scholarly journals Mitochondrial adenosine triphosphatase of the fission yeast Schizosaccharomyces pombe 972h-. Changes in inhibitor sensitivities during the cell cycle indicate similarities and differences in binding sites

1977 ◽  
Vol 162 (3) ◽  
pp. 581-590 ◽  
Author(s):  
D Lloyd ◽  
S W Edwards
Keyword(s):  
Cell Cycle ◽  
Schizosaccharomyces Pombe ◽  
Binding Sites ◽  
Adenosine Triphosphatase ◽  
Specific Activity ◽  
Inhibitor Binding ◽  
Titration Curves ◽  
Catalytic Sites ◽  
Cycle Dependent ◽  
Similar Site

1. We used 11 different inhibitors of energy conservation as inhibitors of ATPase (adenosine triphosphatase) in extracts of Schizosaccharomyces pombe obtained from cells at different stages of the cell cycle. 2. All the inhibitors showed cell-cycle-dependent variations in their I50 values (microng of inhibitor/mg of protein giving 50% inhibition of inhibitor-sensitive ATPase at pH 8.6). 3. From the sensitivity profiles through the cell cycle it was concluded that: (a) oligomycin, venturicidin, triethyltin sulphate and dibutylchloromethyltin chloride all act at closely associated site(s); (b) NN'-dicyclohexylcarbodi-imide and leucinostatin both act at a similar site, which is, however, distinct from that at which other inhibitors of the membrane factor (Fo) act. 4. The variations in I50 values for efrapeptin closely followed changes in specific activity of ATPase, as would be expected for an inhibitor acting at catalytic sites; these fluctuations were different from those for aurovertin, Dio-9, 4-chloro-7-nitrobenzofurazan, quercetin and spegazzinine, all of which show different sensitivity profiles from one another. 5. Anomalous stepwise inhibitor-titration curves were obtained for spegazzinine, NN'-dicyclohexylcarbodiimide, dibutylchloromethyltin chloride and leucinostatin. 6. Possible explanations are proposed for the discontinuous expression of inhibitor-binding sites during the cell cycle.

scholarly journals Time varying mobility and turnover of actomyosin ring components during cytokinesis in Schizosaccharomyces pombe

2020 ◽  
pp. mbc.E20-09-0588
Author(s):  
Anton Kamnev ◽  
Saravanan Palani ◽  
Paola Zambon ◽  
Tom Cheffings ◽  
Nigel Burroughs ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Schizosaccharomyces Pombe ◽  
Fluorescence Recovery After Photobleaching ◽  
Myosin Ii ◽  
Cellular Structures ◽  
Time Varying ◽  
Mobile Fraction ◽  
Novel Strategy ◽  
Cycle Dependent ◽  
Mobile Protein

Cytokinesis in many eukaryotes is dependent on a contractile actomyosin ring (AMR), composed of F-actin, myosin II, and other actin and myosin II regulators. Through fluorescence recovery after photobleaching experiments, many components of the AMR have been shown to be mobile and to undergo constant exchange with the cytosolic pools. However, how the mobility of its components changes at distinct stages of mitosis and cytokinesis has not been addressed. Here, we describe the mobility of eight Schizosaccharomyces pombe AMR proteins at different stages of mitosis and cytokinesis using an approach we have developed. We identified 3 classes of proteins, which showed 1) high (Ain1, Myo2, Myo51), 2) low (Rng2, Mid1, Myp2, Cdc12), and 3) cell cycle dependent (Cdc15) mobile fractions. We observed that the F-BAR protein Cdc15 undergoes a 20∼30% reduction in its mobile fraction after spindle breakdown and initiation of AMR contraction. Moreover, our data indicate that this change in Cdc15 mobility is dependent on the SIN-signalling pathway. Our work offers a novel strategy to estimate cell cycle-dependent mobile protein fractions in cellular structures and provides a valuable dataset, that is of interest to researchers working on cytokinesis.

scholarly journals Subunit composition determines E2F DNA-binding site specificity.

