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PubMedDate not supplied

Chkl binds and phosphorylates BAD protein.

Chk1 (checkpoint kinase 1) is a serine-threonine kinase that is critical for G2/M arrest in response to DNA damage. Chk1 phosphorylates Cdc25C at serine-216, a major regulatory site, in response to DNA damage. Furthermore, Chk1 also phosphorylates Cdc25A on serine 123 which accelerates its degradation through the ubiquitin-proteasome pathway and arrests cells in late G2-phase after DNA damage. In the present study, we demonstrated that Chk1 phosphorylates pro-apoptotic protein BAD (Bcl-2/Bcl-XL-Antagonist, causing cell Death) in vitro. In vitro phosphorylation analysis with various mouse BAD peptides has revealed two phosphorylation sites for Chk1 at serine-155 and serine-170. When wild-type and mutant BAD (S155A) constructs were transfected into 293T cells, an association between BAD and Chk1 was observed by co-immunoprecipitation. In addition, there was an increase in the phosphorylation of serine-155 following DNA damage by adriamycin treatment. Our results suggest that Chk1 associates with BAD and phosphorylates the BAD protein at serine-155. Taken together, our results suggest that Chk1 may inactivate BAD by associating with and phosphorylating residues critical for BAD function in response to DNA damage.

Amino Acid Sequence↗
PubMed2006

Dissecting the roles of checkpoint kinase 1/CDC2 and mitogen-activated protein kinase kinase 1/2/extracellular signal-regulated kinase 1/2 in relation to 7-hydroxystaurosporine-induced apoptosis in human multiple myeloma cells.

The functional roles of Cdc2 and checkpoint kinase 1 (Chk1) in synergistic interactions between 7-hydroxystaurosporine (UCN-01) and mitogen-activated protein kinase kinase 1/2 (MEK1/2) inhibitors [e.g., 2-(2-chloro-4-iodophenylamino)-N-cyclopropylmethoxy-3,4-difluorobenzamide (PD184352)] were examined in human multiple myeloma cells in relation to MEK1/2/ERK1/2 activation and lethality. Time course studies revealed that MEK1/2/extracellular signal-regulated kinase 1/2 (ERK1/2) phosphorylation preceded Cdc2 dephosphorylation (Tyr15) after UCN-01 exposure. Furthermore, enforced expression of Cdc2 or small inducible RNA (siRNA)-mediated Cdc2 knockdown failed to modify ERK1/2 activation status in either the presence or absence of UCN-01, arguing against a causal relationship between these events. However, ectopic expression of Cdc2 sensitized cells to the lethality of UCN-01/MEK inhibitor regimen, whereas Cdc2 knockdown by siRNA significantly diminished the lethal effects of this combination. Conversely, Chk1 knockdown by siRNA enhanced lethality mediated by UCN-01/PD184352. It is interesting that Chk1 knockdown reduced basal ERK1/2 activation and antagonized the ability of UCN-01 to activate ERK1/2. Finally, ectopic expression of constitutively active MEK1 significantly protected cells from the UCN-01/MEK1/2 inhibitor regimen without modifying Cdc2 activation status. Together, these findings indicate that although UCN-01-mediated Chk1 inhibition and Cdc2 activation are unlikely to be responsible for MEK1/2/ERK1/2 activation, both of these events contribute functionally to enhanced lethality in cells coexposed to MEK inhibitors. They also suggest a role for Chk1 in UCN-01-induced ERK1/2 activation, implying the existence of a heretofore unrecognized link between Chk1 and ERK1/2 signaling.

Apoptosis↗
PubMed2006

Influence of chk1 and plk1 silencing on radiation- or cisplatin-induced cytotoxicity in human malignant cells.

