scholarly journals Anoxic Fibroblasts Activate a Replication Checkpoint That Is Bypassed By E1a

2003 ◽  
Vol 23 (24) ◽  
pp. 9032-9045 ◽  
Author(s):  
Lawrence B. Gardner ◽  
Feng Li ◽  
Xuejie Yang ◽  
Chi V. Dang
Keyword(s):  
Cell Cycle ◽  
Cell Cycle Regulation ◽  
S Phase ◽  
Low Oxygen ◽  
Replication Checkpoint ◽  
Phase Arrest ◽  
Distinct Cell ◽  
Rat Fibroblasts ◽  
Transcription Factor E2f ◽  
Cycle Regulation

ABSTRACT Little is known about cell cycle regulation in hypoxic cells, despite its significance. We utilized an experimentally tractable model to study the proliferative responses of rat fibroblasts when rendered hypoxic (0.5% oxygen) or anoxic (<0.01% oxygen). Hypoxic cells underwent G1 arrest, whereas anoxic cells also demonstrated S-phase arrest due to suppression of DNA initiation. Upon reoxygenation, only those cells arrested in G1 were able to resume proliferation. The oncoprotein E1a induced p53-independent apoptosis in anoxic cells, which when suppressed by Bcl-2 permitted proliferation despite anoxia. E1a expression led to marked increases in the transcription factor E2F, and overexpression of E2F-1 allowed proliferation in hypoxic cells, although it had minimal effect on the anoxic suppression of DNA initiation. We thus demonstrate two distinct cell cycle responses to low oxygen and suggest that alterations that lead to increased E2F can overcome hypoxic G1 arrest but that additional alterations, promoted by E1a expression, are necessary for neoplastic cells to proliferate despite anoxia.

scholarly journals Posttranslational Phosphorylation and Ubiquitination of the Saccharomyces cerevisiae Poly(A) Polymerase at the S/G2 Stage of the Cell Cycle

2000 ◽  
Vol 20 (8) ◽  
pp. 2794-2802 ◽  
Author(s):  
Neptune Mizrahi ◽  
Claire Moore
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
Cell Sorting ◽  
S Phase ◽  
Phase Arrest ◽  
Phosphatase Treatment ◽  
M Phase ◽  
64 Kda Protein ◽  
Mitotic Phosphorylation ◽  
Cycle Regulation

ABSTRACT The poly(A) polymerase of the budding yeast Saccharomyces cerevisiae (Pap1) is a 64-kDa protein essential for the maturation of mRNA. We have found that a modified Pap1 of 90 kDa transiently appears in cells after release from α-factor-induced G1 arrest or from a hydroxyurea-induced S-phase arrest. While a small amount of modification occurs in hydroxyurea-arrested cells, fluorescence-activated cell sorting analysis and microscopic examination of bud formation indicate that the majority of modified enzyme is found at late S/G2 and disappears by the time cells have reached M phase. The reduction of the 90-kDa product upon phosphatase treatment indicates that the altered mobility is due to phosphorylation. A preparation containing primarily the phosphorylated Pap1 has no poly(A) addition activity, but this activity is restored by phosphatase treatment. A portion of Pap1 is also polyubiquitinated concurrent with phosphorylation. However, the bulk of the 64-kDa Pap1 is a stable protein with a half-life of 14 h. The timing, nature, and extent of Pap1 modification in comparison to the mitotic phosphorylation of mammalian poly(A) polymerase suggest an intriguing difference in the cell cycle regulation of this enzyme in yeast and mammalian systems.

scholarly journals WEE1 kinase inhibitor AZD1775 induces CDK1 kinase-dependent origin firing in unperturbed G1- and S-phase cells

