scholarly journals Genetically essential and nonessential alpha-tubulin genes specify functionally interchangeable proteins.

1986 ◽  
Vol 6 (11) ◽  
pp. 3722-3733 ◽  
Author(s):  
P J Schatz ◽  
F Solomon ◽  
D Botstein
Keyword(s):  
The Other ◽  
Null Alleles ◽  
Functional Difference ◽  
Nuclear Movement ◽  
Yeast Saccharomyces Cerevisiae ◽  
Spore Viability ◽  
Alpha Tubulin ◽  
Tubulin Genes ◽  
Essential Components ◽  
Meiotic Spindles

Microtubules in yeast are essential components of the mitotic and meiotic spindles and are essential for nuclear movement during cell division and mating. The relative importance in these processes of the two divergent alpha-tubulin genes of the budding yeast Saccharomyces cerevisiae, TUB1 and TUB3, was examined through the construction of null mutations and by increasing their copy number on chromosomes and on plasmids. Experiments with null alleles of TUB3 showed that TUB3 was not essential for mitosis, meiosis, or mating. Null alleles of TUB3, however, did cause several phenotypes, including hypersensitivity to the antimicrotubule drug benomyl and poor spore viability. On the other hand, the TUB1 gene was essential for growth of normal haploid cells. Even in diploids heterozygous for a TUB1 null allele, several dominant phenotypes were evident, including slow growth and poor sporulation. This functional difference between the two genes is apparently due to different levels of expression, because extra copies of either gene could suppress the defects caused by a null mutation in the other. We conclude that in spite of the 10% divergence between the products of the two genes, there is no essential qualitative functional difference between them.

Genetically essential and nonessential alpha-tubulin genes specify functionally interchangeable proteins

1986 ◽  
Vol 6 (11) ◽  
pp. 3722-3733
Author(s):  
P J Schatz ◽  
F Solomon ◽  
D Botstein
Keyword(s):  
The Other ◽  
Null Alleles ◽  
Functional Difference ◽  
Nuclear Movement ◽  
Yeast Saccharomyces Cerevisiae ◽  
Spore Viability ◽  
Alpha Tubulin ◽  
Tubulin Genes ◽  
Essential Components ◽  
Meiotic Spindles

Microtubules in yeast are essential components of the mitotic and meiotic spindles and are essential for nuclear movement during cell division and mating. The relative importance in these processes of the two divergent alpha-tubulin genes of the budding yeast Saccharomyces cerevisiae, TUB1 and TUB3, was examined through the construction of null mutations and by increasing their copy number on chromosomes and on plasmids. Experiments with null alleles of TUB3 showed that TUB3 was not essential for mitosis, meiosis, or mating. Null alleles of TUB3, however, did cause several phenotypes, including hypersensitivity to the antimicrotubule drug benomyl and poor spore viability. On the other hand, the TUB1 gene was essential for growth of normal haploid cells. Even in diploids heterozygous for a TUB1 null allele, several dominant phenotypes were evident, including slow growth and poor sporulation. This functional difference between the two genes is apparently due to different levels of expression, because extra copies of either gene could suppress the defects caused by a null mutation in the other. We conclude that in spite of the 10% divergence between the products of the two genes, there is no essential qualitative functional difference between them.

Insertions of up to 17 amino acids into a region of alpha-tubulin do not disrupt function in vivo

1987 ◽  
Vol 7 (10) ◽  
pp. 3799-3805
Author(s):  
P J Schatz ◽  
G E Georges ◽  
F Solomon ◽  
D Botstein
Keyword(s):  
Amino Acids ◽  
Variable Region ◽  
Yeast Cells ◽  
Meiotic Spindle ◽  
Nuclear Movement ◽  
Yeast Saccharomyces Cerevisiae ◽  
Alpha Tubulin ◽  
Essential Components ◽  
Altered Proteins

