ATM/ATR-independent inhibition of cyclin B accumulation in response to hydroxyurea in nontransformed cell lines is altered in tumour cell lines

Florensa, Roger; Bachs, Oriol; Agell, Neus

doi:10.1038/sj.onc.1207159

Cited by 14 publications

(25 citation statements)

References 56 publications

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“…7 Furthermore, a good correlation between the inhibition of DNA synthesis and the lack of cyclin B1 accumulation was observed in the tumor cell lines. 36 It is well established that the central role of p53 is partly associated with the regulation of cell apoptosis via Caspase-3, Caspase-9, Bcl-2 and Bax. [37][38][39] However, apoptosis with specific regard to Bcl-2 or caspase is also triggered by p53 independent manner, such as Par-4 or p21 WAF1/CIP1 pathway.…”

Section: Discussionmentioning

confidence: 99%

Loss of Cyclin B1 followed by downregulation of Cyclin A/Cdk2, apoptosis and antiproliferation in Hela cell line

Xie¹,

An²,

Kang³

et al. 2005

Intl Journal of Cancer

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Recent studies have shown that Cyclin B1 is overexpressed in various tumor types but present at low levels in normal tissues. To explore the possibility of employing Cyclin B1 as an anticancer target, we knocked down Cyclin B1 in an HeLa cell line using RNA interference (RNAi). Subsequently, we monitored cell cyclerelated molecules by Western blot together with immunofluorescence and determined cell cycle distribution by flow cytometry. XTT and soft agar colony growth experiments were performed to detect cell viability and proliferation. Furthermore, we analyzed cell apoptosis by measuring Bcl-2 and Bax protein level and DNAladder assay. After performing Cyclin B1 RNAi, Cyclin B1, Cyclin A and Cdk2 protein levels were found to be markedly downregulated, whereas Cdc2 was almost unaffected; S-phase fraction increased significantly; HeLa cell viability and cell colony forming ability were markedly diminished after the RNAi; Bcl-2 was noticeably attenuated but Bax was hardly changed; and HeLa cells displayed typical DNA ladder. The loss of Cyclin B1 resulted in the downregulation of Cyclin A and Cdk2, S-phase delay and eventually led to cell apoptosis and the decrease of cell viability and proliferation. Our studies suggest that Cyclin B1 may be a promising anticancer target. ' 2005 Wiley-Liss, Inc.Key words: Cyclin B1; RNA interference (RNAi); Cyclin A; Cyclindependent kinase 2 (Cdk2); apoptosis; HeLaThe prevalent methods employed to treat advanced cervical cancer are chemotherapy, radiation therapy or concomitant chemoradiotherapy, which have exerted immense effect on survival. However, chemotherapy may evoke cytotoxic and sensitization effects, some of which are life threatening. 1 Many attempts have been made to apply to advanced cervical cancer, such as developing therapeutic vaccination against human papillomavirus (HPV) antigens, 2 gene therapy using tissue-specific promoters, 3 adenovirus-mediated p53 delivery 4 and recombinant interleukin-12 adenovirus plus E7 protein.5 Nonetheless, few of them have been satisfying, and thus, it is urgent to explore novel therapy to maximize specificity and minimize off-target effect.Cyclin B1 is the regulatory subunit of Cdk1 serine/threonine kinase, and the accumulation of Cyclin B1 is a prerequisite for mitotic initiation in the late G2 phase in mammalian cells. [6][7][8] Because Cyclin B1 has a direct effect on mitosis, the overexpression of Cyclin B1 may lead to uncontrolled cell proliferation, neoplastic transformation and tumorigenesis.9-11 Hence, Cyclin B1 may be an oncogene in cancer.12 Recent studies have shown that Cyclin B1 is overexpressed in breast cancer, nonsmall cell lung cancer, colorectal cancer, prostate cancer and squamous cell carcinoma of the head and neck but not in normal cells, indicating that it can be a specific anticancer target. [12][13][14][15][16][17] Other studies indicate that tumors overexpressing Cyclin B1 may be resistant to radiotherapy and that Cyclin B1 may be an indicator for the risk of locoregional recurrence and metastasis in hea...

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Section: Discussionmentioning

confidence: 99%

Loss of Cyclin B1 followed by downregulation of Cyclin A/Cdk2, apoptosis and antiproliferation in Hela cell line

Xie¹,

An²,

Kang³

et al. 2005

Intl Journal of Cancer

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“…The presence of an ATR-and ATM-independent pathway activated by DNA synthesis inhibition has also been described (20,21). We have previously described that this pathway leads to a p53-independent down-regulation of cyclin B1 expression (20).…”

Section: Introductionmentioning

confidence: 99%

“…We have previously described that this pathway leads to a p53-independent down-regulation of cyclin B1 expression (20).…”