2019 ◽  
Vol 116 (48) ◽  
pp. 23891-23893 ◽  
Author(s):  
Tatiana N. Moiseeva ◽  
Chenao Qian ◽  
Norie Sugitani ◽  
Hatice U. Osmanbeyoglu ◽  
Christopher J. Bakkenist
Keyword(s):  
Cell Cycle ◽  
Clinical Trials ◽  
Cell Cycle Regulation ◽  
Kinase Inhibitor ◽  
S Phase ◽  
Origin Firing ◽  
Replicative Helicase ◽  
Origin Licensing ◽  
Wee1 Kinase ◽  
Cycle Regulation

WEE1 kinase is a key regulator of the G2/M transition. The WEE1 kinase inhibitor AZD1775 (WEE1i) induces origin firing in replicating cells. We show that WEE1i induces CDK1-dependent RIF1 phosphorylation and CDK2- and CDC7-dependent activation of the replicative helicase. WEE1 suppresses CDK1 and CDK2 kinase activities to regulate the G1/S transition after the origin licensing is complete. We identify a role for WEE1 in cell cycle regulation and important effects of AZD1775, which is in clinical trials.

scholarly journals p21Cip1 and p27Kip1Regulate Cell Cycle Reentry after Hypoxic Stress but Are Not Necessary for Hypoxia-Induced Arrest

2001 ◽  
Vol 21 (4) ◽  
pp. 1196-1206 ◽  
Author(s):  
Susannah L. Green ◽  
Rachel A. Freiberg ◽  
Amato J. Giaccia
Keyword(s):  
Cell Cycle ◽  
Dna Synthesis ◽  
Kinase Inhibitors ◽  
S Phase ◽  
Hypoxic Stress ◽  
Cyclin Dependent Kinase ◽  
Low Oxygen ◽  
Double Knockout ◽  
Phase Arrest ◽  
S Phase Arrest

ABSTRACT We investigated the role of the cyclin-dependent kinase inhibitors p21Cip1 and p27Kip1 in cell cycle regulation during hypoxia and reoxygenation. While moderate hypoxia (1 or 0.1% oxygen) does not significantly impair bromodeoxyuridine incorporation, at very low oxygen tensions (0.01% oxygen) DNA replication is rapidly shut down in immortalized mouse embryo fibroblasts. This S-phase arrest is intact in fibroblasts lacking the cyclin kinase inhibitors p21Cip1 and p27Kip1, indicating that these molecules are not essential elements of the arrest pathway. Hypoxia-induced arrest is accompanied by dephosphorylation of pRb and inhibition of cyclin-dependent kinase 2, which results in part from inhibitory phosphorylation. Interestingly, cells lacking the retinoblastoma tumor suppressor protein also display arrest under hypoxia, suggesting that pRb is not an essential mediator of this response. Upon reoxygenation, DNA synthesis resumes by 3.5 h and reaches aerobic levels by 6 h. Cells lacking p21, however, resume DNA synthesis more rapidly upon reoxygenation than wild-type cells, suggesting that this inhibitor may play a role in preventing premature reentry into the cell cycle upon cessation of the hypoxic stress. While p27 null cells did not exhibit rapid reentry into the cell cycle, cells lacking both p21 and p27 entered S phase even more aggressively than those lacking p21 alone, revealing a possible secondary role for p27 in this response. Cdk2 activity is also restored more rapidly in the double-knockout cells when returned to normoxia. These studies reveal that restoration of DNA synthesis after hypoxic stress, but not the S phase arrest itself, is regulated by p21 and p27.

scholarly journals APC/C and retinoblastoma interaction: cross-talk of retinoblastoma protein with the ubiquitin proteasome pathway

2016 ◽  
Vol 36 (5) ◽  
Author(s):  
Ajeena Ramanujan ◽  
Swati Tiwari
Keyword(s):  
Cell Cycle ◽  
Cell Cycle Regulation ◽  
Alternative Pathway ◽  
Retinoblastoma Protein ◽  
S Phase ◽  
Anaphase Promoting Complex ◽  
Anti Cancer ◽  
Ubiquitin Proteasome ◽  
Proteasome Pathway ◽  
Cycle Regulation

