 50

Author(s):

D J DeMarini ◽

M Winey ◽

D Ursic ◽

F Webb ◽

M R Culbertson

Keyword(s):

Saccharomyces Cerevisiae ◽

Amino Acids ◽

Amino Acid ◽

Gene Product ◽

Signal Sequence ◽

Null Alleles ◽

Nucleoside Triphosphate ◽

Temperature Sensitive ◽

Yeast Saccharomyces Cerevisiae ◽

Amino Terminal

The SEN1 gene, which is essential for growth in the yeast Saccharomyces cerevisiae, is required for endonucleolytic cleavage of introns from all 10 families of precursor tRNAs. A mutation in SEN1 conferring temperature-sensitive lethality also causes in vivo accumulation of pre-tRNAs and a deficiency of in vitro endonuclease activity. Biochemical evidence suggests that the gene product may be one of several components of a nuclear-localized splicing complex. We have cloned the SEN1 gene and characterized the SEN1 mRNA, the SEN1 gene product, the temperature-sensitive sen1-1 mutation, and three SEN1 null alleles. The SEN1 gene corresponds to a 6,336-bp open reading frame coding for a 2,112-amino-acid protein (molecular mass, 239 kDa). Using antisera directed against the C-terminal end of SEN1, we detect a protein corresponding to the predicted molecular weight of SEN1. The SEN1 protein contains a leucine zipper motif, consensus elements for nucleoside triphosphate binding, and a potential nuclear localization signal sequence. The carboxy-terminal 1,214 amino acids of the SEN1 protein are essential for growth, whereas the amino-terminal 898 amino acids are dispensable. A sequence of approximately 500 amino acids located in the essential region of SEN1 has significant similarity to the yeast UPF1 gene product, which is involved in mRNA turnover, and the mouse Mov-10 gene product, whose function is unknown. The mutation that creates the temperature-sensitive sen1-1 allele is located within this 500-amino-acid region, and it causes a substitution for an amino acid that is conserved in all three proteins.

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A presumptive helicase (MOT1 gene product) affects gene expression and is required for viability in the yeast Saccharomyces cerevisiae.

Molecular and Cellular Biology ◽

10.1128/mcb.12.4.1879 ◽

1992 ◽

Vol 12 (4) ◽

pp. 1879-1892 ◽

Cited By ~ 98

Author(s):

J L Davis ◽

R Kunisawa ◽

J Thorner

Keyword(s):

Saccharomyces Cerevisiae ◽

Gene Product ◽

Essential Gene ◽

Single Gene ◽

Temperature Sensitive ◽

Yeast Saccharomyces Cerevisiae ◽

Sequence Element ◽

Cis Acting ◽

Upstream Activating Sequence ◽

Identical Manner

Exposure of a haploid yeast cell to mating pheromone induces transcription of a set of genes. Induction is mediated through a cis-acting DNA sequence found upstream of all pheromone-responsive genes. Although the STE12 gene product binds specifically to this sequence element and is required for maximum levels of both basal and induced transcription, not all pheromone-responsive genes are regulated in an identical manner. To investigate whether additional factors may play a role in transcription of these genes, a genetic screen was used to identify mutants able to express pheromone-responsive genes constitutively in the absence of Ste12. In this way, we identified a recessive, single gene mutation (mot1, for modifier of transcription) which increases the basal level of expression of several, but not all, pheromone-responsive genes. The mot1-1 allele also relaxes the requirement for at least one other class of upstream activating sequence and enhances the expression of another gene not previously thought to be involved in the mating pathway. Cells carrying mot1-1 grow slowly at 30 degrees C and are inviable at 38 degrees C. The MOT1 gene was cloned by complementation of this temperature-sensitive lethality. Construction of a null allele confirmed that MOT1 is an essential gene. MOT1 residues on chromosome XVI and encodes a large protein of 1,867 amino acids which contains all seven of the conserved domains found in known and putative helicases. The product of MOT1 is strikingly homologous to the Saccharomyces cerevisiae SNF2/SW12 and RAD54 gene products over the entire helicase region.

