The Experts below are selected from a list of 1681098 Experts worldwide ranked by ideXlab platform
Cyrus Vaziri - One of the best experts on this subject based on the ideXlab platform.
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the chk1 mediated S phaSe checkpoint targetS initiation factor cdc45 via a cdc25a cdk2 independent mechaniSm
Journal of Biological Chemistry, 2006Co-Authors: Peijun Liu, Mark G Alexandrow, Laura R Barkley, Tovah A Day, Damien M Slater, Heinzpeter Nasheuer, Cyrus VaziriAbstract:DNA damage induced by the carcinogen benzo[a]pyrene dihydrodiol epoxide (BPDE) induceS a Chk1-dependent S-phaSe checkpoint. Here, we have inveStigated the molecular baSiS of BPDE-induced S-phaSe arreSt. Chk1-dependent inhibition of DNA SyntheSiS in BPDE-treated cellS occurred without detectable changeS in Cdc25A levelS, Cdk2 activity, or Cdc7/Dbf4 interaction. OverexpreSSion StudieS Showed that Cdc25A, cyclin A/Cdk2, and Cdc7/Dbf4 were not rate-limiting for DNA SyntheSiS when the BPDE-induced S-phaSe checkpoint waS active. To inveStigate other potential targetS of the S-phaSe checkpoint, we teSted the effectS of BPDE on the chromatin aSSociation of DNA replication factorS. The levelS of chromatin-aSSociated Cdc45 (but not Soluble Cdc45) were reduced concomitantly with BPDE-induced Chk1 activation and inhibition of DNA SyntheSiS. The chromatin aSSociation of Mcm7, Mcm10, and proliferating cell nuclear antigen waS unaffected by BPDE treatment. However, the aSSociation between Mcm7 and Cdc45 in the chromatin fraction waS inhibited in BPDE-treated cellS. Chromatin immunoprecipitation analySeS demonStrated reduced aSSociation of Cdc45 with the β-globin origin of replication in BPDE-treated cellS. The inhibitory effectS of BPDE on DNA SyntheSiS, Cdc45/Mcm7 aSSociationS, and interactionS between Cdc45 and the β-globin locuS were abrogated by the Chk1 inhibitor UCN-01. Taken together, our reSultS Show that the aSSociation between Cdc45 and Mcm7 at originS of replication iS negatively regulated by Chk1 in a Cdk2-independent manner. Therefore, Cdc45 iS likely to be an important target of the Chk1-mediated S-phaSe checkpoint.
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critical role for mouSe huS1 in an S phaSe dna damage cell cycle checkpoint
Molecular and Cellular Biology, 2003Co-Authors: Robert S. Weiss, Philip Leder, Cyrus VaziriAbstract:MouSe HuS1 encodeS an evolutionarily conServed DNA damage reSponSe protein. In thiS Study we examined how targeted deletion of HuS1 affectS cell cycle checkpoint reSponSeS to genotoxic StreSS. Unlike huS1− fiSSion yeaSt (SchizoSaccharomyceS pombe) cellS, which are defective for the G2/M DNA damage checkpoint, HuS1-null mouSe cellS did not inappropriately enter mitoSiS following genotoxin treatment. However, HuS1-deficient cellS diSplayed a Striking S-phaSe DNA damage checkpoint defect. WhereaS wild-type cellS tranSiently repreSSed DNA replication in reSponSe to benzo(a)pyrene dihydrodiol epoxide (BPDE), a genotoxin that cauSeS bulky DNA adductS, HuS1-null cellS maintained relatively high levelS of DNA SyntheSiS following treatment with thiS agent. However, when treated with DNA Strand break-inducing agentS Such aS ionizing radiation (IR), HuS1-deficient cellS Showed intact S-phaSe checkpoint reSponSeS. ConverSely, checkpoint-mediated inhibition of DNA SyntheSiS in reSponSe to BPDE did not require NBS1, a component of the IR-reSponSive S-phaSe checkpoint pathway. Taken together, theSe reSultS demonStrate that HuS1 iS required Specifically for one of two Separable mammalian checkpoint pathwayS that reSpond to diStinct formS of genome damage during S phaSe.
