scholarly journals Effects of a temperature-sensitiveMinutemutation on gene expression inDrosophila melanogaster

1984 ◽  
Vol 43 (3) ◽  
pp. 257-275 ◽  
Author(s):  
Donald A. R. Sinclair ◽  
Thomas A. Grigliatti ◽  
Thomas C. Kaufman
Keyword(s):  
Gene Expression ◽  
Temperature Shift ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Gene Products ◽  
The Past ◽  
Reciprocal Temperature ◽  
Sex Comb ◽  
Mutant Phenotypes

SUMMARYMinute(M) lesions exhibit a striking propensity for interacting with many different mutations. In the past, few attempts have been made to explain these diverse phenomena. This study describes a variety of temperature-sensitive (ts) interactions exhibited by the ts third chromosomeMinutemutationM(3)LS4Q-III(Q-III). Most of these interactions (i.e. those involvingvg, cp, Dl, DfdorLy) reflectQ-III-induced enhancement of the respective mutant phenotypes at the restrictive temperature. However,Q-IIIalso suppresses the extra-sex-comb phenotypes ofPcandMscat 29 °C and evokes lethal and bristle traits when combined withJ34eat the restrictive temperature. All of these interactions are characteristic of non-tsMinutelesions and thus they appear to be correlated with general physiological perturbations associated with theMsyndrome. In addition, our findings show that mutations that affect ribosome production and/or function, namelysu(f)ts67gandbbts−1, exhibit interactions comparable to those elicited byQ-III. Hence, in accordance with previous findings, we argue that most of theQ-IIIinteractions can be attributed to reduced translational capacity at the restrictive temperature. Finally, reciprocal temperature shift studies were used to delineate TSPs for interactions betweenQ-IIIandvg(mid to late second instar),cp(about mid-third instar),Dfd(early third instar) andDl(late second to mid third instar). We believe that these TSPs represent developmental intervals during which the respective gene products are utilized.

scholarly journals GENETIC CONTROL OF THE CELL DIVISION CYCLE IN YEAST: V. GENETIC ANALYSIS OF cdc MUTANTS

Genetics ◽  
1973 ◽  
Vol 74 (2) ◽  
pp. 267-286
Author(s):  
Leland H Hartwell ◽  
Robert K Mortimer ◽  
Joseph Culotti ◽  
Marilyn Culotti
Keyword(s):  
Cell Cycle ◽  
Cell Division ◽  
Nuclear Gene ◽  
Temperature Shift ◽  
Time Lapse ◽  
Cell Division Cycle ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Gene Products ◽  
Diploid Cells

ABSTRACT One hundred and forty-eight temperature-sensitive cell division cycle (cdc) mutants of Saccharomyces cerevisiae have been isolated and characterized. Complementation studies ordered these recessive mutations into 32 groups and tetrad analysis revealed that each of these groups defines a single nuclear gene. Fourteen of these genes have been located on the yeast genetic map. Functionally related cistrons are not tightly clustered. Mutations in different cistrons frequently produce different cellular and nuclear morphologies in the mutant cells following incubation at the restrictive temperature, but all the mutations in the same cistron produce essentially the same morphology. The products of these genes appear, therefore, each to function individually in a discrete step of the cell cycle and they define collectively a large number of different steps. The mutants were examined by time-lapse photomicroscopy to determine the number of cell cycles completed at the restrictive temperature before arrest. For most mutants, cells early in the cell cycle at the time of the temperature shift (before the execution point) arrest in the first cell cycle while those later in the cycle (after the execution point) arrest in the second cell cycle. Execution points for allelic mutations that exhibit first or second cycle arrest are rather similar and appear to be cistron-specific. Other mutants traverse several cycles before arrest, and its suggested that the latter type of response may reveal gene products that are temperature-sensitive for synthesis, whereas the former may be temperature-sensitive for function. The gene products that are defined by the cdc cistrons are essential for the completion of the cell cycle in haploids of a and α mating type and in a/α diploid cells. The same genes, therefore, control the cell cycle in each of these stages of the life cycle.

scholarly journals Inhibition of Endosome Function in CHO Cells Bearing a Temperature-sensitive Defect in the Coatomer (COPI) Component ε-COP