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Regulation of STA1 gene expression by MAT during the life cycle of Saccharomyces cerevisiae

Molecular and Cellular Biology ◽

10.1128/mcb.9.9.3992-3998.1989 ◽

1989 ◽

Vol 9 (9) ◽

pp. 3992-3998

Author(s):

A M Dranginis

Keyword(s):

Saccharomyces Cerevisiae ◽

Mating Type ◽

Yeast Cells ◽

Specific Gene ◽

Activity Levels ◽

Sporulation Medium ◽

Mrna Levels ◽

Yeast Saccharomyces Cerevisiae ◽

Type Locus ◽

Diploid Cells

STA1 encodes a secreted glucoamylase of the yeast Saccharomyces cerevisiae var. diastaticus. Glucoamylase secretion is controlled by the mating type locus MAT; a and alpha haploid yeast cells secrete high levels of the enzyme, but a/alpha diploid cells produce undetectable amounts. It has been suggested that STA1 is regulated by MATa2 (I. Yamashita, Y. Takano, and S. Fukui, J. Bacteriol. 164:769-773, 1985), which is a MAT transcript of previously unknown function. In contrast, this work shows that deletion of the entire MATa2 gene had no effect on STA1 regulation but that deletion of MATa1 sequences completely abolished mating-type control. In all cases, glucoamylase activity levels reflected STA1 mRNA levels. It appears that STA1 is a haploid-specific gene that is regulated by MATa1 and a product of the MAT alpha locus and that this regulation occurs at the level of RNA accumulation. STA1 expression was also shown to be glucose repressible. STA1 mRNA was induced in diploids during sporulation along with SGA, a closely linked gene that encodes an intracellular sporulation-specific glucoamylase of S. cerevisiae. A diploid strain with a MATa1 deletion showed normal induction of STA1 in sporulation medium, but SGA expression was abolished. Therefore, these two homologous and closely linked glucoamylase genes are induced by different mechanisms during sporulation. STA1 induction may be a response to the starvation conditions necessary for sporulation, while SGA induction is governed by the pathway by which MAT regulates sporulation. The strain containing a complete deletion of MATa2 grew, mated, and sporulated normally.

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Temperature sensitive pet mutants in yeast Saccharomyces cerevisiae that lose mitochondrial RNA

Current Genetics ◽

10.1007/bf00378178 ◽

1987 ◽

Vol 11 (5) ◽

pp. 359-367 ◽

Cited By ~ 13

Author(s):

David M. Mueller ◽

Tapan K. Biswas ◽

James Backer ◽

John C. Edwards ◽

Murray Rabinowitz ◽

...

Keyword(s):

Saccharomyces Cerevisiae ◽

Temperature Sensitive ◽

Mitochondrial Rna ◽

Yeast Saccharomyces Cerevisiae ◽

Pet Mutants

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Purification of profilin from Saccharomyces cerevisiae and analysis of profilin-deficient cells.

The Journal of Cell Biology ◽

10.1083/jcb.110.1.105 ◽

1990 ◽

Vol 110 (1) ◽

pp. 105-114 ◽

Cited By ~ 153

Author(s):

B K Haarer ◽

S H Lillie ◽

A E Adams ◽

V Magdolen ◽

W Bandlow ◽

...

Keyword(s):

Saccharomyces Cerevisiae ◽

Cell Wall ◽

Actin Polymerization ◽

Yeast Cells ◽

Predicted Amino Acid Sequence ◽

Temperature Sensitive ◽

Wild Type ◽

Yeast Saccharomyces Cerevisiae ◽

Ellipsoidal Shape ◽

Hydrolysis Of

We have isolated profilin from yeast (Saccharomyces cerevisiae) and have microsequenced a portion of the protein to confirm its identity; the region microsequenced agrees with the predicted amino acid sequence from a profilin gene recently isolated from S. cerevisiae (Magdolen, V., U. Oechsner, G. Müller, and W. Bandlow. 1988. Mol. Cell. Biol. 8:5108-5115). Yeast profilin resembles profilins from other organisms in molecular mass and in the ability to bind to polyproline, retard the rate of actin polymerization, and inhibit hydrolysis of ATP by monomeric actin. Using strains that carry disruptions or deletions of the profilin gene, we have found that, under appropriate conditions, cells can survive without detectable profilin. Such cells grow slowly, are temperature sensitive, lose the normal ellipsoidal shape of yeast cells, often become multinucleate, and generally grow much larger than wild-type cells. In addition, these cells exhibit delocalized deposition of cell wall chitin and have dramatically altered actin distributions.

