scholarly journals MUTATIONS AFFECTING CELL DIVISION IN TETRAHYMENA PYRIFORMIS. I. SELECTION AND GENETIC ANALYSIS

Genetics ◽  
1976 ◽  
Vol 83 (3) ◽  
pp. 489-506
Author(s):  
Joseph Frankel ◽  
Leslie M Jenkins ◽  
F Paul Doerder ◽  
E Marlo Nelsen
Keyword(s):  
Cell Division ◽  
Genetic Analysis ◽  
Single Gene ◽  
Specific Effect ◽  
Tetrahymena Pyriformis ◽  
Recessive Mutation ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Induced Mutations ◽  
Mendelian Segregation

ABSTRACT Fourteen nitrosoguanidine-induced mutations that bring about temperature-sensitive morphological abnormalities resulting from a specific effect on cell division have been isolated as heterozygous phenotypic assortants in Tetrahymena pyriformis syngen 1. Genetic analysis revealed all to be single-gene recessives. Detailed analysis of the kinetics of assortment for one of the mutated alleles revealed a rate (0.0104 pure lines per fission) consistent with that previously observed at other loci in this organism. The mutations fall into six complementation groups (mo1, mo2, mo3, mo6, mo8, and mo12). Homozygotes of mo2 are unconditionally expressed, while all alleles of mo1, mo6, mo8, and mo12 are heat sensitive for division arrest. At the mo3 locus two alleles are heat sensitive, one is primarily cold sensitive, while two are sensitive to both heat and cold. Two out of three combinations of different mo3alleles show conventional Mendelian segregation of conditions of expression. Different alleles of mo1, mo3, mo8, and mo12 also manifest differences in penetrance at the restrictive temperature. Despite these differences involving expression, the abnormal phenotypes themselves are locus-specific and distinctive; in the one case (mo1a and mo1b) in which two alleles manifest somewhat different phenotypes, the F1 between them is intermediate. One additional recessive mutation (fat1) brings about a nonconditional lengthening of the cell cycle, with some arrest of cell division at the restrictive temperature. These findings demonstrate that selection of heterozygotes undergoing phenotypic assortment can be an effective method for obtaining substantial numbers of a desired class of temperature-sensitive mutations in T. pyriformis.

scholarly journals THE SELECTION OF S. CEREVISIAE MUTANTS DEFECTIVE IN THE START EVENT OF CELL DIVISION

Genetics ◽  
1980 ◽  
Vol 95 (3) ◽  
pp. 561-577 ◽  
Author(s):  
Steven I Reed
Keyword(s):  
Cell Division ◽  
Genetic Analysis ◽  
Linkage Map ◽  
Nuclear Genes ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Mating Pheromone ◽  
Preliminary Characterization ◽  
Complementation Groups ◽  
Selection Of

ABSTRACT Thirty-three temperature-sensitive mutations defective in the start event of the cell division cycle of Saccharomyces cereuisiae were isolated and subjected to preliminary characterization. Complementation studies assigned these mutations to four complementation groups, one of which, cdc28, has been described previously. Genetic analysis revealed that these complementation groups define single nuclear genes, unlinked to one another. One of the three newly identified genes, cdc37, has been located in the yeast linkage map on chromosome IV, two meiotic map units distal to hom2.—Each mutation produces stage-specific arrest of cell division at start, the same point where mating pheromone interrupts division. After synchronization at start by incubation at the restrictive temperature, the mutants retain the capacity to enlarge and to conjugate.

scholarly journals Causal relations among cell cycle processes in Tetrahymena pyriformis. An analysis employing temperature-sensitive mutants.

1976 ◽  
Vol 71 (1) ◽  
pp. 242-260 ◽  
Author(s):  
J Frankel ◽  
L M Jenkins ◽  
L E DeBault
Keyword(s):  
Cell Cycle ◽  
Cell Division ◽  
Dna Synthesis ◽  
Tetrahymena Pyriformis ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Developmental Pathway ◽  
Temperature Sensitive Mutants ◽  
Oral Development ◽  
Macronuclear Division