The G2/M checkpoint is an attractive pathway for targeting and sensitizing tumor cells to cancer treatment. Abrogation of the G2/M checkpoint by targeting molecules, such as checkpoint kinase 1 (chk1), increases DNA breakage and sensitizes tumor cells to anti-tumoral agents. However, most of the previously described G2/M abrogators are actually targeting the G2-M border checkpoints rather than mitotic checkpoints. This prompted us to test the effects of combined targeting of chk1 and a critical regulator of mitosis, polo-like kinase 1 (plk1). Chk1 and plk1 were found to be co-expressed in 70% of primary neoplastic tissues we examined. Asynchronized tumor cells were treated with different DNA damaging-agents to activate G1/S, S or G2/M checkpoints. Either chk1 or plk1-specific antisense oligodeoxynucleotides (ASODN) enhanced DNA damaging agent-induced apoptosis. When used in combination, however, chk1- plus plk1-specific ASODN failed to produce synergistic effects. Moreover, selective targeting of plk1 or chk1 in tumor xenografts of mice by oncolytic adenovirus mutants demonstrated potent anti-tumoral efficacy in the presence of low dose cisplatin. Again, combined targeting of chk1 and plk1 did not further enhance anti-tumoral efficacy. We concluded that combined targeting of chk1 and plk1 was not superior to either targeting chk1 or plk1 alone, which suggested that chk1 and plk1 silencing might overlap in their mechanism of action. Whether combined targeting of chk1 with other, more specific mitotic regulators would synergistically sensitize tumor to anti-neoplastic therapeutics needs to be further clarified.

Animals↗
PubMed2004

Mechanism of cell cycle arrest by sulfoquinovosyl monoacylglycerol with a C18-saturated fatty acid (C18-SQMG).

We have screened the inhibitors of mammalian DNA polymerases from natural products, and in the process found that either sulfoglycolipids or sulfoquinovosyl monoacylglycerol with a C18-saturated fatty acid (C18-SQMG), potently and selectively inhibited the activity of mammalian DNA polymerase (pol) and moderately the pol alpha. C18-SQMG was a cancer cell growth suppressor and a promissive anti-tumor agent. The purpose of this study was to elucidate the cell growth inhibition mechanism of C18-SQMG using HeLa cells. Analyses of the cell cycle and cyclin expression suggested that C18-SQMG arrested the cell cycle at intra-S phase, and the inhibition manner of DNA replication by C18-SQMG was similar to that by hydroxyurea. However, the DNA replication block by C18-SQMG did not induce degradation of Cdc25A protein, which was required for the replication block by hydroxyurea. C18-SQMG somewhat delayed mitosis because it induced phosphorylation of protein kinases, such as checkpoint kinases 1 and 2. These results suggest that C18-SQMG at first blocked DNA replication at the S phase by inhibiting replicative DNA polymerases, such as alpha, and then as the result of the inhibition, the other checkpoint signals associated with the pol might have responded.

Cell Cycle↗
PubMed2002

The identification and characterization of the marine natural product scytonemin as a novel antiproliferative pharmacophore.

Marine natural products provide a rich source of chemical diversity that can be used to design and develop new, potentially useful therapeutic agents. We report here that scytonemin, a pigment isolated from cyanobacteria, is the first described small molecule inhibitor of human polo-like kinase, a serine/threonine kinase that plays an integral role in regulating the G(2)/M transition in the cell cycle. Scytonemin inhibited polo-like kinase 1 activity in a concentration-dependent manner with an IC(50) of 2 microM against the recombinant enzyme. Biochemical analysis showed that scytonemin reduced GST-polo-like kinase 1 activity in a time-independent fashion, suggesting reversibility, and with a mixed-competition mechanism with respect to ATP. Although scytonemin was less potent against protein kinase A and Tie2, a tyrosine kinase, it did inhibit other cell cycle-regulatory kinases like Myt1, checkpoint kinase 1, cyclin-dependent kinase 1/cyclin B, and protein kinase Cbeta2 with IC(50) values similar to that seen for polo-like kinase 1. Consistent with these effects, scytonemin effectively attenuated, without chemical toxicity, the growth factor- or mitogen-induced proliferation of three cell types commonly implicated in inflammatory hyperproliferation. Similarly, scytonemin (up to 10 microM) was not cytotoxic to nonproliferating endotoxin-stimulated human monocytes. In addition, Jurkat T cells treated with scytonemin were induced to undergo apoptosis in a non-cell cycle-dependent manner consistent with its activities on multiple kinases. Here we propose that scytonemin's dimeric structure, unique among natural products, may be a valuable template for the development of more potent and selective kinase inhibitors used for the treatment of hyperproliferative disorders.