1997 ◽  
Vol 139 (7) ◽  
pp. 1747-1759 ◽  
Author(s):  
Elizabeth Daro ◽  
David Sheff ◽  
Marie Gomez ◽  
Thomas Kreis ◽  
Ira Mellman
Keyword(s):  
Mammalian Cells ◽  
Secretory Pathway ◽  
Semliki Forest Virus ◽  
Temperature Shift ◽  
Endocytic Pathway ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Intact Cells ◽  
Early Endosomes ◽  
Mutant Phenotypes

Recent evidence has suggested that subunits of the coatomer protein (COPI) complexes are functionally associated with endosomes in mammalian cells. We now provide genetic evidence that COPI plays a role in endocytosis in intact cells. The ldlF mutant CHO cell line bears a temperature-sensitive defect in the COPI subunit ε-COP. In addition to exhibiting conditional defects in the secretory pathway, we find that the cells are also defective at mediating endosome-associated functions. As found for cells microinjected with anti-COPI antibodies, ldlF cells at the restrictive temperature could not be infected by vesicular stomatitis (VSV) or Semliki Forest virus (SFV) that require delivery to acidic endosomes to penetrate into the cytosol. Although there was no temperature-sensitive defect in the internalization of receptor-bound transferrin (Tfn), Tfn recycling and accumulation of HRP were markedly inhibited at the restrictive temperature. Sorting of receptor-bound markers such as EGF to lysosomes was also reduced, although delivery of fluid-phase markers was only partially inhibited. In addition, lysosomes redistributed from their typical perinuclear location to the tips of the ldlF cells. Mutant phenotypes began to emerge within 2 h of temperature shift, the time required for the loss of detectable ε-COP, suggesting that the endocytic defects were not secondary to a block in the secretory pathway. Importantly, the mutant phenotypes were also corrected by transfection of wild-type ε-COP cDNA demonstrating that they directly or indirectly reflected the ε-COP defect. Taken together, the results suggest that ε-COP acts early in the endocytic pathway, most likely inhibiting the normal sorting and recycling functions of early endosomes.

CDC68, a yeast gene that affects regulation of cell proliferation and transcription, encodes a protein with a highly acidic carboxyl terminus

1991 ◽  
Vol 11 (11) ◽  
pp. 5718-5726
Author(s):  
A Rowley ◽  
R A Singer ◽  
G C Johnston
Keyword(s):  
Gene Expression ◽  
Cell Cycle ◽  
Cycle Performance ◽  
Carboxyl Terminus ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Negative Effects ◽  
Yeast Saccharomyces Cerevisiae ◽  
Acid Protein ◽  
Mutant Cells

The cell cycle of the budding yeast Saccharomyces cerevisiae has been investigated through the study of conditional cdc mutations that specifically affect cell cycle performance. Cells bearing the cdc68-1 mutation (J. A. Prendergast, L. E. Murray, A. Rowley, D. R. Carruthers, R. A. Singer, and G. C. Johnston, Genetics 124:81-90, 1990) are temperature sensitive for the performance of the G1 regulatory event, START. Here we describe the CDC68 gene and present evidence that the CDC68 gene product functions in transcription. CDC68 encodes a 1,035-amino-acid protein with a highly acidic and serine-rich carboxyl terminus. The abundance of transcripts from several unrelated genes is decreased in cdc68-1 mutant cells after transfer to the restrictive temperature, while at least one transcript, from the HSP82 gene, persists in an aberrant fashion. Thus, the cdc68-1 mutation has both positive and negative effects on gene expression. Our findings complement those of Malone et al. (E. A. Malone, C. D. Clark, A. Chiang, and F. Winston, Mol. Cell. Biol. 11:5710-5717, 1991), who have independently identified the CDC68 gene (as SPT16) as a transcriptional suppressor of delta-insertion mutations. Among transcripts that rapidly become depleted in cdc68-1 mutant cells are those of the G1 cyclin genes CLN1, CLN2, and CLN3/WHI1/DAF1, whose activity has been previously shown to be required for the performance of START. The decreased abundance of cyclin transcripts in cdc68-1 mutant cells, coupled with the suppression of cdc68-1-mediated START arrest by the CLN2-1 hyperactive allele of CLN2, shows that the CDC68 gene affects START through cyclin gene expression.

scholarly journals GENETIC AND DEVELOPMENTAL ANALYSIS OF A TEMPERATURE-SENSITIVE MINUTE MUTATION OF DROSOPHILA MELANOGASTER