Utilization of temperature-sensitive mutants of Tetrahymena pyriformis affected in cell division or developmental pathway selection has permitted elucidation of causal dependencies interrelating micronuclear and macronuclear replication and division, oral development, and cytokinesis. In those mutants in which cell division is specifically blocked at restrictive temperatures, micronuclear division proceeds with somewhat accelerated periodicity but maintains normal coupling to predivision oral development. Macronuclear division is almost totally suppressed in an early acting mutant (mola) that prevents formation of the fission zone, and is variably affected in other mutants (such as mo3) that allow the fission zone to form but arrest constriction. However, macronuclear DNA synthesis can proceed for about four cycles in the nondividing mutant cells. A second class of mutants (psm) undergoes a switch of developmental pathway such that cells fail to enter division but instead repeatedly carry out an unusual type of oral replacement while growing in nutrient medium at the restrictive temperature. Under these circumstances no nuclei divide, yet macronuclear DNA accumulation continues. These results suggest that (a) macronuclear division is stringently affected by restriction of cell division, (b) micronuclear division and replication can continue in cells that are undergoing the type of oral development that is characteristic of division cycles, and (c) macronuclear DNA synthesis can continue in growing cells regardless of their developmental status. The observed relationships among events are consistent with the further suggestion that the cell cycle in this organism may consist of separate clusters of events. with a varying degree of coupling among clusters. A minimal model of the Tetrahymena cell cycle that takes these phenomena into account is suggested.

Test for temporal or spatial restrictions in gene product function during the cell division cycle

1983 ◽  
Vol 3 (7) ◽  
pp. 1255-1265
Author(s):  
S K Dutcher ◽  
L H Hartwell
Keyword(s):  
Cell Division ◽  
Gene Product ◽  
Gene Mutations ◽  
Cell Division Cycle ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Product Function ◽  
Complete Cell ◽  
Relationship Of ◽  
Gene Product Function

The ability of a functional gene to complement a nonfunctional gene may depend upon the intracellular relationship of the two genes. If so, the function of the gene product in question must be limited in time or in space. CDC (cell division cycle) gene products of Saccharomyces cerevisiae control discrete steps in cell division; therefore, they constitute reasonable candidates for genes that function with temporal or spatial restrictions. In an attempt to reveal such restrictions, we compared the ability of a CDC gene to complement a temperature-sensitive cdc gene in diploids where the genes are located within the same nucleus to complementation in heterokaryons where the genes are located in different nuclei. In CDC X cdc matings, complementation was monitored in rare heterokaryons by assaying the production of cdc haploid progeny (cytoductants) at the restrictive temperature. The production of cdc cytoductants indicates that the cdc nucleus was able to complete cell division at the restrictive temperature and implies that the CDC gene product was provided by the other nucleus or by cytoplasm in the heterokaryon. Cytoductants from cdc28 or cdc37 crosses were not efficiently produced, suggesting that these two genes are restricted spatially or temporally in their function. We found that of the cdc mutants tested 33 were complemented; cdc cytoductants were recovered at least as frequently as CDC cytoductants. A particularly interesting example was provided by the CDC4 gene. Mutations in CDC4 were found previously to produce a defect in both cell division and karyogamy. Surprisingly, the cell division defect of cdc4 nuclei is complemented by CDC4 nuclei in a heterokaryon, whereas the karyogamy defect is not.

scholarly journals Cell cycle arrest of cdc mutants and specificity of the RAD9 checkpoint.

Genetics ◽  
1993 ◽  
Vol 134 (1) ◽  
pp. 63-80 ◽  
Author(s):  
T A Weinert ◽  
L H Hartwell
Keyword(s):  
Cell Cycle ◽  
Cell Division ◽  
Dna Replication ◽  
Cell Cycle Control ◽  
Dna Lesions ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
A Cell ◽  
G2 Phase ◽  
Rad Mutants

Abstract In eucaryotes a cell cycle control called a checkpoint ensures that mitosis occurs only after chromosomes are completely replicated and any damage is repaired. The function of this checkpoint in budding yeast requires the RAD9 gene. Here we examine the role of the RAD9 gene in the arrest of the 12 cell division cycle (cdc) mutants, temperature-sensitive lethal mutants that arrest in specific phases of the cell cycle at a restrictive temperature. We found that in four cdc mutants the cdc rad9 cells failed to arrest after a shift to the restrictive temperature, rather they continued cell division and died rapidly, whereas the cdc RAD cells arrested and remained viable. The cell cycle and genetic phenotypes of the 12 cdc RAD mutants indicate the function of the RAD9 checkpoint is phase-specific and signal-specific. First, the four cdc RAD mutants that required RAD9 each arrested in the late S/G2 phase after a shift to the restrictive temperature when DNA replication was complete or nearly complete, and second, each leaves DNA lesions when the CDC gene product is limiting for cell division. Three of the four CDC genes are known to encode DNA replication enzymes. We found that the RAD17 gene is also essential for the function of the RAD9 checkpoint because it is required for phase-specific arrest of the same four cdc mutants. We also show that both X- or UV-irradiated cells require the RAD9 and RAD17 genes for delay in the G2 phase. Together, these results indicate that the RAD9 checkpoint is apparently activated only by DNA lesions and arrests cell division only in the late S/G2 phase.