Antineoplastic Agents↗
PubMed2006

DNA methyltransferase 1 knockdown activates a replication stress checkpoint.

DNA methyltransferase 1 (DNMT1) is an important component of the epigenetic machinery and is responsible for copying DNA methylation patterns during cell division. Coordination of DNA methylation and DNA replication is critical for maintaining epigenetic programming. Knockdown of DNMT1 leads to inhibition of DNA replication, but the mechanism has been unclear. Here we show that depletion of DNMT1 with either antisense or small interfering RNA (siRNA) specific to DNMT1 activates a cascade of genotoxic stress checkpoint proteins, resulting in phosphorylation of checkpoint kinases 1 and 2 (Chk1 and -2), gammaH2AX focus formation, and cell division control protein 25a (CDC25a) degradation, in an ataxia telangiectasia mutated-Rad3-related (ATR)-dependent manner. siRNA knockdown of ATR blocks the response to DNMT1 depletion; DNA synthesis continues in the absence of DNMT1, resulting in global hypomethylation. Similarly, the response to DNMT1 knockdown is significantly attenuated in human mutant ATR fibroblast cells from a Seckel syndrome patient. This response is sensitive to DNMT1 depletion, independent of the catalytic domain of DNMT1, as indicated by abolition of the response with ectopic expression of either DNMT1 or DNMT1 with the catalytic domain deleted. There is no response to short-term treatment with 5-aza-deoxycytidine (5-aza-CdR), which causes demethylation by trapping DNMT1 in 5-aza-CdR-containing DNA but does not cause disappearance of DNMT1 from the nucleus. Our data are consistent with the hypothesis that removal of DNMT1 from replication forks is the trigger for this response.

Animals↗
PubMed2005

Monochloramine inhibits ultraviolet B-induced p53 activation and DNA repair response in human fibroblasts.

Monochloramine (NH2Cl) is one of the inflammation-derived oxidants, and has various effects on cell cycle, apoptosis and signal transduction. We studied the effects of NH2Cl on DNA repair response induced by ultraviolet B (UVB) irradiation in normal human diploid fibroblasts, TIG-1. TIG-1 irradiated with 20 mJ/cm2 UVB showed marked increase in thymine dimer, which decreased by about 50% after 24 h. This decrease in thymine dimer was significantly attenuated (P < 0.05) by the pretreatment of NH2Cl (200 microM), which indicated DNA repair inhibition. UVB induced p53 phosphorylation at Ser15, Ser20 and Ser37, and p53 accumulation, and NH2Cl also inhibited these changes. Consequently, UVB-induced increase in the downstream effectors of p53, namely p21Cip1 and Gadd45a, were almost completely inhibited by NH2Cl. Immunoprecipitation study indicated that the association of p53 and MDM2, an E3 ubiquitin ligase for p53, did not change substantially by NH2Cl and/or UVB. The phosphorylation of p53 (Ser15 and Ser37) by UVB is catalyzed by ATR (ataxia telangiectasia mutated and Rad3 related kinase), which works as DNA damage sensor, and ATR also phosphorylates checkpoint kinase 1(Chk1) at Ser345. NH2Cl also inhibited the phosphorylation of Chk1 (Ser345). As UVB-induced DNA damage is repaired by nucleotide excision repair (NER) in human cells, these findings indicated that NH2Cl inhibited NER through the inhibition of p53 phosphorylation and accumulation, and NH2Cl probably impaired DNA damage recognition and/or ATR activation. NH2Cl may facilitate carcinogenesis through the inhibition of NER that repairs DNA damages from various carcinogens.