Section: Introductionmentioning

confidence: 99%

Different S/M Checkpoint Responses of Tumor and Non–Tumor Cell Lines to DNA Replication Inhibition

et al. 2007

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Cell cycle checkpoint abrogation, especially the inhibition of Chk1 in combination with DNA-damaging treatments, has been proposed as a promising way of sensitizing cancer cells. However, less is known about the possibility to selectively affect tumor cells when they are treated with agents that block DNA synthesis in combination with replication checkpoint inhibitors. Here, we present clear insights in the different responses of tumor and non-transformed cells to the inhibition of DNA replication with hydroxyurea in combination with checkpoint abrogation via inhibition of Ataxia telangiectasia-mutated-(ATM) and Rad3-related/ATM (ATR/ ATM) and Chk1 kinases. Interestingly, we find that nontransformed cell lines activate ATR/ATM-and Chk1-independent pathways in response to replication inhibition to prevent mitotic entry with unreplicated DNA. In contrast, tumor cell lines such as HCT116 and HeLa cells rely entirely on Chk1 activity for a proper response to replication inhibitors. Our results show that p38 is activated in response to hydroxyurea treatment and collaborates with Chk1 to prevent mitotic entry in non-transformed cell lines by maintaining cyclin B1/Cdk1 complexes inactive. Furthermore, DNA replication arrest down-regulates cyclin B1 promoter activity in non-transformed cells, but not in tumor cells in a Chk1-and p38-independent way. Thus, our data show that non-transformed cells present a more robust DNA replication checkpoint response compared with tumor cells that involves activation of the p38 pathway. We show that some of these responses to replication block can be lost in tumor cells, causing a defective checkpoint and providing a rationale for tumor-selective effects of combined therapies. [Cancer Res 2007;67(24):11648-56]

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“…Cell culture studies indicate that cyclin B1 accumulates during the S and G 2 phases of the cell cycle, and these changes are associated with an increase in cyclin B1 mRNA levels (14) and a decrease in protein degradation (15)(16)(17)(18). Treatment of the cells with a DNA-damaging agent, such as hydroxyurea or UV, decreases the level of cyclin B1 as well as mRNA for cyclin B1, and these treatments prevent entry into mitosis by both an ataxia-telangiectasia mutated (ATM)/ATM-and Rad3-related (ATR)-dependent and an ATM/ATR-independent mechanism (15).…”

Section: Introductionmentioning

confidence: 99%

“…Treatment of the cells with a DNA-damaging agent, such as hydroxyurea or UV, decreases the level of cyclin B1 as well as mRNA for cyclin B1, and these treatments prevent entry into mitosis by both an ataxia-telangiectasia mutated (ATM)/ATM-and Rad3-related (ATR)-dependent and an ATM/ATR-independent mechanism (15).…”

Section: Introductionmentioning

confidence: 99%

Effect of Caffeine on the ATR/Chk1 Pathway in the Epidermis of UVB-Irradiated Mice

et al. 2008

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Administration of caffeine was shown in earlier studies to enhance UVB-induced apoptosis and inhibit UVB-induced carcinogenesis in hairless SKH-1 mice. Here, we describe a potential mechanism for these in vivo effects. A single irradiation of mouse skin with UVB activated the ataxiatelangiectasia mutated-and Rad3-related (ATR) pathway, causing a severalfold increase in keratinocytes with phosphoChk1 (Ser 345 ) and a marked decrease in mitotic keratinocytes with cyclin B1 compared with baseline. When given in the drinking water for 1 to 2 weeks before UVB, caffeine (0.4 mg/mL) markedly inhibited the UVB-induced phosphorylation of Chk1 on Ser 345 and caused premature expression of cyclin B1 in the epidermis. Normal keratinocytes had delayed mitotic entry for >10 h following UVB. Caffeine administration reduced this mitotic delay to only 4 h and caused markedly increased apoptosis by 6 to 10 h after UVB. p53 knockout mice were used to determine the role of p53 in these processes. Irradiation with UVB markedly decreased the number of mitotic keratinocytes with cyclin B1 in p53 knockout mice, and topical caffeine immediately after UVB abrogated this response and increased UVB-induced apoptosis severalfold. These effects of caffeine in knockout mice were substantially greater than in wild-type mice. The ability of caffeine to promote the deletion of p53 À/À keratinocytes may be relevant to its inhibitory effect on UVB-induced skin cancer. Our studies indicate that administration of caffeine enhances the removal of DNA-damaged cells by inhibiting the ATRmediated phosphorylation of Chk1 and prematurely increasing the number of cyclin B1-containing cells that undergo lethal mitosis. [Cancer Res 2008;68(7):2523-9]

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ATM/ATR-independent inhibition of cyclin B accumulation in response to hydroxyurea in nontransformed cell lines is altered in tumour cell lines

Cited by 14 publications

References 56 publications

Loss of Cyclin B1 followed by downregulation of Cyclin A/Cdk2, apoptosis and antiproliferation in Hela cell line

Loss of Cyclin B1 followed by downregulation of Cyclin A/Cdk2, apoptosis and antiproliferation in Hela cell line

Different S/M Checkpoint Responses of Tumor and Non–Tumor Cell Lines to DNA Replication Inhibition

Effect of Caffeine on the ATR/Chk1 Pathway in the Epidermis of UVB-Irradiated Mice

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