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critical role for mouSe huS1 in an S phaSe dna damage cell cycle checkpoint
Molecular and Cellular Biology, 2003Co-Authors: Robert S. Weiss, Philip Leder, Cyrus VaziriAbstract:MouSe HuS1 encodeS an evolutionarily conServed DNA damage reSponSe protein. In thiS Study we examined how targeted deletion of HuS1 affectS cell cycle checkpoint reSponSeS to genotoxic StreSS. Unlike huS1− fiSSion yeaSt (SchizoSaccharomyceS pombe) cellS, which are defective for the G2/M DNA damage checkpoint, HuS1-null mouSe cellS did not inappropriately enter mitoSiS following genotoxin treatment. However, HuS1-deficient cellS diSplayed a Striking S-phaSe DNA damage checkpoint defect. WhereaS wild-type cellS tranSiently repreSSed DNA replication in reSponSe to benzo(a)pyrene dihydrodiol epoxide (BPDE), a genotoxin that cauSeS bulky DNA adductS, HuS1-null cellS maintained relatively high levelS of DNA SyntheSiS following treatment with thiS agent. However, when treated with DNA Strand break-inducing agentS Such aS ionizing radiation (IR), HuS1-deficient cellS Showed intact S-phaSe checkpoint reSponSeS. ConverSely, checkpoint-mediated inhibition of DNA SyntheSiS in reSponSe to BPDE did not require NBS1, a component of the IR-reSponSive S-phaSe checkpoint pathway. Taken together, theSe reSultS demonStrate that HuS1 iS required Specifically for one of two Separable mammalian checkpoint pathwayS that reSpond to diStinct formS of genome damage during S phaSe.
Jiri Lukas - One of the best experts on this subject based on the ideXlab platform.
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human aSf1 regulateS the flow of S phaSe hiStoneS during replicational StreSS
Molecular Cell, 2005Co-Authors: Anja Groth, Jiri Lukas, Jiri Bartek, Dominique Raygallet, Jeanpierre Quivy, Genevieve AlmouzniAbstract:Maintenance of chromoSomal integrity requireS tight coordination of hiStone bioSyntheSiS with DNA replication. Here, we Show that extractS from human cellS expoSed to replication StreSS diSplay an increaSed capacity to Support replication-coupled chromatin aSSembly. While in unperturbed S phaSe, hASf1 exiSted in equilibrium between an active form and an inactive hiStone-free pool, replication StreSS mobilized the majority of hASf1 into an active multichaperone complex together with hiStoneS. ThiS active multichaperone complex waS limiting for chromatin aSSembly in S phaSe extractS, and hASf1 waS required for the enhanced aSSembly activity in cellS expoSed to replication StreSS. ConSiStently, SiRNA-mediated knockdown of hASf1 impaired the kineticS of S phaSe progreSSion. Together, theSe data SuggeSt that hASf1 provideS the cellS with a buffering SyStem for hiStone exceSS generated in reSponSe to Stalled replication and explainS how mammalian cellS maintain a critical "active" hiStone pool available for depoSition during recovery from replication StreSSeS.
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checking on dna damage in S phaSe
Nature Reviews Molecular Cell Biology, 2004Co-Authors: Jiri Bartek, Claudia Lukas, Jiri LukasAbstract:The preciSe replication of the genome and the continuouS Surveillance of itS integrity are eSSential for Survival and the avoidance of variouS diSeaSeS. CellS reSpond to DNA damage by activating a complex network of the So-called checkpoint pathwayS to delay their cell-cycle progreSSion and repair the defectS. In thiS review we integrate findingS on the emerging mechaniSmS of activation, the Signalling pathwayS and the Spatio-temporal organization of the intra-S-phaSe DNA-damage checkpoint and itS impact on the cell-cycle machinery, and diScuSS itS biological Significance.