Blotting, Western↗
PubMed2004

Chk1, but not Chk2, is involved in the cellular response to DNA damaging agents: differential activity in cells expressing or not p53.

Mammalian Chk1 and Chk2 protein kinases are two important components of the G(2) DNA damage checkpoint. They are activated by upstream kinases (ataxia telangectasia mutated gene and ATM and Rad 3 related gene) and interfere with the activity of the cdc2/cyclinB1 complex, necessary for the G(2)-M transition, through the inactivation of the cdc25 phosphatases (cdc25A and cdc25C). To understand the role of Chk1 and Chk2 in the cellular response to different anticancer agents, we knocked down the expression of each protein or simultaneously of both proteins by using the small interfering RNA technique in the HCT-116 colon carcinoma cell line and in its isogenic systems in which p53 and p21 have been inactivated by targeted homologous recombination. We here show that inhibition of Chk1 but not of Chk2 in p21(-/-) and p53(-/-) cells caused a greater abrogation of G(2) block induced by ionizing radiation and cis-diamine-dichloroplatinum treatments and a greater sensitization to the same treatments than in the parental cell line with p53 and p21 wild type proteins. These data further emphasise the role of Chk1 as a molecular target to inhibit in tumors with a defect in the G(1) checkpoint with the aim of increasing the selectivity and specificity of anticancer drug treatments.

Antineoplastic Agents↗
PubMed2005

New opportunities in chemosensitization and radiosensitization: modulating the DNA-damage response.

Many current cancer treatments, including certain classes of chemotherapeutics and radiation, induce cytotoxicity by damaging DNA. However, many cancers are resistant to these therapies, which represents a significant challenge in the clinic. Thus, modulating DNA-damage responses to selectively enhance the sensitivity of cancer cells to these therapies is highly desirable. When DNA damage is detected, DNA checkpoint mechanisms are activated to halt cells at various phases of the cell cycle. Simultaneously, DNA-damage sensors transduce signals to activate DNA-repair mechanisms via de novo expression or post-translational modification of enzymes required for DNA repair. p53 is the major player in a checkpoint that arrests cells at the G1/S boundary, while checkpoint kinase (Chk)1 is critical for the G2/M checkpoint and also the S checkpoint that prevents cell cycle progression after replication defects (intra-S-phase checkpoint) or S/M uncoupling (S/M checkpoint). Poly(ADP-ribose) polymerase is involved in sensing DNA single-strand breaks and inducing DNA repair via poly(ADP-ribosyl)ating various DNA-binding and DNA-repair proteins. In this review, strategies for implementing small-molecule inhibitors of poly(ADP-ribose) polymerase and Chk1, which are emerging as potential adjuncts to current therapies, are discussed.

Antineoplastic Agents↗
PubMed2002

Disruption of the checkpoint kinase 1/cell division cycle 25A pathway abrogates ionizing radiation-induced S and G2 checkpoints.

Checkpoint kinase (Chk)1 is an evolutionarily conserved protein kinase that was first identified in fission yeast as an essential component of the DNA damage checkpoint. In mice, Chk1 provides an essential function in the absence of environmentally imposed genotoxic stress. Here we show that human cells lacking Chk1 exhibit defects in both the ionizing radiation (IR)-induced S and G(2) checkpoints. In addition, loss of Chk1 resulted in the accumulation of a hypophosphorylated form of the Cdc25A protein phosphatase, and Chk1-deficient cells failed to degrade Cdc25A after IR. The IR-induced S and G(2) checkpoints were partially restored in Chk1-deficient cells when Cdc25A accumulation was interfered with. Finally, Cdc25A was phosphorylated by Chk1 in vitro on similar sites phosphorylated in vivo, including serine-123. These findings indicate that Chk1 directly phosphorylates Cdc25A during an unperturbed cell cycle, and that phosphorylation of Cdc25A by Chk1 is required for cells to delay cell cycle progression in response to double-strand DNA breaks.

Cell Cycle↗
PubMed2003

Breast cancer-specific gene 1 interacts with the mitotic checkpoint kinase BubR1.

