scholarly journals Temperature-sensitive mutations that cause stage-specific defects in Zebrafish fin regeneration.

Genetics ◽  
1995 ◽  
Vol 141 (4) ◽  
pp. 1583-1595 ◽  
Author(s):  
S L Johnson ◽  
J A Weston
Keyword(s):  
Critical Period ◽  
Slow Growth ◽  
Temperature Shift ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Subsequent Growth ◽  
Fin Regeneration ◽  
Bone Differentiation ◽  
Early Stages ◽  
Wound Epithelium

Abstract When amputated, the fins of adult zebrafish rapidly regenerate the missing tissue. Fin regeneration proceeds through several stages, including wound healing, establishment of the wound epithelium, recruitment of the blastema from mesenchymal cells underlying the wound epithelium, and differentiation and outgrowth of the regenerate. We screened for temperature-sensitive mutations that affect the regeneration of the fin. Seven mutations were identified, including five that fail to regenerate their fins, one that causes slow growth during regeneration, and one that causes dysmorphic bumps or tumors to develop in the regenerating fin. reg5 mutants fail to regenerate their caudal fins, whereas reg6 mutants develop dysmorphic bumps in their regenerates at the restrictive temperature. Temperature-shift experiments indicate that reg5 and reg6 affect different stages of regeneration. The critical period for reg5 occurs during the early stages of regeneration before or during establishment of the blastema, resulting in defects in subsequent growth of the blastema and failure to differentiate bone-forming cells. The critical period for reg6 occurs after the onset of bone differentiation and during early stages of regenerative outgrowth. Both reg5 and reg6 also show temperature-sensitive defects in embryonic development or in ontogenetic outgrowth of the juvenile fin.

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 MEIOTIC RECOMBINATION AND DNA SYNTHESIS IN A NEW CELL CYCLE MUTANT OF SACCHAROMYCES CEREVISIAE

Genetics ◽  
1978 ◽  
Vol 90 (1) ◽  
pp. 49-68
Author(s):  
Yona Kassir ◽  
Giora Simchen
Keyword(s):  
Cell Cycle ◽  
Dna Replication ◽  
Dna Synthesis ◽  
Nuclear Division ◽  
Temperature Shift ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Permissive Temperature ◽  
Essential Function ◽  
Normal Meiosis

ABSTRACT Vegetative cells carrying the new temperature-sensitive mutation cdc40 arrest at the restrictive temperature with a medial nuclear division phenotype. DNA replication is observed under these conditions, but most cells remain sensitive to hydroxyurea and do not complete the ongoing cell cycle if the drug is present during release from the temperature block. It is suggested that the cdc40 lesion affects an essential function in DNA synthesis. Normal meiosis is observed at the permissive temperature in cdc40 homozygotes. At the restrictive temperature, a full round of premeiotic DNA replication is observed, but neither commitment to recombination nor later meiotic events occur. Meiotic cells that are already committed to the recombination process at the permissive temperature do not complete it if transferred to the restrictive temperature before recombination is realized. These temperature shift-up experiments demonstrate that the CDC40 function is required for the completion of recombination events, as well as for the earlier stage of recombination commitment. Temperature shift-down experiments with cdc40 homozygotes suggest that meiotic segregation depends on the final events of recombination rather than on commitment to recombination.

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.

Conditional absence of mitosis-specific antigens in a temperature-sensitive embryonic-arrest mutant of Caenorhabditis elegans

1987 ◽  
Vol 87 (2) ◽  
pp. 305-314 ◽  
Author(s):  
R.M. Hecht ◽  
M. Berg-Zabelshansky ◽  
P.N. Rao ◽  
F.M. Davis
Keyword(s):  
Caenorhabditis Elegans ◽  
Mammalian Cells ◽  
Temperature Shift ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
C Elegans ◽  
Phosphatase Treatment ◽  
M Phase ◽  
A Cell ◽  
Embryonic Arrest

A monoclonal antibody, specific to phosphoproteins in mitotic HeLa cells was found to crossreact with a similar set of proteins in embryos of the nematode, Caenorhabditis elegans. In C. elegans, as in mammalian cells, the highly conserved antigenic epitope is associated with a family of high molecular weight polypeptides. The antigenic reactivity of these multiple proteins also depends on their phosphorylation, since antibody binding is reduced after alkaline phosphatase treatment. The antigens are detected at the centrosomes, and in the nuclear region and surrounding cytoplasm of mitotic cells. The significance of these antigens is emphasized by their absence at restrictive temperature in embryos of the temperature-sensitive embryonic-arrest mutant, emb-29V. Furthermore, temperature shift-down experiments suggest that the emb-29 mutation defines a cell division cycle function that affects an essential activity required for progression into M phase.

scholarly journals Residual cell division measurements are unreliable as indicators of the timing of events in the Saccharomyces cerevisiae cell cycle