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e2f dependent accumulation of hemi1 regulateS S phaSe entry by inhibiting apc cdh1
Nature Cell Biology, 2002Co-Authors: Jerry Y Hsu, Claus Storgaard Sorensen, Julie D R Reimann, Jiri Lukas, P JacksonAbstract:Emi1 promoteS mitotic entry in XenopuS laeviS embryoS by inhibiting the APCCdc20 ubiquitination complex to allow accumulation of cyclin B. We Show here that human Emi1 (hEmi1) functionS to promote cyclin A accumulation and S phaSe entry in Somatic cellS by inhibiting the APCCdh1 complex. At the G1–S tranSition, hEmi1 iS tranScriptionally induced by the E2F tranScription factor, much like cyclin A. hEmi1 overexpreSSion accelerateS S phaSe entry and can override a G1 block cauSed by overexpreSSion of Cdh1 or the E2F-inhibitor p105 retinoblaStoma protein (pRb). Depleting cellS of hEmi1 through RNA interference preventS accumulation of cyclin A and inhibitS S phaSe entry. TheSe data SuggeSt that E2F can activate both tranScription of cyclin A and the hEmi1-dependent Stabilization of APCCdh1 targetS, Such aS cyclin A, to promote S phaSe entry.
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mammalian g1 and S phaSe checkpointS in reSponSe to dna damage
Current Opinion in Cell Biology, 2001Co-Authors: Jiri Bartek, Jiri LukasAbstract:The ability to preServe genomic integrity iS a fundamental feature of life. Recent findingS regarding the molecular baSiS of the cell-cycle checkpoint reSponSeS of mammalian cellS to genotoxic StreSS have converged into a two-wave concept of the G1 checkpoint, and Shed light on the So-far eluSive intra-S-phaSe checkpoint. Rapidly operating caScadeS that target the Cdc25A phoSphataSe appear central in both the initiation wave of the G1 checkpoint (preceding the p53-mediated maintenance wave) and the tranSient intra-S-phaSe reSponSe. Multiple linkS between defectS in the G1/S checkpointS, genomic inStability and oncogeneSiS are emerging, aS are new challengeS and hopeS raiSed by thiS knowledge.
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cyclin e induced S phaSe without activation of the prb e2f pathway
Genes & Development, 1997Co-Authors: Jiri Lukas, Steven I Reed, Kristian Helin, Thomas Herzinger, Klaus Hansen, Maria Cristina Moroni, Dalia Resnitzky, Jiri BartekAbstract:In cellS of higher eukaryoteS, cyclin D-dependent kinaSeS Cdk4 and Cdk6 and, poSSibly, cyclin E-dependent Cdk2 poSitively regulate the G1- to S-phaSe tranSition, by phoSphorylating the retinoblaStoma protein (pRb), thereby releaSing E2F tranScription factorS that control S-phaSe geneS. Here we performed microinjection and tranSfection experimentS uSing rat R12 fibroblaStS, their derivativeS conditionally overexpreSSing cyclinS D1 or E, and human U-2-OS cellS, to explore the action of G1 cyclinS and the relationShip of E2F and cyclin E in S-phaSe induction. We demonStrate that ectopic expreSSion of cyclin E, but not cyclin D1, can override G1 arreSt impoSed by either the p16INK4a Cdk inhibitor Specific for Cdk4 and Cdk6 or a novel phoSphorylation-deficient mutant pRb. Several complementary approacheS to aSSeSS E2F activation, including quantitative reporter aSSayS in live cellS, Showed that the cyclin E-induced S phaSe and completion of the cell diviSion cycle can occur in the abSence of E2F-mediated tranSactivation. Together with the ability of cyclin E to overcome a G1 block induced by expreSSion of dominant-negative mutant DP-1, a heterodimeric partner of E2FS, theSe reSultS provide evidence for a cyclin E-controlled S phaSe-promoting event in Somatic cellS downStream of or parallel to phoSphorylation of pRb and independent of E2F activation. They furthermore indicate that a lack of E2F-mediated tranSactivation can be compenSated by hyperactivation of thiS cyclin E-controlled event.
Paul Nurse - One of the best experts on this subject based on the ideXlab platform.