Microtubules in yeasts are essential components of the mitotic and meiotic spindle and are necessary for nuclear movement during cell division and mating. The yeast Saccharomyces cerevisiae has two alpha-tubulin genes, TUB1 and TUB3, either of which alone is sufficient for these processes when present in a high enough copy number. Comparisons of sequences from several species reveals the presence of a variable region near the amino terminus of alpha-tubulin proteins. We perturbed the structure of this region in TUB3 by inserting into it 3, 9, or 17 amino acids and tested the ability of these altered proteins to function as the only alpha-tubulin protein in yeast cells. We found that each of these altered proteins was sufficient on its own for mitotic growth, mating, and methods of yeast. We conclude that this region can tolerate considerable variation without losing any of the highly conserved functions of alpha-tubulin. Our results suggest that variability in this region occurs because it can be tolerated, not because it specifies an important function for the protein.

scholarly journals Insertions of up to 17 amino acids into a region of alpha-tubulin do not disrupt function in vivo.

1987 ◽  
Vol 7 (10) ◽  
pp. 3799-3805 ◽  
Author(s):  
P J Schatz ◽  
G E Georges ◽  
F Solomon ◽  
D Botstein
Keyword(s):  
Amino Acids ◽  
Variable Region ◽  
Yeast Cells ◽  
Meiotic Spindle ◽  
Nuclear Movement ◽  
Yeast Saccharomyces Cerevisiae ◽  
Alpha Tubulin ◽  
Essential Components ◽  
Altered Proteins

Microtubules in yeasts are essential components of the mitotic and meiotic spindle and are necessary for nuclear movement during cell division and mating. The yeast Saccharomyces cerevisiae has two alpha-tubulin genes, TUB1 and TUB3, either of which alone is sufficient for these processes when present in a high enough copy number. Comparisons of sequences from several species reveals the presence of a variable region near the amino terminus of alpha-tubulin proteins. We perturbed the structure of this region in TUB3 by inserting into it 3, 9, or 17 amino acids and tested the ability of these altered proteins to function as the only alpha-tubulin protein in yeast cells. We found that each of these altered proteins was sufficient on its own for mitotic growth, mating, and methods of yeast. We conclude that this region can tolerate considerable variation without losing any of the highly conserved functions of alpha-tubulin. Our results suggest that variability in this region occurs because it can be tolerated, not because it specifies an important function for the protein.

Two functional alpha-tubulin genes of the yeast Saccharomyces cerevisiae encode divergent proteins

1986 ◽  
Vol 6 (11) ◽  
pp. 3711-3721
Author(s):  
P J Schatz ◽  
L Pillus ◽  
P Grisafi ◽  
F Solomon ◽  
D Botstein
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Budding Yeast ◽  
Amino Acid Sequences ◽  
Yeast Cells ◽  
Nucleotide Sequencing ◽  
Cell Fractionation ◽  
Gene Products ◽  
Yeast Saccharomyces Cerevisiae ◽  
Alpha Tubulin ◽  
Tubulin Genes

Two alpha-tubulin genes from the budding yeast Saccharomyces cerevisiae were identified and cloned by cross-species DNA homology. Nucleotide sequencing studies revealed that the two genes, named TUB1 and TUB3, encoded gene products of 447 and 445 amino acids, respectively, that are highly homologous to alpha-tubulins from other species. Comparison of the sequences of the two genes revealed a 19% divergence between the nucleotide sequences and a 10% divergence between the amino acid sequences. Each gene had a single intervening sequence, located at an identical position in codon 9. Cell fractionation studies showed that both gene products were present in yeast microtubules. These two genes, along with the TUB2 beta-tubulin gene, probably encode the entire complement of tubulin in budding yeast cells.

scholarly journals Two functional alpha-tubulin genes of the yeast Saccharomyces cerevisiae encode divergent proteins.

