Dataset Information

C/EBP? Is a Transcriptional Regulator of Wee1 at the G?/M Phase of the Cell Cycle.

ABSTRACT: The CCAAT/enhancer-binding protein ? (C/EBP?) is a transcription factor that regulates cellular proliferation, differentiation, apoptosis and tumorigenesis. Although the pro-oncogenic roles of C/EBP? have been implicated in various human cancers, how it contributes to tumorigenesis or tumor progression has not been determined. Immunohistochemistry with human non-small cell lung cancer (NSCLC) tissues revealed that higher levels of C/EBP? protein were expressed compared to normal lung tissues. Knockdown of C/EBP? by siRNA reduced the proliferative capacity of NSCLC cells by delaying the G?/M transition in the cell cycle. In C/EBP?-knockdown cells, a prolonged increase in phosphorylation of cyclin dependent kinase 1 at tyrosine 15 (Y15-pCDK1) was displayed with simultaneously increased Wee1 and decreased Cdc25B expression. Chromatin immunoprecipitation (ChIP) analysis showed that C/EBP? bound to distal promoter regions of WEE1 and repressed WEE1 transcription through its interaction with histone deacetylase 2. Treatment of C/EBP?-knockdown cells with a Wee1 inhibitor induced a decrease in Y15-pCDK1 and recovered cells from G?/M arrest. In the xenograft tumors, the depletion of C/EBP? significantly reduced tumor growth. Taken together, these results indicate that Wee1 is a novel transcription target of C/EBP? that is required for the G?/M phase of cell cycle progression, ultimately regulating proliferation of NSCLC cells.

SUBMITTER: Lee JH

PROVIDER: S-EPMC6407104 | biostudies-literature | 2019 Feb

REPOSITORIES: biostudies-literature

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C/EBPβ Is a Transcriptional Regulator of Wee1 at the G₂/M Phase of the Cell Cycle.

Lee Ji Hae JH Sung Jee Young JY Choi Eun Kyung EK Yoon Hyun-Kyoung HK Kang Bo Ram BR Hong Eun Kyung EK Park Byung-Kiu BK Kim Yong-Nyun YN Rho Seung Bae SB Yoon Kyungsil K

Cells 20190211 2

The CCAAT/enhancer-binding protein β (C/EBPβ) is a transcription factor that regulates cellular proliferation, differentiation, apoptosis and tumorigenesis. Although the pro-oncogenic roles of C/EBPβ have been implicated in various human cancers, how it contributes to tumorigenesis or tumor progression has not been determined. Immunohistochemistry with human non-small cell lung cancer (NSCLC) tissues revealed that higher levels of C/EBPβ protein were expressed compared to normal lung tissues. Kn ...[more]

PMID: 30754676

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Project description:BackgroundNeuroblastoma is responsible for 15% of all childhood cancer deaths. Despite advances in treatment and disease management, the overall 5-year survival rates remain poor in high-risk disease (25-40%). MiR-497 was previously identified by our laboratory as a member of a miRNA expression signature, predictive of neuroblastoma patient survival and has been reported as a tumor suppressor in a variety of other cancers. WEE1, a tyrosine kinase regulator of the cell cycle and predicted target of miR-497, has emerged as an oncogene in several cancer types and therefore represents an attractive potential target for novel therapy approaches in high-risk neuroblastoma. Our aim was to investigate the potential tumor suppressive role of miR-497 in high-risk neuroblastoma.MethodsExpression levels of miR-497 and WEE1 in tissues and cells were determined using RT-PCR. The effect of miR-497 and siWEE1 on cell viability was evaluated using MTS assays, apoptosis levels were determined using FACS analysis of Annexin V/PI stained cells, and target protein expression was determined using western blot. Luciferase reporter plasmids were constructed to confirm direct targeting. Results were reported as mean±S.E.M and differences were tested for significance using 2-tailed Students t-test.ResultsWe determined that miR-497 expression was significantly lower in high-risk MYCN amplified (MNA) tumors and that low miR-497 expression was associated with worse EFS and OS in our cohort. Over-expression of miR-497 reduced cell viability and increased apoptosis in MNA cells. We identified WEE1 as a novel target for miR-497 in neuroblastoma. Furthermore, our analysis showed that high WEE1 levels are significantly associated with poor EFS and OS in neuroblastoma and that siRNA knockdown of WEE1 in MNA cell lines results in significant levels of apoptosis, supporting an oncogenic role of WEE1 in neuroblastoma. Cisplatin (CDDP) treatment of both miR-497 over-expressing cells and WEE1 inhibited cells, resulted in a significant increase in apoptosis in MNA cells, describing a synergistic effect and therefore a potential therapeutic for high-risk neuroblastoma.ConclusionOur study's results are consistent with miR-497 being a candidate tumor suppressor in neuroblastoma, through the direct targeting of WEE1. These findings re-enforce the proposal of WEE1 as a therapeutic target in neuroblastoma.

Project description:Circadian clocks are endogenous time-keeping systems that temporally organize biological processes. Gating of cell cycle events by a circadian clock is a universal observation that is currently considered a mechanism serving to protect DNA from diurnal exposure to ultraviolet radiation or other mutagens. In this study, we put forward another possibility: that such gating helps to insulate the circadian clock from perturbations induced by transcriptional inhibition during the M phase of the cell cycle. We introduced a periodic pulse of transcriptional inhibition into a previously published mammalian circadian model and simulated the behavior of the modified model under both constant darkness and light-dark cycle conditions. The simulation results under constant darkness indicated that periodic transcriptional inhibition could entrain/lock the circadian clock just as a light-dark cycle does. At equilibrium states, a transcriptional inhibition pulse of certain periods was always locked close to certain circadian phases where inhibition on Per and Bmal1 mRNA synthesis was most balanced. In a light-dark cycle condition, inhibitions imposed at different parts of a circadian period induced different degrees of perturbation to the circadian clock. When imposed at the middle- or late-night phase, the transcriptional inhibition cycle induced the least perturbations to the circadian clock. The late-night time window of least perturbation overlapped with the experimentally observed time window, where mitosis is most frequent. This supports our hypothesis that the circadian clock gates the cell cycle M phase to certain circadian phases to minimize perturbations induced by the latter. This study reveals the hidden effects of the cell division cycle on the circadian clock and, together with the current picture of genome stability maintenance by circadian gating of cell cycle, provides a more comprehensive understanding of the phenomenon of circading gating of cell cycle.

Dataset Information

C/EBP? Is a Transcriptional Regulator of Wee1 at the G?/M Phase of the Cell Cycle.

Publications

C/EBPβ Is a Transcriptional Regulator of Wee1 at the G₂/M Phase of the Cell Cycle.

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