The abnormal expression of breast cancer-specific gene 1 (BCSG1) in malignant mammary epithelial cells is highly associated with the development and progression of breast cancer. A series of in vitro and in vivo studies performed in our laboratory and others have demonstrated that BCSG1 expression significantly stimulates proliferation, invasion, and metastasis of breast cancer cells. However, currently little is known about how BCSG1 exerts its oncogenic functions. To elucidate the cellular mechanisms underlying the effects of BCSG1 in breast cancer cells, we used a yeast two-hybrid system to screen for proteins that could associate with BCSG1. Through this screening, we identified the mitotic checkpoint protein BubR1 as a novel binding partner of BCSG1. The specific association of BCSG1 with BubR1 in breast cancer cells was demonstrated by immunoprecipitation and GST pull-down assays. Intriguingly, experiments conducted in four different cell lines all showed that exogenous expressions of BCSG1 consistently reduce the cellular levels of the BubR1 protein without affecting BubR1 mRNA expression. The tendency of endogenous BCSG1 expression coinciding with lower BubR1 protein levels was also observed in seven out of eight breast cancer cell lines. We further showed that the reducing effect of BCSG1 on BubR1 protein expression could be prevented by treating BCSG1-transfected cells with MG-132, a selective 26S proteasome inhibitor, implying that the proteasome machinery may be involved in the BCSG1-induced reduction of the BubR1 protein. Accompanied with a reduction of BubR1 protein level, BCSG1 expression resulted in multinucleation of breast cancer cells upon treatment with spindle inhibitor nocodazole, indicating an impaired mitotic checkpoint. Taken together, our novel findings suggest that BCSG1 may accelerate the progression of breast cancer at least in part by compromising the mitotic checkpoint control through inactivation of BubR1.

Cell Cycle Proteins↗
PubMed2003

Differential activation of the DNA replication checkpoint contributes to asynchrony of cell division in C. elegans embryos.

BACKGROUND: Acquisition of lineage-specific cell cycle duration is a central feature of metazoan development. The mechanisms by which this is achieved during early embryogenesis are poorly understood. In the nematode Caenorhabditis elegans, differential cell cycle duration is apparent starting at the two-cell stage, when the larger anterior blastomere AB divides before the smaller posterior blastomere P(1). How anterior-posterior (A-P) polarity cues control this asynchrony remains to be elucidated. RESULTS: We establish that early C. elegans embryos possess a hitherto unrecognized DNA replication checkpoint that relies on the PI-3-like kinase atl-1 and the kinase chk-1. We demonstrate that preferential activation of this checkpoint in the P(1) blastomere contributes to asynchrony of cell division in two-cell-stage wild-type embryos. Furthermore, we show that preferential checkpoint activation is largely abrogated in embryos that undergo equal first cleavage following inactivation of Galpha signaling. CONCLUSION: Our findings establish that differential checkpoint activation contributes to acquisition of distinct cell cycle duration in two-cell-stage C. elegans embryos and suggest a novel mechanism coupling asymmetric division to acquisition of distinct cell cycle duration during development.

Animals↗
PubMed1999

Drosophila grapes/CHK1 mutants are defective in cyclin proteolysis and coordination of mitotic events.

The Drosophila grapes (grp) gene, which encodes a homolog of the Schizosaccharomyces pombe Chk1 kinase, provides a cell-cycle checkpoint that delays mitosis in response to inhibition of DNA replication [1]. Grp is also required in the undisturbed early embryonic cycles: in its absence, mitotic abnormalities appear in cycle 12 and chromosomes fail to fully separate in subsequent cycles [2] [3]. In other systems, Chk1 kinase phosphorylates and suppresses the activity of Cdc25 phosphatase: the resulting failure to remove inhibitory phosphate from cyclin-dependent kinase 1 (Cdk1) prevents entry into mitosis [4] [5]. Because in Drosophila embryos Cdk1 lacks inhibitory phosphate during cycles 11-13 [6], it is not clear that known actions of Grp/Chk1 suffice in these cycles. We found that the loss of grp compromised cyclin A proteolysis and delayed mitotic disjunction of sister chromosomes. These defects occurred before previously reported grp phenotypes. We conclude that Grp activates cyclin A degradation, and functions to time the disjunction of chromosomes in the early embryo. As cyclin A destruction is required for sister chromosome separation [7], a failure in Grp-promoted cyclin destruction can also explain the mitotic phenotype. The mitotic failure described previously for cycle 12 grp embryos might be a more severe form of the phenotypes that we describe in earlier embryos and we suggest that the underlying defect is reduced degradation of cyclin A.