Genetics ◽  
1981 ◽  
Vol 97 (3-4) ◽  
pp. 581-606 ◽  
Author(s):  
Donald A R Sinclair ◽  
David T Suzuki ◽  
Thomas A Grigliatti
Keyword(s):  
Larval Instar ◽  
Temperature Shift ◽  
Heat Pulse ◽  
Sensitive Period ◽  
Temperature Sensitive ◽  
Direct Effects ◽  
First Instar ◽  
Reciprocal Temperature ◽  
Morphological Defects ◽  
Developmental Analysis

ABSTRACT A temperature-sensitive (ts) third chromosome Minute (M) mutation, designated Q-III, has been recovered and characterized. Q-III heterozygotes raised at 29" exhibit all of the dominant traits of M mutants including small bristles, rough eyes, prolonged development, reduced viability 2nd interactions with several unrelated mutations. Q-III homozygotes raised at 29° are lethal; death occurs primarily during the first larval instar. When raised at 22°, Q-Ill heterozygotes are phenotypically normal and Q-III homozygotes display moderate Mtraits. In addition, Q-IIIelicits ts sterility and maternal-effect lethality. As it true of Mlesions, the dominant traits of Q-111 are not expressed in triploid females raised at 29°. Complementation tests suggest that Q-III is a ts allele of M(3)LS4, which is located in 3L near the centromere.——Reciprocal temperature-shift experiments revealed that the temperature-sensitive period (TSP) of Q-111 lethality is polyphasic, extending from the first instar to the latter half of pupation. Heat-pulse experiments further resolved this into two post-embryonic TSPs: one occurring during the latter half of the second larval instar, and the other extending from the larval/pupal boundary to the second half of pupation. In addition, heat pulses elicited a large number of striking adult phenotypes in Q-III individuals. These included pattern alterations such as deficiencies and duplications and cther morphological defects in structures produced by the eye-antennal, leg, wing and genital imaginal discs and the abdominal histoblasts. Each defect or pattern alteration is associated with a specific TSP during development.——We favor the interpretation that most of the major Q-III defects, particularly the structural duplications and deficiencies, result from temperature-induced cell death in mitotically active imaginal anlagen, while the small macrochaete phene probably results from the direct effects of Q-III on bristle synthesis. The hypothesis that the Q-III locus specifices a component required for protein synthesis is discussed, and it is concluded that this hypothesis can account for the pleiotropy of Q-III, and that perhaps it can be extended to M loci in general.

scholarly journals Diverse natural variants suppress mutations in hundreds of essential genes

2020 ◽  
Author(s):  
Leopold Parts ◽  
Amandine Batté ◽  
Maykel Lopes ◽  
Michael W. Yuen ◽  
Meredith Laver ◽  
...  
Keyword(s):  
Wild Strain ◽  
Essential Genes ◽  
Temperature Sensitive ◽  
Gene Products ◽  
Fitness Effects ◽  
Monogenic Diseases ◽  
Interaction Partners ◽  
Causal Genes ◽  
Natural Variants ◽  
Mutant Phenotypes

AbstractThe consequence of a mutation can be influenced by the context in which it operates. For example, loss of gene function may be tolerated in one genetic background, but lead to lethality in another. The extent to which mutant phenotypes are malleable, the complexity of the architecture of modifiers, and the identities of causal genes and pathways remain largely unknown. Here, we measure the fitness effects of ~1,500 temperature sensitive alleles of yeast essential genes in the context of variation from ten different natural genetic backgrounds, and map the modifiers for 19 combinations. Altogether, fitness defects for 183 of the 530 tested genes (35%) could be suppressed by standing genetic variation in at least one wild strain. Suppression was generally driven by gain-of-function of a single, strong modifier gene. The validated causes included both variants in protein interaction partners or pathway members suppressing specific genes, as well as general modifiers altering the effect of many temperature sensitive alleles. The emerging frequency of suppression and range of possible suppression mechanisms suggest that a substantial fraction of monogenic diseases could be repressed by modulating other gene products.

scholarly journals DETERMINATION OF THE ORDER OF GENE FUNCTION IN THE YEAST NUCLEAR DIVISION PATHWAY USING cs AND ts MUTANTS

Genetics ◽  
1982 ◽  
Vol 100 (4) ◽  
pp. 565-577
Author(s):  
Don Moir ◽  
David Botstein
Keyword(s):  
Gene Function ◽  
Nuclear Division ◽  
Temperature Shift ◽  
Yeast Saccharomyces Cerevisiae ◽  
Lethal Mutations ◽  
Reciprocal Temperature ◽  
Cold Sensitive ◽  
Cell Cycle Pathway ◽  
Mutant Phenotypes