1997 ◽  
Vol 17 (12) ◽  
pp. 6994-7007 ◽  
Author(s):  
Y Tao ◽  
R F Kassatly ◽  
W D Cress ◽  
J M Horowitz
Keyword(s):  
Cell Cycle ◽  
Dna Binding ◽  
Binding Sites ◽  
Dna Bending ◽  
Susceptibility Gene ◽  
Specific Sequence ◽  
Cellular Genes ◽  
Cycle Dependent

The product of the retinoblastoma (Rb) susceptibility gene, Rb-1, regulates the activity of a wide variety of transcription factors, such as E2F, in a cell cycle-dependent fashion. E2F is a heterodimeric transcription factor composed of two subunits each encoded by one of two related gene families, denoted E2F and DP. Five E2F genes, E2F-1 through E2F-5, and two DP genes, DP-1 and DP-2, have been isolated from mammals, and heterodimeric complexes of these proteins are expressed in most, if not all, vertebrate cells. It is not yet clear whether E2F/DP complexes regulate overlapping and/or specific cellular genes. Moreover, little is known about whether Rb regulates all or a subset of E2F-dependent genes. Using recombinant E2F, DP, and Rb proteins prepared in baculovirus-infected cells and a repetitive immunoprecipitation-PCR procedure (CASTing), we have identified consensus DNA-binding sites for E2F-1/DP-1, E2F-1/DP-2, E2F-4/DP-1, and E2F-4/DP-2 complexes as well as an Rb/E2F-1/DP-1 trimeric complex. Our data indicate that (i) E2F, DP, and Rb proteins each influence the selection of E2F-binding sites; (ii) E2F sites differ with respect to their intrinsic DNA-bending properties; (iii) E2F/DP complexes induce distinct degrees of DNA bending; and (iv) complex-specific E2F sites selected in vitro function distinctly as regulators of cell cycle-dependent transcription in vivo. These data indicate that the specific sequence of an E2F site may determine its role in transcriptional regulation and suggest that Rb/E2F complexes may regulate subsets of E2F-dependent cellular genes.

scholarly journals Mitochondrial adenosine triphosphatase of the fission yeast, Schizosaccharomyces pombe 972h-. Changes in activity and inhibitor-sensitivity in response to catabolite repression

1976 ◽  
Vol 160 (2) ◽  
pp. 335-342 ◽  
Author(s):  
D Lloyd ◽  
S W Edwards
Keyword(s):  
Atpase Activity ◽  
Schizosaccharomyces Pombe ◽  
Adenosine Triphosphatase ◽  
Specific Activity ◽  
Early Stage ◽  
Gel Filtration ◽  
Growth Cycle ◽  
Mitochondrial Atpase ◽  
Submitochondrial Particles ◽  
Fourfold Increase

1. The specific activity of mitochondrial ATPase (adenosine triphosphatase) in extracts of Schizosaccharomyces pombe decreased 2.5-fold as the glucose concentration in the growth medium decreased from 50mM to 15mM. 2. During the late exponential phase of growth, ATPase activity doubled. 3. Sensitivity to oligomycin and Dio-9 as measured by values for I50(mug of inhibitor/mg of protein giving 50% inhibition) at pH 6.8 increased sixfold and ninefold respectively during the initial decrease in ATPase activity, and this degree of sensitivity was maintained for the remainder of the growth cycle. 4. Increased sensitivity to NN′-dicyclohexylcarbodi-imide, triethyltin and venturicidin was also observed during the early stage of glucose de-repression. 5. Smaller increases in sensitivity to efrapeptin, aurovertin, 7-chloro-4-nitrobenzo-2-oxa-1,3-diaz-le, quercetin and spegazzinine also occurred. 6. The ATPase of glycerol-grown cells was less sensitive to inhibitors than that of glucose-repressed cells; change in values for I50 were not so marked during the growth cycle of cells growing with glycerol. 7. When submitochondrial particles from glycerol-grown cells were tested by passage through Sephadex G-50, a fourfold increase in activity was accompanied by increased inhibitor resistance. 8. Gel filtration of submitochondrial particles from glucose-de-repressed cells gave similar results, whereas loss of ATPase occurred in submitochondrial particles from glucose-repressed cells. 9. It is proposed that alterations in sensitivity to inhibitors at different stages of glucose derepression may be partly controlled by a naturally occuring inhibitor of ATPase. 10. The inhibitors tested may be classififed into two groups on the basis of alterations of sensitivity of the ATPase during physiological modification: (a) oligomycin, Dio-9, NN′-dicyclohexylcarbodi-imide, venturicidin and triethyltin, and (b) efrapeptin, aurovertin, 7-chloro-4-nitrobenzo-2-oxa-1,3-diazole, quercetin and spegazzinine.