Anaphase↗
PubMed2004

Knockdown of Chk1, Wee1 and Myt1 by RNA interference abrogates G2 checkpoint and induces apoptosis.

Mammalian cells undergo cell cycle arrest in response to DNA damage due to the existence of multiple checkpoint response mechanisms. One such checkpoint pathway operating at the G(1) phase is frequently lost in cancer cells due to mutation of the p53 tumor suppressor gene. However, cancer cells often arrest at the G(2) phase upon DNA damage, due to activation of another checkpoint pathway that prevents the activation Cdc2 kinase. The kinases, Chk1, Wee1, and Myt1 are key regulators of this G(2) checkpoint, which act directly or indirectly to inhibit Cdc2 activity. Here we show that RNA interference (RNAi)-mediated downregulation of Wee1 kinase abrogated an Adriamycin trade mark -induced G(2) checkpoint in human cervical carcinoma Hela cells that are defective in G(1) checkpoint response. Wee1 downregulation sensitized HeLa cells to Adriamycin trade mark -induced apoptosis. Downregulation of Chk1 kinase in Hela cells also caused a significant amount of cell death in dependent of DNA damage. In contrast, Myt1 downregulation also abrogated Adriamycin trade mark -induced G(2) arrest but did not cause substantial apoptosis. Reduction in Wee1, Chk1, or Myt1 levels did not sensitize normal human mammary epithelial cells (HMEC) cells to Adriamycin trade mark -induced apoptosis unlike the situation in Hela cells. Our study reveals distinct roles for Chk1, Wee1, and Myt1 in G(2) checkpoint regulation. The data reported here support the attractiveness of Wee1 and Chk1 is as molecular targets for abrogating the G(2) DNA damage checkpoint arrest, a situation that may selectively sensitize p53-deficient tumor cells to radiation or chemotherapy treatment.

Apoptosis↗
PubMed2006

Low dietary folate initiates intestinal tumors in mice, with altered expression of G2-M checkpoint regulators polo-like kinase 1 and cell division cycle 25c.

Clinical reports have suggested that low dietary folate increases risk for colorectal cancer. Animal studies for investigation of folate and tumorigenesis have used carcinogen induction or mice with germ-line mutations. We have developed a new spontaneous tumor model in which mice, with or without a null allele in a key folate-metabolizing enzyme, methylenetetrahydrofolate reductase (Mthfr), develop intestinal tumors due to low dietary folate alone. On folate-deficient diets, 12.5% of Mthfr(+/+) mice and 28.1% of Mthfr(+/-) mice developed tumors; mice on control diets were negative. Dietary and genotype effects on tumor development were significant. To investigate mechanisms of folate-dependent tumorigenesis, we examined levels of DNA damage and gene expression of two genes involved in DNA damage response and G(2)-M checkpoint regulation, polo-like kinase 1 (Plk1) and cell division cycle 25c (Cdc25c). Folate deficiency increased DNA damage and decreased expression of both genes (assessed by quantitative reverse transcription-PCR and immunofluorescence) in normal intestine compared with levels in mice on control diets. An immunofluorescence assay for CDC25c activity (phosphorylated CDC2) also found CDC25c activity to be decreased in folate-deficient normal intestine. In tumors, however, Plk1 and Cdc25c mRNA were found to be higher (11- and 3-fold, respectively) compared with normal intestine from folate-deficient mice; immunofluorescence studies of PLK1, CDC25c, and phosphorylated CDC2 supported these findings. Our data suggest that folate deficiency can initiate tumor development, that Mthfr mutation can enhance this phenomenon, and that altered expression of Plk1 and Cdc25c may contribute to folate-dependent intestinal tumorigenesis.

