Cdh1 Regulates Cell Cycle through Modulating the Claspin/Chk1 and the Rb/E2F1 Pathways

APC/Cdh1 is a major cell cycle regulator and its function has been implicated in DNA damage repair; however, its exact role remains unclear. Using affinity purification coupled with mass spectrometry, we identified Claspin as a novel Cdh1-interacting protein and further demonstrated that Claspin is a novel Cdh1 ubiquitin substrate. As a result, inactivation of Cdh1 leads to activation of the Claspin/Chk1 pathway. Previously, we demonstrated that Rb interacts with Cdh1 to influence its ability to degrade Skp2. Here, we report that Cdh1 reciprocally regulates the Rb pathway through competing with E2F1 to bind the hypophosphorylated form of Rb. Although inactivation of Cdh1 in HeLa cells, with defective p53/Rb pathways, led to premature S phase entry, acute depletion of Cdh1 in primary human fibroblasts resulted in premature senescence. Acute loss of many other major tumor suppressors, including PTEN and VHL, also induces premature senescence in a p53- or Rb-dependent manner. Similarly, we showed that inactivation of the p53/Rb pathways by overexpression of SV40 LT-antigen partially reversed Cdh1 depletion–induced growth arrest. Therefore, loss of Cdh1 is only beneficial to cells with abnormal p53 and Rb pathways, which helps explain why Cdh1 loss is not frequently found in many tumors.

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CHFR regulates chemoresistance in triple-negative breast cancer through destabilizing ZEB1

Cell Death and Disease ◽

10.1038/s41419-021-04114-8 ◽

2021 ◽

Vol 12 (9) ◽

Author(s):

Hong Luo ◽

Zhicheng Zhou ◽

Shan Huang ◽

Mengru Ma ◽

Manyu Zhao ◽

...

Keyword(s):

Breast Cancer ◽

Cell Cycle ◽

Triple Negative Breast Cancer ◽

Fatty Acid Synthase ◽

Triple Negative ◽

Trichostatin A ◽

Affinity Purification ◽

Negative Regulator ◽

Dependent Manner ◽

Breast Cancers

AbstractFailures to treat triple-negative breast cancer (TNBC) are mainly due to chemoresistance or radioresistance. We and others previously discovered that zinc finger E-box-binding homeobox 1 (ZEB1) is a massive driver causing these resistance. However, how to dynamically modulate the intrinsic expression of ZEB1 during cell cycle progression is elusive. Here integrated affinity purification combined with mass spectrometry and TCGA analysis identify a cell cycle-related E3 ubiquitin ligase, checkpoint with forkhead and ring finger domains (CHFR), as a key negative regulator of ZEB1 in TNBC. Functional studies reveal that CHFR associates with and decreases ZEB1 expression in a ubiquitinating-dependent manner and that CHFR represses fatty acid synthase (FASN) expression through ZEB1, leading to significant cell death of TNBC under chemotherapy. Intriguingly, a small-molecule inhibitor of HDAC under clinical trial, Trichostatin A (TSA), increases the expression of CHFR independent of histone acetylation, thereby destabilizes ZEB1 and sensitizes the resistant TNBC cells to conventional chemotherapy. In patients with basal-like breast cancers, CHFR levels significantly correlates with survival. These findings suggest the therapeutic potential for targeting CHFR-ZEB1 signaling in resistant malignant breast cancers.

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Cell Cycle Association of the Retinoblastoma Protein Rb and the Histone Demethylase LSD1 with the Epstein-Barr Virus Latency Promoter Cp

Journal of Virology ◽

10.1128/jvi.01412-07 ◽

2008 ◽

Vol 82 (7) ◽

pp. 3428-3437 ◽

Cited By ~ 23

Author(s):

Charles M. Chau ◽

Zhong Deng ◽

Hyojueng Kang ◽

Paul M. Lieberman

Keyword(s):

Cell Cycle ◽

Epstein Barr Virus ◽

Affinity Purification ◽

Histone H3 ◽

Histone Demethylase ◽

Stable Complex ◽

Dependent Manner ◽

Barr Virus ◽

A Cell ◽

Epstein Barr

ABSTRACT The Epstein-Barr virus C promoter (Cp) regulates the major multicistronic transcript encoding the EBNA-LP, 1, 2, and 3 genes required for B-cell proliferation during latency. The growth-transforming potential of these viral genes suggests that they must be tightly regulated with the host cell cycle and differentiation process. To better understand Cp regulation, we used DNA affinity purification to identify cellular and viral proteins that bind to Cp in latently infected cells. Several previously unknown factors were identified, including the cell cycle regulatory proteins E2F1 and Rb. E2F1 bound to a specific site in Cp located in the core Cp region 3′ of the known EBNA2-responsive RBP-Jk (CSL, CBF1) binding site. The histone H3 K4 demethylase LSD1 (BCC110) was also identified by DNA affinity and was shown to form a stable complex with Rb. Coimmunoprecipitation assays demonstrated that E2F1, Rb, and LSD1 bind to Cp in a cell cycle-dependent manner. Rb and LSD1 binding to Cp increased after the S phase, corresponding to a decrease in histone H3 K4 methylation and Cp transcription. Coimmunoprecipitation and immunofluorescence assays reveal that LSD1 interacts with Rb. Surprisingly, LSD1 did not coimmunoprecipitate with E2F1, suggesting that it associates with Rb independently of E2F1. Depletion of LSD1 by small interfering RNAs inhibited Cp basal transcription levels, and overexpression of LSD1 altered the cell cycle profile in p53-positive (p53+), but not p53-negative (p53−), HCT cells. These findings indicate that Cp is a cell cycle-regulated promoter that is under the control of Rb and the histone demethylase LSD1 in multiple latency types.