ABSTRACT Cold-sensitive (cs) and heat-sensitive (ts) conditional-lethal mutations that affect specifically the cell division cycle of budding yeast (Saccharomyces cerevisiae) were used to determine the order of gene function. Reciprocal temperature-shift experiments using cs-ts double mutants revealed a detailed order of function among genes whose execution points and mutant phenotypes are very similar. The data suggest that the nuclear branch of the overall cell-cycle pathway itself contains at least one branch.

scholarly journals An rne-1 pnp-7 Double Mutation Suppresses the Temperature-Sensitive Defect of lacZ Gene Expression in a divE Mutant

1998 ◽  
Vol 180 (6) ◽  
pp. 1389-1395 ◽  
Author(s):  
Toshiko Aiso ◽  
Reiko Ohki
Keyword(s):  
Gene Expression ◽  
Escherichia Coli ◽  
Protein Synthesis ◽  
Temperature Shift ◽  
Mrna Degradation ◽  
Northern Hybridization ◽  
Temperature Sensitive ◽  
Degradation Pathways ◽  
Lacz Gene ◽  
Double Mutation

ABSTRACT A divE mutant, which has a temperature-sensitive mutation in the tRNA1 Ser gene, exhibits differential loss of the synthesis of certain proteins, such as β-galactosidase and succinate dehydrogenase, at nonpermissive temperatures. In Escherichia coli, the UCA codon is recognized only by tRNA1 Ser. Several genes containing UCA codons are normally expressed after a temperature shift to 42°C in the divE mutant. Therefore, it is unlikely that the defect in protein synthesis at 42°C is simply caused by a defect in the decoding function of the mutant tRNA1 Ser. In this study, we sought to determine the cause of the defect in lacZ gene expression in thedivE mutant. It has also been shown that the defect inlacZ gene expression is accompanied by a decrease in the amount of lacZ mRNA. To examine whether inactivation of mRNA degradation pathways restores the defect in lacZ gene expression, we constructed divE mutants containingrne-1, rnb-500, and pnp-7 mutations in various combinations. We found that the defect was almost completely restored by introducing an rne-1 pnp-7 double mutation into the divE mutant. Northern hybridization analysis showed that the rne-1 mutation stabilized lacZ mRNA, whereas the pnp-7 mutation stabilized mutant tRNA1 Ser, at 44°C. We present a mechanism that may explain these results.

scholarly journals Incubation at the nonpermissive temperature induces deficiencies in UV resistance and mutagenesis in mouse mutant cells expressing a temperature-sensitive ubiquitin-activating enzyme (E1).

1997 ◽  
Vol 17 (3) ◽  
pp. 1484-1489 ◽  
Author(s):  
H Ikehata ◽  
S Kaneda ◽  
F Yamao ◽  
T Seno ◽  
T Ono ◽  
...  
Keyword(s):  
Dna Repair ◽  
Mammalian Cells ◽  
Uv Light ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Nonpermissive Temperature ◽  
Conjugation System ◽  
Ubiquitin Conjugating Enzyme ◽  
Mutant Phenotypes ◽  
Mutant Cells

In temperature-sensitive (ts) mutants of mouse FM3A cells, the levels of mutagenesis and survival of cells treated with DNA-damaging agents have been difficult to assess because they are killed after their mutant phenotypes are expressed at the nonpermissive temperature. To avoid this difficulty, we incubated the ts mutant cells at the restrictive temperature, 39 degrees C, for only a limited period after inducing DNA damage. We used ts mutants defective in genes for ubiquitin-activating enzyme (E1), DNA polymerase alpha, and p34(cdc2) kinase. Whereas the latter two showed no effect, E1 mutants were sensitized remarkably to UV light if incubated at 39 degrees C for limited periods after UV exposure. Eighty-five percent of the sensitization occurred within the first 12 h of incubation at 39 degrees C, and more than 36 h at 39 degrees C did not produce any further sensitization. Moreover, while the 39 degrees C incubation gave E1 mutants a moderate spontaneous mutator phenotype, the same treatment significantly diminished the level of UV-induced 6-thioguanine resistance mutagenesis and extended the time necessary for expression of the mutation phenotype. These characteristics of E1 mutants are reminiscent of the defective DNA repair phenotypes of Saccharomyces cerevisiae rad6 mutants, which have defects in a ubiquitin-conjugating enzyme (E2), to which E1 is known to transfer ubiquitin. These results demonstrate the involvement of E1 in eukaryotic DNA repair and mutagenesis and provide the first direct evidence that the ubiquitin-conjugation system contributes to DNA repair in mammalian cells.