scholarly journals RPC53 encodes a subunit of Saccharomyces cerevisiae RNA polymerase C (III) whose inactivation leads to a predominantly G1 arrest.

1992 ◽  
Vol 12 (10) ◽  
pp. 4314-4326 ◽  
Author(s):  
C Mann ◽  
J Y Micouin ◽  
N Chiannilkulchai ◽  
I Treich ◽  
J M Buhler ◽  
...  
Keyword(s):  
Cell Division ◽  
Amino Acid ◽  
Rna Polymerase ◽  
Essential Gene ◽  
Temperature Sensitive ◽  
G1 Arrest ◽  
Restrictive Temperature ◽  
Amino Acid Residues ◽  
Gene Encoding ◽  
Carboxy Terminal

RPC53 is shown to be an essential gene encoding the C53 subunit specifically associated with yeast RNA polymerase C (III). Temperature-sensitive rpc53 mutants were generated and showed a rapid inhibition of tRNA synthesis after transfer to the restrictive temperature. Unexpectedly, the rpc53 mutants preferentially arrested their cell division in the G1 phase as large, round, unbudded cells. The RPC53 DNA sequence is predicted to code for a hydrophilic M(r)-46,916 protein enriched in charged amino acid residues. The carboxy-terminal 136 amino acids of C53 are significantly similar (25% identical amino acid residues) to the same region of the human BN51 protein. The BN51 cDNA was originally isolated by its ability to complement a temperature-sensitive hamster cell mutant that undergoes a G1 cell division arrest, as is true for the rpc53 mutants.

DIPLOID SPORE FORMATION AND OTHER MEIOTIC EFFECTS OF TWO CELL-DIVISION-CYCLE MUTATIONS OF SACCHAROMYCES CEREVISIAE

Genetics ◽  
1980 ◽  
Vol 96 (4) ◽  
pp. 859-876 ◽  
Author(s):  
David Schild ◽  
Breck Byers
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Division ◽  
Spindle Pole Body ◽  
Spindle Pole ◽  
Cell Division Cycle ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Single Spore ◽  
Diploid Cells ◽  
Meiosis I

ABSTRACT The meiotic effects of two cell-division-cycle mutations of Saccharomyces cerevisiae (cdc5 and cdc14) have been examined. These mutations were isolated by L. H. Hartwell and his colleagues and characterized as defective in mitosis, causing a temperature-sensitive arrest in late nuclear division. When subjected to the restrictive temperature in meiosis, diploid cells homozygous for either of these mutations generally proceeded through premeiotic DNA synthesis and commitment to meiotic levels of recombination, but then arrested at a stage following spindle pole body (SPB) duplication and separation. The two SPBs lacked the interconnection by spindle microtubules typical of the complete meiosis I spindle. Challenge of these homozygotes by a semi-restrictive temperature often caused the production of asci containing two diploid spores. Genetic analysis of the viable pairs of spores revealed that each spore had become homozygous for centromere-linked markers significantly more frequently than for distal markers, indicating that the two spores each contained pairs of sister centromeres that had co-segregated in the reductional division of meiosis I. Ultrastructural analysis of the cdc5 homozygote demonstrated that these cells had completed meiosis I and formed two meiosis II spindles, but that the latter remained unusually short. This resulted in the encapsulation of both poles of each spindle within a single spore wall. These mutations therefore are defective in both meiotic divisions, as well as in the mitotic division described originally.

scholarly journals Interaction between the plasmid R6K andEscherichia coliwith defective DNA polymerase I

1978 ◽  
Vol 32 (1) ◽  
pp. 25-35 ◽  
Author(s):  
D. J. Tweats ◽  
J. T. Smith
Keyword(s):  
Cell Division ◽  
Dna Polymerase ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Dna Polymerase I ◽  
E Coli ◽  
R Factor ◽  
Polymerase I