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The Spatial and temporal organization of origin firing during the S-phaSe of fiSSion yeaSt
Genome research, 2015Co-Authors: Atanas Kaykov, Paul NurseAbstract:EukaryoteS duplicate their genomeS uSing multiple replication originS, but the organization of origin firing along chromoSomeS and during S-phaSe iS not well underStood. USing fiSSion yeaSt, we report the firSt genome-wide analySiS of the Spatial and temporal organization of replication origin firing, analyzed uSing Single DNA moleculeS that can approach the full length of chromoSomeS. At S-phaSe onSet, originS fire randomly and SparSely throughout the chromoSomeS. Later in S-phaSe, cluSterS of fired originS appear embedded in the SparSer regionS, which form the baSiS of nuclear replication foci. The formation of cluSterS requireS proper hiStone methylation and acetylation, and their locationS are not inherited between cell cycleS. The rate of origin firing increaSeS gradually, peaking juSt before mid S-phaSe. Toward the end of S-phaSe, nearly all the available originS within the unreplicated regionS are fired, contributing to the timely completion of genome replication. We propoSe that the majority of originS do not fire aS a part of a determiniStic program. InStead, origin firing, both individually and aS cluSterS, Should be viewed aS being moStly StochaStic.
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cyclin dependent kinaSe inhibitS reinitiation of a normal S phaSe program during g2 in fiSSion yeaSt
Molecular and Cellular Biology, 2009Co-Authors: Lee Kiang, Christian Heichinger, Stephen Watt, Jurg Bahler, Paul NurseAbstract:To achieve faithful replication of the genome once in each cell cycle, reinitiation of S phaSe iS prevented in G(2) and originS are reStricted from refiring within S phaSe. We have inveStigated the block to rereplication during G(2) in fiSSion yeaSt. The DNA SyntheSiS that occurS when G(2)/M cyclin-dependent kinaSe (CDK) activity iS depleted haS been aSSumed to be repeated roundS of S phaSe without mitoSiS, but thiS haS not been demonStrated to be the caSe. We Show here that on G(2)/M CDK depletion in G(2), repeated S phaSeS are induced, which are correlated with normal G(1)/S tranScription and attainment of doublingS in cell Size. MoStly normal mitotic S-phaSe originS are utilized, although at different efficiencieS, and replication iS eSSentially equal acroSS the genome. We conclude that CDK inhibitS reinitiation of S phaSe during G(2), and if G(2)/M CDK iS depleted, replication reSultS from induction of a largely normal S-phaSe program with only Small differenceS in origin uSage and efficiency.
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pre meiotic S phaSe iS linked to reductional chromoSome Segregation and recombination
Nature, 2001Co-Authors: Yoshinori Watanabe, Masayuki Yamamoto, Shihori Yokobayashi, Paul NurseAbstract:MeioSiS iS initiated from G1 of the cell cycle and iS characterized by a pre-meiotic S phaSe followed by two SucceSSive nuclear diviSionS. The firSt of theSe, meioSiS I, differS from mitoSiS in having a reductional pattern of chromoSome Segregation. Here we Show that meioSiS can be initiated from G2 in fiSSion yeaSt cellS by ectopically activating the meioSiS-inducing network. The SubSequent meioSiS I occurS without a pre-meiotic S phaSe and with decreaSed recombination, and exhibitS a mitotic pattern of equational chromoSome Segregation. The SubSequent meioSiS II reSultS in random chromoSome Segregation. ThiS behaviour iS Similar to that obServed in cellS lacking the meiotic coheSin Rec8 (refS 3, 4), which becomeS aSSociated with chromoSomeS at G1/S phaSe, including the inner centromere, a region that iS probably critical for SiSter-centromere orientation. If the expreSSion of Rec8 iS delayed to S phaSe/G2, then the centromereS behave equationally. We propoSe that the preSence of Rec8 in chromatin iS required at the pre-meiotic S phaSe to conStruct centromereS that behave reductionally and chromoSome armS capable of a high level of recombination, and that thiS explainS why meioSiS iS initiated from G1 of the cell cycle.
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the fiSSion yeaSt cdc18 gene product coupleS S phaSe to Start and mitoSiS
Cell, 1993Co-Authors: Thomas J Kelly, Steven G Martin, Susan L Forsburg, Robert J Stephen, Alicia Russo, Paul NurseAbstract:Commitment to the cell cycle in fiSSion yeaSt requireS the function of the cdc10+ tranScriptional activator at START. The product of the cdc18+ gene iS a major downStream target of cdc10+, and tranScription of cdc18+ iS activated by cdc10+ during paSSage through START. The cdc18+ function iS required for entry into S phaSe. In addition, the product of the cdc18+ gene iS part of the checkpoint control that preventS mitoSiS from occurring until S phaSe iS completed. ThuS, cdc18+ playS a key role in coupling S phaSe to START and mitoSiS.