scholarly journals Temperature Sensitivity Conferred byligAAlleles from Psychrophilic Bacteria upon Substitution in Mesophilic Bacteria and a Yeast Species

2016 ◽  
Vol 82 (6) ◽  
pp. 1924-1932 ◽  
Author(s):  
Jarosław A. Pankowski ◽  
Stephanie M. Puckett ◽  
Francis E. Nano
Keyword(s):  
Yeast Species ◽  
Essential Gene ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Gene Products ◽  
Psychrophilic Bacteria ◽  
Content Type ◽  
Francisella Novicida ◽  
Yeast Viability ◽  
Eukaryotic Microbes

ABSTRACTWe have assembled a collection of 13 psychrophilicligAalleles that can serve as genetic elements for engineering mesophiles to a temperature-sensitive (TS) phenotype. When theseligAalleles were substituted intoFrancisella novicida, they conferred a TS phenotype with restrictive temperatures between 33 and 39°C. When theF. novicidaligAhybrid strains were plated above their restrictive temperatures, eight of them generated temperature-resistant variants. For two alleles, the mutations that led to temperature resistance clustered near the 5′ end of the gene, and the mutations increased the predicted strength of the ribosome binding site at least 3-fold. FourF. novicida ligAhybrid strains generated no temperature-resistant variants at a detectable level. These results suggest that multiple mutations are needed to create temperature-resistant variants of theseligAgene products. OneligAallele was isolated from aColwelliaspecies that has a maximal growth temperature of 12°C, and this allele supported growth ofF. novicidaonly as a hybrid between the psychrophilic and theF. novicidaligAgenes. However, the full psychrophilic gene alone supported the growth ofSalmonella enterica, imparting a restrictive temperature of 27°C. We also tested twoligAalleles from twoPseudoalteromonasstrains for their ability to support the viability of aSaccharomyces cerevisiaestrain that lacked its essential gene,CDC9, encoding an ATP-dependent DNA ligase. In both cases, the psychrophilic bacterial alleles supported yeast viability and their expression generated TS phenotypes. This collection ofligAalleles should be useful in engineering bacteria, and possibly eukaryotic microbes, to predictable TS phenotypes.

scholarly journals GENETIC ANALYSIS OF TWO ALLELIC TEMPERATURE-SENSITIVE MUTANTS OF DROSOPHILA MELANOGASTER BOTH OF WHICH ARE ZYGOTIC AND MATERNAL-EFFECT LETHALS

Genetics ◽  
1978 ◽  
Vol 89 (2) ◽  
pp. 341-353
Author(s):  
Allen Shearn ◽  
Grafton Hersperger ◽  
Evelyn Hersperger
Keyword(s):  
Drosophila Melanogaster ◽  
Maternal Effect ◽  
Larval Instar ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Gene Products ◽  
Larval Stages ◽  
Temperature Sensitive Mutants ◽  
Small Disc ◽  
Disc Cells

ABSTRACT After fertilization, the development of a zygote depends upon both gene products synthesized by its maternal parent and gene products synthesized by the zygote itself. To analyze genetically the relative contributions of these two sources of gene products, several laboratories have been isolating two classes of mutants of Drosophila melanogaster: maternal-effect lethals and zygotic lethals. This report concerns the analysis of two temperature-sensitive mutants, OX736hs and PC025hs, which were isolated as alleles of a small-disc mutant, l(3)1902. These alleles are not only zygotic lethals, but also maternal-effect lethals. They have temperature-sensitive periods during larval life and during oogenesis. Mutant larvae exposed continuously to restrictive temperature have small discs. One- or two-day exposures to the restrictive temperature administered during the third larval instar lead to a homeotic transformation of the midlegs and hindlegs to the pattern characteristic of the forelegs. Mutant females exposed to the restrictive temperature during oogenesis produce eggs that can develop until gastrulation, but do not hatch. —The existence of these mutants, and one that was recently described by another group, implies that there may be a class of genes, heretofore unrecognized, whose products are synthesized during oogenesis, are essential for embryogenesis and are also synthesized during larval stages within imaginal disc cells.

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