Animals↗
PubMed2006

A stronger DNA damage-induced G2 checkpoint due to over-activated CHK1 in the absence of PARP-1.

Poly(ADP-ribose) polymerase 1 (PARP-1) is involved in multi-pathways to respond to DNA damage. Lack of or inhibition of PARP-1 activity leads to slow progress of cell cycle and sensitization of cells to different stresses. Recently, it was reported that besides the Ku dependent main nonhomologous end joining (NHEJ) pathway, there is a PARP-1 dependent complementary NHEJ pathway to repair DNA double strand break (DSB). Here we show that compared with PARP-1+/+ cells, PARP-1-/- cells display a much stronger G2 checkpoint response following ionizing radiation (IR). Treatment with Chk1 siRNA abolishes the stronger G2 checkpoint response and sensitizes PARP-1-/- cells to IR. These data indicate that the stronger G2 checkpoint response in PARP-1-/- cells is CHK1 dependent, which protects cells from IR induced killing. We also show that 4-Amino-1,8-naphthalimide (4-AN, inhibitor of PARP) but not methoxyamine (inhibitor of base excision repair (BER)), affects IR induced G2 arrest and cell sensitivity in PARP-1+/+ cells, resulting in the phenotypes similar to those of PARP-1-/- cells. These results indicate that DSB repair from the complementary NHEJ pathway of PARP-1, but not single strand break (SSB) repair from the BER function of PARP-1, may play an essential role in the over-activated CHK1 regulated G2 checkpoint response and radiosensitivity in PARP-1-/- cells.

1-Naphthylamine↗
PubMed1999

Role of c-Jun N-terminal kinase 1 (JNK1) in cell cycle checkpoint activated by the protease inhibitor N-acetyl-leucinyl-leucinyl-norleucinal.

The cysteine protease inhibitor N-acetyl-leucinyl-leucinyl-norleucinal (LLnL) inhibited the growth of the Calu-1 lung carcinoma cells and induced a prolonged cell cycle arrest in the S phase. c-Jun N-terminal kinases (JNKs) participate in cellular responses to mitogenic stimuli, environmental stresses, and apoptotic signals but its role in cell cycle checkpoint control has not been elucidated. In this report, we examined the role of JNK in LLnL-induced S phase checkpoint by overexpression of a dominant-negative mutant of JNK1 (JNK1-APF) in Calu-1 cells. Expression of high levels of JNK1-APF blocked the growth-inhibitory effects of LLnL and abrogated S phase arrest induced by LLnL. These results support the role of JNK in the activation of cell cycle checkpoint induced by LLnL.

Anisomycin↗
PubMedDate not supplied

P21waf-1-Chk1 pathway monitors G1 phase microtubule integrity and is crucial for restriction point transition.

Microtubule-disruption (MTD) is often thought to arrest the mammalian cell cycle only during mitosis. However, MTD has also been demonstrated to arrest cells during interphase at a G(1)-phase point we call G(1)MTA. Microtubule integrity is now shown to be required for progression past G(1)MTA and the mammalian restriction-point. Neither p21(waf1) nor p27(kip1) are required for MTD-induced G(1)-arrest. Only p21(waf1) is crucial for normal G(1)MTA passage. The p21(waf1)-Chk1-cdc25C-cdc2-checkpoint-pathway is implicated in monitoring this passage. P21(waf1) deletion deregulates G(1)MTA transition and decreases MTD-G(1) arrest, possibly via Chk1 disregulation. Oncogene-induced overexpression of p21(waf1) produced opposite effects on the Chk1-cdc25C-cdc2 pathway and enhanced MTD-G(1) arrest. G(1)MTA thus represents a novel facet of mammalian G(1)/S checkpoint.

Animals↗