The ubiquitin (Ub) ligase anaphase promoting complex/cyclosome (APC/C) and the tumour suppressor retinoblastoma protein (pRB) play key roles in cell cycle regulation. APC/C is a critical regulator of mitosis and G1-phase of the cell cycle whereas pRB keeps a check on proliferation by inhibiting transition to the S-phase. APC/C and pRB interact with each other via the co-activator of APC/C, FZR1, providing an alternative pathway of regulation of G1 to S transition by pRB using a post-translational mechanism. Both pRB and FZR1 have complex roles and are implicated not only in regulation of cell proliferation but also in differentiation, quiescence, apoptosis, maintenance of chromosomal integrity and metabolism. Both are also targeted by transforming viruses. We discuss recent advances in our understanding of the involvement of APC/C and pRB in cell cycle based decisions and how these insights will be useful for development of anti-cancer and anti-viral drugs.

Meiosis-specific cell cycle regulation in maturing Xenopus oocytes

1994 ◽  
Vol 107 (11) ◽  
pp. 3005-3013 ◽  
Author(s):  
K. Ohsumi ◽  
W. Sawada ◽  
T. Kishimoto
Keyword(s):  
Cell Cycle ◽  
Meiotic Division ◽  
Cell Cycle Regulation ◽  
Xenopus Oocytes ◽  
Kinase Activity ◽  
S Phase ◽  
Cell Cycles ◽  
Minute Interval ◽  
Cycle Regulation ◽  
Meiosis I

Meiotic cell cycles differ from mitotic cell cycles in that the former lack S-phase in the interphase between meiosis I and meiosis II. To obtain clues for mechanisms involved in the cell cycle regulation unique to meiosis, we have examined changes in chromosomal morphology and H1 kinase activity during a meiotic period from metaphase I (MI) to metaphase II (MII) in Xenopus oocytes. Using populations of oocytes that underwent germinal vesicle breakdown (GVBD) within a 10 minute interval, we found that the kinase activity declined gradually during the 60 minute period after GVBD and then increased steadily during the following 80 minute interval, showing remarkable differences from the rapid drop and biphasic increase of the kinase activity in intermitotic periods (Solomon et al. (1990) Cell 63, 1013–1024; Dasso and Newport (1990) Cell 61, 811–823). We also found that the exit from MI lagged, by more than 30 minutes, behind the time of lowest H1 kinase activity, whereas the two events took place concomitantly at the end of meiosis II and mitosis. Consequently, the H1 kinase activity was already increasing during the first meiotic division. When H1 kinase activation at MII was delayed by a transient inhibition of protein synthesis after GVBD, oocytes were able to support formation of interphase nuclei and DNA replication between the first meiotic division and the MII arrest, indicating that the cell cycle entered S-phase between meiosis I and meiosis II. These results strongly suggest that the machinery required for entering S-phase has been established in maturing oocytes by the end of meiosis I.(ABSTRACT TRUNCATED AT 250 WORDS)

The Cyclin Interactor p26INCA1 Regulates the Hematopoietic Stem Cell Pool Via CDK Inhibition.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 637-637
Author(s):  
Nicole Baeumer ◽  
Sven Diederichs ◽  
Steffen Koschmieder ◽  
Boris V. Skryabin ◽  
Feng Zhang ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Bone Marrow ◽  
Stem Cell ◽  
Cell Cycle Regulation ◽  
Kinase Activity ◽  
Bone Marrow Cells ◽  
S Phase ◽  
Hematopoietic Stem ◽  
Cdk2 Activity ◽  
Cycle Regulation