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Replication slippage between distant short repeats in Saccharomyces cerevisiae depends on the direction of replication and the RAD50 and RAD52 genes.

Molecular and Cellular Biology ◽

10.1128/mcb.15.10.5607 ◽

1995 ◽

Vol 15 (10) ◽

pp. 5607-5617 ◽

Cited By ~ 66

Author(s):

H T Tran ◽

N P Degtyareva ◽

N N Koloteva ◽

A Sugino ◽

H Masumoto ◽

...

Keyword(s):

Saccharomyces Cerevisiae ◽

Genome Stability ◽

Genome Instability ◽

Temperature Sensitive ◽

Direct Repeats ◽

Dna Polymerase Delta ◽

Dna Double Strand Breaks ◽

Yeast Saccharomyces Cerevisiae ◽

Replication Slippage ◽

Random Sequences

Small direct repeats, which are frequent in all genomes, are a potential source of genome instability. To study the occurrence and genetic control of repeat-associated deletions, we developed a system in the yeast Saccharomyces cerevisiae that was based on small direct repeats separated by either random sequences or inverted repeats. Deletions were examined in the LYS2 gene, using a set of 31- to 156-bp inserts that included inserts with no apparent potential for secondary structure as well as two quasipalindromes. All inserts were flanked by 6- to 9-bp direct repeats of LYS2 sequence, providing an opportunity for Lys+ reversion via precise excision. Reversions could arise by extended deletions involving either direct repeats or random sequences and by -1-or +2-bp frameshift mutations. The deletion breakpoints were always associated with short (3- to 9-bp) perfect or imperfect direct repeats. Compared with the POL+ strain, deletions between small direct repeats were increased as much as 100-fold, and the spectrum was changed in a temperature-sensitive DNA polymerase delta pol3-t mutant, suggesting a role for replication. The type of deletion depended on orientation relative to the origin of replication. On the basis of these results, we propose (i) that extended deletions between small repeats arise by replication slippage and (ii) that the deletions occur primarily in either the leading or lagging strand. The RAD50 and RAD52 genes, which are required for the recombinational repair of many kinds of DNA double-strand breaks, appeared to be required also for the production of up to 90% of the deletions arising between separated repeats in the pol3-t mutant, suggesting a newly identified role for these genes in genome stability and possibly replication.

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Cloning and characterization of DST2, the gene for DNA strand transfer protein beta from Saccharomyces cerevisiae.

Molecular and Cellular Biology ◽

10.1128/mcb.11.5.2583 ◽

1991 ◽

Vol 11 (5) ◽

pp. 2583-2592 ◽

Cited By ~ 57

Author(s):

C C Dykstra ◽

K Kitada ◽

A B Clark ◽

R K Hamatake ◽

A Sugino

Keyword(s):

Saccharomyces Cerevisiae ◽

Intragenic Recombination ◽

Temperature Sensitive ◽

Strand Transfer ◽

Gene Encoding ◽

Yeast Saccharomyces Cerevisiae ◽

Prolonged Incubation ◽

Transfer Protein ◽

Dna Strand

The gene encoding the 180-kDa DNA strand transfer protein beta from the yeast Saccharomyces cerevisiae was identified and sequenced. This gene, DST2 (DNA strand transferase 2), was located on chromosome VII. dst2 gene disruption mutants exhibited temperature-sensitive sporulation and a 50% longer generation time during vegetative growth than did the wild type. Spontaneous mitotic recombination in the mutants was reduced severalfold for both intrachromosomal recombination and intragenic gene conversion. The mutants also had reduced levels of the intragenic recombination that is induced during meiosis. Meiotic recombinants were, however, somewhat unstable in the mutants, with a decrease in recombinants and survival upon prolonged incubation in sporulation media. spo13 or spo13 rad50 mutations did not relieve the sporulation defect of dst2 mutations. A dst1 dst2 double mutant has the same phenotype as a dst2 single mutant. All phenotypes associated with the dst2 mutations could be complemented by a plasmid containing DST2.

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