David J Hall - One of the best experts on this subject based on the ideXlab platform.
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an early S phaSe checkpoint iS regulated by the e2f1 tranScription factor
Biochemical and Biophysical Research Communications, 1999Co-Authors: Matthew C Stubbs, Gordon D Strachan, David J HallAbstract:AbStract The E2F1 tranScription factor regulateS tranSit of cellS through the S phaSe checkpoint, dependent on itS aSSociation with cyclin A/cdk2. ExpreSSion in cellS of a mutant E2F1 lacking the cyclin A/cdk2 binding domain leadS to partial arreSt of cellS at the S phaSe checkpoint. When Subconfluent growing cellS expreSSing thiS mutant E2F1 are analyzed in detail, it iS Shown here that they diSplay a Significantly reduced incorporation of 3 H-thymidine into the DNA of each S phaSe cell, compared to control cellS or to cellS overexpreSSing full-length E2F1. Further, when cellS are blocked at the G1/S phaSe border and releaSed, there iS a clear reduction in the amount of 3 H-thymidine incorporated into the DNA of S phaSe cellS by 1.5 hourS poSt releaSe. ConSidering a normal 6 hour S phaSe duration, the reSultS Show that the S phaSe checkpoint mediated by E2F1 iS not a late S phaSe event but an early one.
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expreSSion of a deletion mutant of the e2f1 tranScription factor in fibroblaStS lengthenS S phaSe and increaSeS SenSitivity to S phaSe Specific toxinS
Cancer Research, 1995Co-Authors: Thomas J Logan, Devon L Evans, W E Mercer, Maryann Bjornsti, David J HallAbstract:To better underStand how the E2F1 tranScription factor contributeS to the proceSS of cell proliferation, NIH-3T3 cell lineS were generated that conStitutively expreSS either the wild-type E2F1 protein or an amino terminal deletion mutant, termed E2F1d87. Proliferating E2F1d87-expreSSing cellS exhibit a Significant lengthening of S phaSe relative to control and E2F1 cell lineS and are hyperSenSitive to the cytotoxic effectS of the S phaSe-Specific antitumor drug camptothecin. ThiS SenSitivity iS aSSociated with an increaSe in drug-induced p53 and WAF1 levelS. The E2F1 and E2F1d87 cell lineS are both able to initiate, but not complete, S phaSe under conditionS of Serum Starvation. However, quantitation of DNA SyntheSiS, during culture in Serum-deprived media, indicateS that the E2F1d87 cell line SyntheSizeS more DNA/cell aS compared to the E2F1 cell line. ConSiStent with thiS relative increaSe in DNA SyntheSiS, the E2F1d87 cell line undergoeS camptothecin-induced apoptoSiS when cultured under conditionS of Serum Starvation, while the control and E2F1 cell lineS are unaffected by drug treatment under the Same conditionS. ThuS, the SenSitivity of the E2F1d87 cell line to camptothecin iS not dependent on cell proliferation. The data preSented here SuggeSt that cell cycle parameterS can be manipulated in order to enhance SenSitivity of a cell to the toxic effectS of Specific chemotherapeutic agentS.
Jiri Bartek - One of the best experts on this subject based on the ideXlab platform.
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human aSf1 regulateS the flow of S phaSe hiStoneS during replicational StreSS
Molecular Cell, 2005Co-Authors: Anja Groth, Jiri Lukas, Jiri Bartek, Dominique Raygallet, Jeanpierre Quivy, Genevieve AlmouzniAbstract:Maintenance of chromoSomal integrity requireS tight coordination of hiStone bioSyntheSiS with DNA replication. Here, we Show that extractS from human cellS expoSed to replication StreSS diSplay an increaSed capacity to Support replication-coupled chromatin aSSembly. While in unperturbed S phaSe, hASf1 exiSted in equilibrium between an active form and an inactive hiStone-free pool, replication StreSS mobilized the majority of hASf1 into an active multichaperone complex together with hiStoneS. ThiS active multichaperone complex waS limiting for chromatin aSSembly in S phaSe extractS, and hASf1 waS required for the enhanced aSSembly activity in cellS expoSed to replication StreSS. ConSiStently, SiRNA-mediated knockdown of hASf1 impaired the kineticS of S phaSe progreSSion. Together, theSe data SuggeSt that hASf1 provideS the cellS with a buffering SyStem for hiStone exceSS generated in reSponSe to Stalled replication and explainS how mammalian cellS maintain a critical "active" hiStone pool available for depoSition during recovery from replication StreSSeS.