Abstract Cell cycle progression is driven by the kinase activity of cyclin/CDK complexes. Dysregulation of the cell cycle leads to altered cell growth and contributes to tumorigenesis. Recently, we identified p26INCA1 as novel interaction partner of Cyclin A1/CDK2. Here, we characterize the phenotype of Inca1-null mice to uncover the cellular and molecular function of Inca1. Inca1-knockout mice were viable and fertile. FACS analyses revealed that aging mutant animals harbored an increased hematopoietic stem cell (HSC) pool. Bone marrow cells of young mice exhibited enhanced clonogenic replating efficiency in colony formation assays as compared to wildtype mice. Weekly administration of the myeloablative agent 5-fluorouracil (5-FU) led to a significantly shorter life span of Inca1−/ − mice compared to wildtype littermates. The increased 5-FU toxicity might thus be related to a higher number of cycling HSC in Inca1−/ − bone marrow. Analysis of the impact of Inca1 on cell cycle regulation demonstrated that the fraction of Inca1−/ − embryonic fibroblasts (MEFs) in S phase was significantly increased. Ectopic INCA1 expression reduced proliferation and colony formation of proliferating cells such as primary bone marrow cells, HeLa, HuTu80 and 32D cell lines. Serum starvation rapidly induced and mitogenic signals inhibited Inca1 expression providing a further link to cell cycle regulation. To identify the molecular mechanism of cell cycle regulation by Inca1, we investigated the influence of Inca1 on the direct inhibition of CDK2. In spleen lysates from Inca1-deficient mice, cellular CDK2 kinase activity towards Histone H1 was significantly induced compared to lysates of wildtype littermates. In in vitro kinase assays, recombinant INCA1 strongly inhibited CDK2 activity. In addition, we hypothesized that other cyclin kinase inhibitors (CKI) could partially compensate in vivo for the loss of Inca1 function. p21cip1/waf1 mRNA and protein expression were induced in Inca1−/ − MEFs compared to wildtype cells hinting at a partial compensation of the loss of Inca1 by induction of p21. Loss of Inca1 combined with p21 knockdown synergistically increased S-phase. These results indicate that Inca1 could be functionally related to p21 and that the rather mild phenotype observed in Inca1−/ − mice and the modest differences in Cdk activity observed in cell lysates lacking Inca1 could be due to compensatory induction of the CKI p21. In summary, loss of Inca1 increased cell proliferation, replating efficiency, S-phase progression, and Cdk2 activity whereas gain of Inca1 suppressed these cell functions. Inca1 expression was induced during cell cycle arrest. We conclude that Inca1 could be a novel cell cycle suppressor regulating the quiescence of HSCs through the inhibition of Cdk2.

scholarly journals A Sequence Element Downstream of the Yeast HTB1 Gene Contributes to mRNA 3′ Processing and Cell Cycle Regulation

2002 ◽  
Vol 22 (24) ◽  
pp. 8415-8425 ◽  
Author(s):  
Susan G. Campbell ◽  
Marcel li del Olmo ◽  
Paul Beglan ◽  
Ursula Bond
Keyword(s):  
Cell Cycle ◽  
Site Selection ◽  
Cleavage Site ◽  
Cell Cycle Regulation ◽  
Binding Activity ◽  
S Phase ◽  
Sequence Element ◽  
Reading Frame ◽  
Protein Factor ◽  
Cycle Regulation

ABSTRACT Histone mRNAs accumulate in the S phase and are rapidly degraded as cells progress into the G2 phase of the cell cycle. In Saccharomyces cerevisiae, fusion of the 3′ untranslated region and downstream sequences of the yeast histone gene HTB1 to a neomycin phosphotransferase open reading frame is sufficient to confer cell cycle regulation on the resulting chimera gene (neo-HTB1). We have identified a sequence element, designated the distal downstream element (DDE), that influences both the 3′-end cleavage site selection and the cell cycle regulation of the neo-HTB1 mRNA. Mutations in the DDE, which is located approximately 110 nucleotides downstream of the HTB1 gene, lead to a delay in the accumulation of the neo-HTB1 mRNA in the S phase and a lack of mRNA turnover in the G2 phase. The DDE is transcribed as part of the primary transcript and binds a protein factor(s). Maximum binding is observed in the S phase of the cell cycle, and mutations that affect the turnover of the HTB1 mRNA alter the binding activity. While located in the same general region, mutations that affect 3′-end cleavage site selection act independently from those that alter the cell cycle regulation.

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