SUMMARYInitial experiments demonstrated that the plasmid R6K cannot be transferred to or maintained readily in theE. coliDNA polymerase I deficient strain JG138polA1. Results withE. coliMM386polA12(R6K), which has a temperature sensitive polymerase I enzyme, showed cell division becomes abnormal when the polymerase I enzyme of the host bacteria is inactivated at the restrictive temperature. Under conditions of polymerase I deficiency, R6K replication, as measured by monitoring R-factor-mediated β-lactamase activity, also becomes abnormal with the loss of multiple R6K copies per cell and the apparent maintenance of a single R-factor copy per cell.

scholarly journals Genetic Analysis of Influenza Virus NS1 Gene: a Temperature-Sensitive Mutant Shows Defective Formation of Virus Particles

2005 ◽  
Vol 79 (24) ◽  
pp. 15246-15257 ◽  
Author(s):  
Urtzi Garaigorta ◽  
Ana M. Falcón ◽  
Juan Ortín
Keyword(s):  
Amino Acid ◽  
Genetic Analysis ◽  
Terminal Region ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Mutant Library ◽  
Ns1 Protein ◽  
Neuraminidase Treatment ◽  
Virus Particles ◽  
Ns1 Gene

ABSTRACT To perform a genetic analysis of the influenza A virus NS1 gene, a library of NS1 mutants was generated by PCR-mediated mutagenesis. A collection of mutant ribonucleic proteins containing the nonstructural genes was generated from the library that were rescued for an infectious virus mutant library by a novel RNP competition virus rescue procedure. Several temperature-sensitive (ts) mutant viruses were obtained by screening of the mutant library, and the sequences of their NS1 genes were determined. Most of the mutations identified led to amino acid exchanges and concentrated in the N-terminal region of the protein, but some of them occurred in the C-terminal region. Mutant 11C contained three mutations that led to amino acid exchanges, V18A, R44K, and S195P, all of which were required for the ts phenotype, and was characterized further. Several steps in the infection were slightly altered: (i) M1, M2, NS1, and neuraminidase (NA) accumulations were reduced and (ii) NS1 protein was retained in the nucleus in a temperature-independent manner, but these modifications could not justify the strong virus titer reduction at restrictive temperature. The most dramatic phenotype was the almost complete absence of virus particles in the culture medium, in spite of normal accumulation and nucleocytoplasmic export of virus RNPs. The function affected in the 11C mutant was required late in the infection, as documented by shift-up and shift-down experiments. The defect in virion production was not due to reduced NA expression, as virus yield could not be rescued by exogenous neuraminidase treatment. All together, the analysis of 11C mutant phenotype may indicate a role for NS1 protein in a late event in virus morphogenesis.

scholarly journals The RNA helicase Ddx52 functions as a growth switch in juvenile zebrafish

Development ◽  
2021 ◽  
Author(s):  
Tzu-Lun Tseng ◽  
Ying-Ting Wang ◽  
Chang-Yu Tsao ◽  
Yi-Teng Ke ◽  
Yi-Ching Lee ◽  
...  
Keyword(s):  
Positional Cloning ◽  
Single Gene ◽  
Rna Helicase ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Juvenile Growth ◽  
Peter Pan ◽  
Organism Growth ◽  
Juvenile Stages ◽  
Sensitive Allele

Vertebrate animals usually display robust growth trajectories during juvenile stages, and reversible suspension of this growth momentum by a single genetic determinant has not been reported. Here, we report a single genetic factor that is essential for juvenile growth in zebrafish. Using a forward genetic screen, we recovered a temperature-sensitive allele, pan (after Peter Pan), that suspends whole-organism growth at juvenile stages. Remarkably, even after growth is halted for a full 8-week period, pan mutants are able to resume a robust growth trajectory after release from the restrictive temperature, eventually growing into fertile adults without apparent adverse phenotypes. Positional cloning and complementation assays revealed that pan encodes a probable ATP-Dependent RNA Helicase (DEAD-Box Helicase 52; ddx52) that maintains the level of 47S precursor ribosomal RNA. Furthermore, genetic silencing of ddx52 and pharmacological inhibition of bulk RNA transcription similarly suspend the growth of flies, zebrafish and mice. Our findings reveal evidence that safe, reversible pauses of juvenile growth can be mediated by targeting the activity of a single gene, and that its pausing mechanism has high evolutionary conservation.

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.

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