scholarly journals Cell cycle expression of two replicative DNA polymerases alpha and delta from Schizosaccharomyces pombe.

1993 ◽  
Vol 4 (2) ◽  
pp. 145-157 ◽  
Author(s):  
H Park ◽  
S Francesconi ◽  
T S Wang
Keyword(s):  
Cell Cycle ◽  
Schizosaccharomyces Pombe ◽  
Dna Polymerases ◽  
Single Copy ◽  
Upstream Region ◽  
Vitro Assay ◽  
Expression Of Genes ◽  
Genes Encoding ◽  
Cycle Dependent

We have investigated the expression of two Schizosaccharomyces pombe replicative DNA polymerases alpha and delta during the cell cycle. The pol alpha+ and pol delta+ genes encoding DNA polymerases alpha and delta were isolated from S. pombe. Both pol alpha+ and pol delta+ genes are single copy genes in haploid cells and are essential for cell viability. In contrast to Saccharomyces cerevisiae homologs, the steady-state transcripts of both S. pombe pol alpha+ and pol delta+ genes were present throughout the cell cycle. Sequence analysis of the pol alpha+ and pol delta+ genes did not reveal the Mlu I motifs in their upstream sequences that are involved in cell cycle-dependent transcription of S. cerevisiae DNA synthesis genes as well as the S. pombe cdc22+ gene at the G1/S boundary. However, five near-match Mlu I motifs were found in the upstream region of the pol alpha+ gene. S. pombe DNA polymerases alpha and delta proteins were also expressed constantly throughout the cell cycle. In addition, the enzymatic activity of the S. pombe DNA polymerase alpha measured by in vitro assay was detected at all stages of the cell cycle. Thus, these S. pombe replicative DNA polymerases, like that of S. pombe cdc17+ gene, are expressed throughout the cell cycle at the transcriptional and protein level. These results indicate that S. pombe has at least two regulatory modes for the expression of genes involved in DNA replication and DNA precursor synthesis.

Transcription of the E2F-1 gene is rendered cell cycle dependent by E2F DNA-binding sites within its promoter

1994 ◽  
Vol 14 (10) ◽  
pp. 6607-6615
Author(s):  
E Neuman ◽  
E K Flemington ◽  
W R Sellers ◽  
W G Kaelin
Keyword(s):  
Cell Cycle ◽  
Binding Sites ◽  
Mammalian Cells ◽  
Mrna Abundance ◽  
Dna Binding Sites ◽  
Cell Cycle Dependence ◽  
Transcription Factor E2f ◽  
Cycle Dependent

The cell cycle-regulatory transcription factor E2F-1 is regulated by interactions with proteins such as the retinoblastoma gene product and by cell cycle-dependent alterations in E2F-1 mRNA abundance. To better understand this latter phenomenon, we have isolated the human E2F-1 promoter. The human E2F-1 promoter, fused to a luciferase cDNA, gave rise to cell cycle-dependent luciferase activity upon transfection into mammalian cells in a manner which paralleled previously reported changes in E2F-1 mRNA abundance. The E2F-1 promoter contains four potential E2F-binding sites organized as two imperfect palindromes. Gel shift and transactivation studies suggested that these sites can bind to E2F in vitro and in vivo. Mutation of the two E2F palindromes abolished the cell cycle dependence of the E2F-1 promoter. Thus, E2F-1 appears to be regulated at the level of transcription, and this regulation is due, at least in part, to binding of one or more E2F family members to the E2F-1 promoter.

scholarly journals Transcription of the E2F-1 gene is rendered cell cycle dependent by E2F DNA-binding sites within its promoter.