1986 ◽  
Vol 6 (11) ◽  
pp. 3711-3721 ◽  
Author(s):  
P J Schatz ◽  
L Pillus ◽  
P Grisafi ◽  
F Solomon ◽  
D Botstein
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Budding Yeast ◽  
Amino Acid Sequences ◽  
Yeast Cells ◽  
Nucleotide Sequencing ◽  
Cell Fractionation ◽  
Gene Products ◽  
Yeast Saccharomyces Cerevisiae ◽  
Alpha Tubulin ◽  
Tubulin Genes

Two alpha-tubulin genes from the budding yeast Saccharomyces cerevisiae were identified and cloned by cross-species DNA homology. Nucleotide sequencing studies revealed that the two genes, named TUB1 and TUB3, encoded gene products of 447 and 445 amino acids, respectively, that are highly homologous to alpha-tubulins from other species. Comparison of the sequences of the two genes revealed a 19% divergence between the nucleotide sequences and a 10% divergence between the amino acid sequences. Each gene had a single intervening sequence, located at an identical position in codon 9. Cell fractionation studies showed that both gene products were present in yeast microtubules. These two genes, along with the TUB2 beta-tubulin gene, probably encode the entire complement of tubulin in budding yeast cells.

scholarly journals Recombination of Ty elements in yeast can be induced by a double-strand break.

Genetics ◽  
1995 ◽  
Vol 140 (1) ◽  
pp. 67-77 ◽  
Author(s):  
A Parket ◽  
O Inbar ◽  
M Kupiec
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
Strand Break ◽  
Double Strand Break ◽  
The Other ◽  
Direct Repeats ◽  
Yeast Saccharomyces Cerevisiae ◽  
Low Frequencies ◽  
Ty Elements ◽  
Main Family

Abstract The Ty retrotransposons are the main family of dispersed repeated sequences in the yeast Saccharomyces cerevisiae. These elements are flanked by a pair of long terminal direct repeats (LTRs). Previous experiments have shown that Ty elements recombine at low frequencies, despite the fact that they are present in 30 copies per genome. This frequency is not highly increased by treatments that cause DNA damage, such as UV irradiation. In this study, we show that it is possible to increase the recombination level of a genetically marked Ty by creating a double-strand break in it. This break is repaired by two competing mechanisms: one of them leaves a single LTR in place of the Ty, and the other is a gene conversion event in which the marked Ty is replaced by an ectopically located one. In a strain in which the marked Ty has only one LTR, the double-strand break is repaired by conversion. We have also measured the efficiency of repair and monitored the progression of the cells through the cell-cycle. We found that in the presence of a double-strand break in the marked Ty, a proportion of the cells is unable to resume growth.

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

1992 ◽  
Vol 12 (5) ◽  
pp. 2154-2164 ◽  
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.

Tandem arrangement of tubulin genes in the protozoan parasite Leishmania enriettii

1983 ◽  
Vol 3 (6) ◽  
pp. 1070-1076
Author(s):  
S M Landfear ◽  
D McMahon-Pratt ◽  
D F Wirth
Keyword(s):  
Tandem Repeat ◽  
Blot Hybridization ◽  
Protozoan Parasite ◽  
Southern Blot Hybridization ◽  
Beta Tubulin ◽  
Developmentally Regulated ◽  
Alpha Tubulin ◽  
Tubulin Genes ◽  
Southern Blot Hybridization Analysis ◽  
Leishmania Enriettii

The arrangement of developmentally regulated alpha- and beta-tubulin genes has been studied in the parasitic protozoan Leishmania enriettii by using Southern blot hybridization analysis. The alpha-tubulin genes occur in a tandem repeat whose monomeric unit may be represented by a 2-kilobase PstI fragment. Similarly, the beta-tubulin genes probably occur in a separate tandem repeat consisting of approximately 4-kilobase units unlinked to the alpha-tubulin repeats.