1983 ◽  
Vol 64 (1) ◽  
pp. 307-322
Author(s):  
K.M. Richmond ◽  
D.H. Williamson
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
Cell Division ◽  
Temperature Shift ◽  
Growth Conditions ◽  
Population Doubling ◽  
Population Doubling Time ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Point Data

We report here an analysis of the execution point of the temperature-sensitive Saccharomyces cerevisiae cell cycle mutant, cdc27-47. When a logarithmically growing culture was shifted from standard growth conditions (strain 27.8B growing in YEPD at 25 degrees C) to the restrictive temperature cell division ceased abruptly and reproducibly within one population doubling time, the extent of cell division indicating an execution point early in the cell cycle. Approximately 50% of stationary-phase cells were able to divide when refed with fresh medium at 37 degrees C, showing that the execution point could be passed before ‘start’. This makes the sharp cut-off in cell division difficult to explain. This difficulty was compounded by observations of the cell cycle stage at which individual cells acquired the capacity to divide at 37 degrees C. Half the cells that were budded at the time of a temperature shift-up formed three division-blocked cells, and in 11 of these 13 cases, two were descended from the original mother cell and one from the original bud. Thus, mother and daughter cells pass the execution point independently; daughters usually during G1, and mothers usually in the budded phase of the previous cycle. The sharp cut-off in cell division is therefore spurious, and a mechanism is proposed to account for it, which has implications for the interpretation of the execution points of other cdc mutants. In addition, the expression of the cdc27-47 execution point was modified by both genetic and environmental factors, being affected by carbon source, by the petite condition, and by genetic background. This illustrates the difficulties of interpreting execution point data and the dangers of extrapolation of cell cycle parameters between strains and growth conditions.

scholarly journals Sld2, Which Interacts with Dpb11 inSaccharomyces cerevisiae, Is Required for Chromosomal DNA Replication

1998 ◽  
Vol 18 (10) ◽  
pp. 6102-6109 ◽  
Author(s):  
Yoichiro Kamimura ◽  
Hiroshi Masumoto ◽  
Akio Sugino ◽  
Hiroyuki Araki
Keyword(s):  
Dna Replication ◽  
Temperature Shift ◽  
Growth Defect ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Chromosomal Dna ◽  
Autonomously Replicating Sequence ◽  
Complementation Groups ◽  
Synthetically Lethal ◽  
Dna Polymerase Ii

ABSTRACT The DPB11 gene, which genetically interacts with DNA polymerase II (ɛ), one of three replicative DNA polymerases, is required for DNA replication and the S phase checkpoint inSaccharomyces cerevisiae. To identify factors interacting with Dbp11, we have isolated sld (synthetically lethal withdpb11-1) mutations which fall into six complementation groups (sld1 to -6). In this study, we characterized SLD2, encoding an essential 52-kDa protein. High-copy SLD2 suppressed the thermosensitive growth defect caused by dpb11-1. Conversely, high-copy DPB11suppressed the temperature-sensitive growth defect caused bysld2-6. The interaction between Sld2 and Dpb11 was detected in a two-hybrid assay. This interaction was evident at 25°C but not at 34°C when Sld2-6 or Dpb11-1 replaced its wild-type protein. No interaction between Sld2-6 and Dpb11-1 could be detected even at 25°C. Immunoprecipitation experiments confirmed that Dpb11 physically interacts with Sld2. sld2-6 cells were defective in DNA replication at the restrictive temperature, as were dpb11-1cells. Further, in dpb11-1 and sld2-6 cells, the bubble-shaped replication intermediates formed in the region of the autonomously replicating sequence reduced quickly after a temperature shift. These results strongly suggest the involvement of the Dpb11-Sld2 complex in a step close to the initiation of DNA replication.

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.

scholarly journals The Putative Nucleic Acid Helicase Sen1p Is Required for Formation and Stability of Termini and for Maximal Rates of Synthesis and Levels of Accumulation of Small Nucleolar RNAs inSaccharomyces cerevisiae

1998 ◽  
Vol 18 (12) ◽  
pp. 6885-6896 ◽  
Author(s):  
Theodore P. Rasmussen ◽  
Michael R. Culbertson
Keyword(s):  
Nucleic Acid ◽  
Temperature Shift ◽  
Northern Blotting ◽  
Rnase H ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Type I ◽  
Small Nucleolar Rnas ◽  
Rna Helicases ◽  
Nucleolar Rnas