1994 ◽  
Vol 14 (10) ◽  
pp. 6607-6615 ◽  
Author(s):  
E Neuman ◽  
E K Flemington ◽  
W R Sellers ◽  
W G Kaelin
Keyword(s):  
Cell Cycle ◽  
Binding Sites ◽  
Mammalian Cells ◽  
Mrna Abundance ◽  
Dna Binding Sites ◽  
Cell Cycle Dependence ◽  
Transcription Factor E2f ◽  
Cycle Dependent

The cell cycle-regulatory transcription factor E2F-1 is regulated by interactions with proteins such as the retinoblastoma gene product and by cell cycle-dependent alterations in E2F-1 mRNA abundance. To better understand this latter phenomenon, we have isolated the human E2F-1 promoter. The human E2F-1 promoter, fused to a luciferase cDNA, gave rise to cell cycle-dependent luciferase activity upon transfection into mammalian cells in a manner which paralleled previously reported changes in E2F-1 mRNA abundance. The E2F-1 promoter contains four potential E2F-binding sites organized as two imperfect palindromes. Gel shift and transactivation studies suggested that these sites can bind to E2F in vitro and in vivo. Mutation of the two E2F palindromes abolished the cell cycle dependence of the E2F-1 promoter. Thus, E2F-1 appears to be regulated at the level of transcription, and this regulation is due, at least in part, to binding of one or more E2F family members to the E2F-1 promoter.

Cell-cycle–dependent oscillation of GATA2 expression in hematopoietic cells

Blood ◽  
2007 ◽  
Vol 109 (10) ◽  
pp. 4200-4208 ◽  
Author(s):  
Shinichiro Koga ◽  
Nobuhiro Yamaguchi ◽  
Tomoko Abe ◽  
Masayoshi Minegishi ◽  
Shigeru Tsuchiya ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Fluorescent Protein ◽  
Specific Activity ◽  
Expression Profiles ◽  
Hematopoietic Cells ◽  
Hematopoietic Stem ◽  
Altered Expression ◽  
Cyclin A2 ◽  
Cycle Dependent

Abstract In vitro manipulation of hematopoietic stem cells (HSCs) is a key issue in both transplantation therapy and regenerative medicine, and thus new methods are required to achieve HSC expansion with self-renewal. GATA2 is a transcription factor controlling pool size of HSCs. Of interest, continuous overexpression of GATA2 does not induce HSC proliferation. In this report, we demonstrate that GATA2 expression, in leukemic and normal hematopoietic cells, oscillates during the cell cycle, such that expression is high in S phase but low in G1/S and M phase. GATA2 binding to target Bcl-X gene also oscillates in accordance with GATA2 expression. Using a green fluorescent protein (GFP)–GATA2 fusion protein, we demonstrate cell-cycle–specific activity of proteasome-dependent degradation of GATA2. Immunoprecipitation/immunoblotting analysis demonstrated phosphorylation of GATA2 at cyclin-dependent kinase (Cdk)–consensus motifs, S/T0P+1, and interaction of GATA2 with Cdk2/cyclin A2–, Cdk2/cyclin A2–, and Cdk4/cyclin D1–phosphorylated GATA2 in vitro. Mutants in phosphorylation motifs exhibited altered expression profiles of GFP-GATA2 domain fusion proteins. These results indicate that GATA2 phosphorylation by Cdk/cyclin systems is responsible for the cell-cycle–dependent regulation of GATA2 expression, and suggest the possibility that a cell-cycle–specific “on-off” response of GATA2 expression may control hematopoietic-cell proliferation and survival.

scholarly journals Cell cycle-dependent nucleosome occupancy at cohesin binding sites in yeast chromosomes

Genomics ◽  
2008 ◽  
Vol 91 (3) ◽  
pp. 274-280 ◽  
Author(s):  
Jie Liu ◽  
Daniel M. Czajkowsky ◽  
Shoudan Liang ◽  
Zhifeng Shao
Keyword(s):  
Cell Cycle ◽  
Binding Sites ◽  
Nucleosome Occupancy ◽  
Cycle Dependent
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