A mutation in the alpha 1-tubulin gene of Chlamydomonas reinhardtii confers resistance to anti-microtubule herbicides

1993 ◽  
Vol 106 (1) ◽  
pp. 209-218 ◽  
Author(s):  
S.W. James ◽  
C.D. Silflow ◽  
P. Stroom ◽  
P.A. Lefebvre
Keyword(s):  
Chlamydomonas Reinhardtii ◽  
Resistance Mutation ◽  
In Vitro Translation ◽  
Predicted Amino Acid Sequence ◽  
Two Dimensional ◽  
Wild Type ◽  
Tubulin Gene ◽  
Alpha Tubulin ◽  
Tubulin Genes

A mutation in the alpha 1-tubulin gene of Chlamydomonas reinhardtii was isolated by using the amiprophos-methyl-resistant mutation apm1-18 as a background to select new mutants that showed increased resistance to the drug. The upA12 mutation caused twofold resistance to amiprophos-methyl and oryzalin, and twofold hypersensitivity to the microtubule-stabilizing drug taxol, suggesting that the mutation enhanced microtubule stability. The resistance mutation was semi-dominant and mapped to the same interval on linkage group III as the alpha 1-tubulin gene. Two-dimensional gel immunoblots of proteins in the mutant cells revealed two electrophoretically altered alpha-tubulin isoforms, one of which was acetylated and incorporated into microtubules in the axoneme. The mutant isoforms co-segregated with the drug-resistance phenotypes when mutant upA12 was backcrossed to wild-type cells. Two-dimensional gel analysis of in vitro translation products showed that the non-acetylated variant alpha-tubulin was a primary gene product. DNA sequence analysis of the alpha 1-tubulin genes from mutant and wild-type cells revealed a single missense mutation, which predicted a change in codon 24 from tyrosine in wild type to histidine in mutant upA12. This alteration in the predicted amino acid sequence corroborated the approximately +1 basic charge shift observed for the variant alpha-tubulins. The mutant allele of the alpha 1-tubulin gene was designated tua1-1.

Overexpression of yeast homologs of the mammalian checkpoint gene RCC1 suppresses the class of alpha-tubulin mutations that arrest with excess microtubules.

Genetics ◽  
1994 ◽  
Vol 137 (2) ◽  
pp. 381-392 ◽  
Author(s):  
D Kirkpatrick ◽  
F Solomon
Keyword(s):  
Cell Cycle ◽  
Microtubule Assembly ◽  
Functional Requirements ◽  
Cellular Regulation ◽  
Yeast Saccharomyces Cerevisiae ◽  
Alpha Tubulin ◽  
Class 1 ◽  
Cold Sensitive ◽  
Growth Phenotype ◽  
Cytoplasmic Structures

Abstract Microtubules in eukaryotic cells participate in a variety of nuclear and cytoplasmic structures, reflecting functional requirements and cell cycle position. We are studying the cellular regulation of microtubule assembly and organization in the yeast Saccharomyces cerevisiae. We screened for genes that when overexpressed suppress the growth phenotype of conditional mutants in alpha-tubulin that arrest with excess microtubules at the nonpermissive temperature (class 2 mutations). Here we describe one such suppressing element, called ATS1 (for Alpha Tubulin Suppressor). Overexpression of this gene rescues both the growth and microtubule phenotypes of all class 2 mutations, but not the cold-sensitive mutations that arrest with no microtubules (class 1 mutations). Deletion of ATS1 confers a modest slow growth phenotype which is slightly enhanced in strains containing both a deletion of ATS1 and a class 2 tub 1 mutation. The predicted ATS1 protein contains 333 amino acids and has considerable structural homology to the products of both the mammalian mitotic control gene RCC1 and the S. cerevisiae gene SRM1/PRP20. Overexpression of SRM1/PRP20 also suppresses class 2 mutants. The results suggest that this family of genes may participate in regulatory interactions between microtubules and the cell cycle.

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