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checking on dna damage in S phaSe
Nature Reviews Molecular Cell Biology, 2004Co-Authors: Jiri Bartek, Claudia Lukas, Jiri LukasAbstract:The preciSe replication of the genome and the continuouS Surveillance of itS integrity are eSSential for Survival and the avoidance of variouS diSeaSeS. CellS reSpond to DNA damage by activating a complex network of the So-called checkpoint pathwayS to delay their cell-cycle progreSSion and repair the defectS. In thiS review we integrate findingS on the emerging mechaniSmS of activation, the Signalling pathwayS and the Spatio-temporal organization of the intra-S-phaSe DNA-damage checkpoint and itS impact on the cell-cycle machinery, and diScuSS itS biological Significance.
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mammalian g1 and S phaSe checkpointS in reSponSe to dna damage
Current Opinion in Cell Biology, 2001Co-Authors: Jiri Bartek, Jiri LukasAbstract:The ability to preServe genomic integrity iS a fundamental feature of life. Recent findingS regarding the molecular baSiS of the cell-cycle checkpoint reSponSeS of mammalian cellS to genotoxic StreSS have converged into a two-wave concept of the G1 checkpoint, and Shed light on the So-far eluSive intra-S-phaSe checkpoint. Rapidly operating caScadeS that target the Cdc25A phoSphataSe appear central in both the initiation wave of the G1 checkpoint (preceding the p53-mediated maintenance wave) and the tranSient intra-S-phaSe reSponSe. Multiple linkS between defectS in the G1/S checkpointS, genomic inStability and oncogeneSiS are emerging, aS are new challengeS and hopeS raiSed by thiS knowledge.
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cyclin e induced S phaSe without activation of the prb e2f pathway
Genes & Development, 1997Co-Authors: Jiri Lukas, Steven I Reed, Kristian Helin, Thomas Herzinger, Klaus Hansen, Maria Cristina Moroni, Dalia Resnitzky, Jiri BartekAbstract:In cellS of higher eukaryoteS, cyclin D-dependent kinaSeS Cdk4 and Cdk6 and, poSSibly, cyclin E-dependent Cdk2 poSitively regulate the G1- to S-phaSe tranSition, by phoSphorylating the retinoblaStoma protein (pRb), thereby releaSing E2F tranScription factorS that control S-phaSe geneS. Here we performed microinjection and tranSfection experimentS uSing rat R12 fibroblaStS, their derivativeS conditionally overexpreSSing cyclinS D1 or E, and human U-2-OS cellS, to explore the action of G1 cyclinS and the relationShip of E2F and cyclin E in S-phaSe induction. We demonStrate that ectopic expreSSion of cyclin E, but not cyclin D1, can override G1 arreSt impoSed by either the p16INK4a Cdk inhibitor Specific for Cdk4 and Cdk6 or a novel phoSphorylation-deficient mutant pRb. Several complementary approacheS to aSSeSS E2F activation, including quantitative reporter aSSayS in live cellS, Showed that the cyclin E-induced S phaSe and completion of the cell diviSion cycle can occur in the abSence of E2F-mediated tranSactivation. Together with the ability of cyclin E to overcome a G1 block induced by expreSSion of dominant-negative mutant DP-1, a heterodimeric partner of E2FS, theSe reSultS provide evidence for a cyclin E-controlled S phaSe-promoting event in Somatic cellS downStream of or parallel to phoSphorylation of pRb and independent of E2F activation. They furthermore indicate that a lack of E2F-mediated tranSactivation can be compenSated by hyperactivation of thiS cyclin E-controlled event.