ABSTRACT Sen1p from Saccharomyces cerevisiae is a nucleic acid helicase related to DEAD box RNA helicases and type I DNA helicases. The temperature-sensitive sen1-1 mutation located in the helicase motif alters the accumulation of pre-tRNAs, pre-rRNAs, and some small nuclear RNAs. In this report, we show that cells carryingsen1-1 exhibit altered accumulation of several small nucleolar RNAs (snoRNAs) immediately upon temperature shift. Using Northern blotting, RNase H cleavage, primer extension, and base compositional analysis, we detected three forms of the snoRNA snR13 in wild-type cells: an abundant TMG-capped 124-nucleotide (nt) mature form (snR13F) and two less abundant RNAs, including a heterogeneous population of ∼1,400-nt 3′-extended forms (snR13R) and a 108-nt 5′-truncated form (snR13T) that is missing 16 nt at the 5′ end. A subpopulation of snR13R contains the same 5′ truncation. Newly synthesized snR13R RNA accumulates with time at the expense of snR13F following temperature shift of sen1-1 cells, suggesting a possible precursor-product relationship. snR13R and snR13T both increase in abundance at the restrictive temperature, indicating that Sen1p stabilizes the 5′ end and promotes maturation of the 3′ end. snR13F contains canonical C and D boxes common to many snoRNAs. The 5′ end of snR13T and the 3′ end of snR13F reside within C2U4 sequences that immediately flank the C and D boxes. A mutation in the 5′ C2U4 repeat causes underaccumulation of snR13F, whereas mutations in the 3′ C2U4 repeat cause the accumulation of two novel RNAs that migrate in the 500-nt range. At the restrictive temperature, double mutants carrying sen1-1 and mutations in the 3′ C2U4 repeat show reduced accumulation of the novel RNAs and increased accumulation of snR13R RNA, indicating that Sen1p and the 3′ C2U4 sequence act in a common pathway to facilitate 3′ end formation. Based on these findings, we propose that Sen1p and the C2U4 repeats that flank the C and D boxes promote maturation of the 3′ terminus and stability of the 5′ terminus and are required for maximal rates of synthesis and levels of accumulation of mature snR13F.

scholarly journals hyp Loci Control Cell Pattern Formation in the Vegetative Mycelium of Aspergillus nidulans

Genetics ◽  
1998 ◽  
Vol 148 (2) ◽  
pp. 669-680
Author(s):  
Susan G W Kaminskyj ◽  
John E Hamer
Keyword(s):  
Aspergillus Nidulans ◽  
Tip Growth ◽  
Nuclear Division ◽  
Apical Cell ◽  
Control Cell ◽  
Temperature Shift ◽  
Growth Cycle ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Apical Cells

Abstract Aspergillus nidulans grows by apical extension of multinucleate cells called hyphae that are subdivided by the insertion of crosswalls called septa. Apical cells vary in length and number of nuclei, whereas subapical cells are typically 40 μm long with three to four nuclei. Apical cells have active mitotic cycles, whereas subapical cells are arrested for growth and mitosis until branch formation reinitiates tip growth and nuclear divisions. This multicellular growth pattern requires coordination between localized growth, nuclear division, and septation. We searched a temperature-sensitive mutant collection for strains with conditional defects in growth patterning and identified six mutants (designated hyp for hypercellular). The identified hyp mutations are nonlethal, recessive defects in five unlinked genes (hypA-hypE). Phenotypic analyses showed that these hyp mutants have aberrant patterns of septation and show defects in polarity establishment and tip growth, but they have normal nuclear division cycles and can complete the asexual growth cycle at restrictive temperature. Temperature shift analysis revealed that hypD and hypE play general roles in hyphal morphogenesis, since inactivation of these genes resulted in a general widening of apical and subapical cells. Interestingly, loss of hypA or hypB function lead to a cessation of apical cell growth but activated isotropic growth and mitosis in subapical cells. The inferred functions of hypA and hypB suggest a mechanism for coordinating apical growth, subapical cell arrest, and mitosis in A. nidulans.

STUDIES ON THE DEVELOPMENTAL CHARACTERISTICS OF FUSED MUTANTS OF DROSOPHILA MELANOGASTER

1979 ◽  
Vol 21 (3) ◽  
pp. 335-346 ◽  
Author(s):  
Glen G. Wurst ◽  
William P. Hanratty
Keyword(s):  
Posterior Part ◽  
Temperature Shift ◽  
Developmental Time ◽  
Wing Disc ◽  
Sensitive Period ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Affected Tissue ◽  
Developmental Characteristics ◽  
Fused Gene

The development of the lethal and visible effects of 16 independently arising fused mutations was studied. No complementation was observed in any of the 120 heteroallelic combinations of fused mutations. Temperature shift experiments indicated that there is coordinance of expression of all the pleiotropic fused effects, with regard to both temperature sensitive period and degree of expressivity. These studies showed that the expressivity of both the lethal and visible fused effects are directly related to pupal developmental time spent at the restrictive temperature. Gynandromorph studies indicated that all visible effects of the fused mutation develop autonomously and that the thoracic effects are localized in the posterior part of the anterior developmental compartment of the wing disc. These data support the conclusion that the fused locus contains a single functional unit. The data also suggest that the fused gene is expressed simultaneously in each affected tissue of the fly and that the fused product may perform a similar function in each affected part of the fly.

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