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DNA Damage-Induced Cell Cycle Checkpoint Control Requires CtIP, a Phosphorylation-Dependent Binding Partner of BRCA1 C-Terminal Domains

Molecular and Cellular Biology ◽

10.1128/mcb.24.21.9478-9486.2004 ◽

2004 ◽

Vol 24 (21) ◽

pp. 9478-9486 ◽

Cited By ~ 246

Author(s):

Xiaochun Yu ◽

Junjie Chen

Keyword(s):

Cell Cycle ◽

Dna Damage ◽

Molecular Mechanisms ◽

Cell Cycle Checkpoint ◽

Dependent Manner ◽

Checkpoint Control ◽

Interacting Protein ◽

Binding Partner ◽

Cycle Checkpoint ◽

Multiple Cell

ABSTRACT The BRCA1 C-terminal (BRCT) domain has recently been implicated as a phospho-protein binding domain. We demonstrate here that a CTBP-interacting protein CtIP interacts with BRCA1 BRCT domains in a phosphorylation-dependent manner. The CtIP/BRCA1 complex only exists in G2 phase and is required for DNA damage-induced Chk1 phosphorylation and the G2/M transition checkpoint. However, the CtIP/BRCA1 complex is not required for the damage-induced G2 accumulation checkpoint, which is controlled by a separate BRCA1/BACH1 complex. Taken together, these data not only implicate CtIP as a critical player in cell cycle checkpoint control but also provide molecular mechanisms by which BRCA1 controls multiple cell cycle transitions after DNA damage.

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Deregulated expression of E2F-1 induces S-phase entry and leads to apoptosis.

Molecular and Cellular Biology ◽

10.1128/mcb.14.12.8166 ◽

1994 ◽

Vol 14 (12) ◽

pp. 8166-8173 ◽

Cited By ~ 298

Author(s):

B Shan ◽

W H Lee

Keyword(s):

Cell Cycle ◽

Cell Death ◽

Cell Cycle Progression ◽

Apoptotic Cell ◽

Expression System ◽

Critical Role ◽

S Phase ◽

Dependent Manner ◽

Cycle Progression ◽

Phase Entry

E2F-1, the first gene product identified among a family of E2F transcription factors, is thought to play a critical role in G1/S progression of the cell cycle. Transcriptional activities of E2F are modulated during the cell cycle, mainly by the formation of complexes between E2F and several key regulators of cell cycle such as the retinoblastoma protein and related proteins. To further understand the roles of E2F in the cell cycle progression, we have overexpressed exogenous E2F-1 by using a tetracycline-controlled expression system. We have found that the induced expression of E2F-1 in Rat-2 fibroblasts promotes S-phase entry and subsequently leads to apoptosis. The apoptosis occurs in an E2F-1 dose-dependent manner. Cells resistant to the induction of apoptosis have lost the ability to express exogenous E2F-1. Cells growing in low serum are more sensitive to the E2F-1-mediated cell death. Overexpression of E2F-1 mutants that impair DNA binding or transactivation does not alter cell cycle progression or induce apoptosis. These results define a novel pathway to apoptosis and demonstrate that premature S-phase entry is associated with apoptotic cell death.

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Deregulated expression of E2F-1 induces S-phase entry and leads to apoptosis

Molecular and Cellular Biology ◽

10.1128/mcb.14.12.8166-8173.1994 ◽

1994 ◽

Vol 14 (12) ◽

pp. 8166-8173 ◽

Cited By ~ 2

Author(s):

B Shan ◽

W H Lee

Keyword(s):

Cell Cycle ◽

Cell Death ◽

Cell Cycle Progression ◽

Apoptotic Cell ◽

Expression System ◽

Critical Role ◽

S Phase ◽

Dependent Manner ◽

Cycle Progression ◽

Phase Entry

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The Epstein-Barr Virus Protein BRLF1 Activates S Phase Entry through E2F1 Induction

Journal of Virology ◽

10.1128/jvi.73.8.6540-6550.1999 ◽

1999 ◽

Vol 73 (8) ◽

pp. 6540-6550 ◽

Cited By ~ 40

Author(s):

Jennifer J. Swenson ◽

Amy E. Mauser ◽

William K. Kaufmann ◽

Shannon C. Kenney

Keyword(s):

Cell Cycle ◽

Epstein Barr Virus ◽

Human Fibroblasts ◽

S Phase ◽

Lytic Replication ◽

Osteosarcoma Cell ◽

Barr Virus ◽

Infected Cells ◽

Epstein Barr ◽

Phase Entry

ABSTRACT The Epstein-Barr Virus (EBV) immediate-early protein BRLF1 is one of two transactivators which mediate the switch from latent to lytic replication in EBV-infected cells. DNA viruses often modulate the function of critical cell cycle proteins to maximize the efficiency of virus replication. Here we have examined the effect of BRLF1 on cell cycle progression. A replication-deficient adenovirus expressing BRLF1 (AdBRLF1) was used to infect normal human fibroblasts and various epithelial cell lines. BRLF1 expression induced S phase entry in contact-inhibited fibroblasts and in the human osteosarcoma cell line U-2 OS. AdBRLF1 infection produced a dramatic increase in the level of E2F1 but not E2F4. In contrast, the levels of Rb, p107, and p130 were decreased in AdBRLF1-infected cells. Electrophoretic mobility shift assays confirmed an increased level of free E2F1 in the AdBRLF1-infected human fibroblasts. Consistent with the previously described effect of E2F1, AdBRLF1-infected fibroblasts had increased levels of p53 and p21 and died by apoptosis. BRLF1-induced activation of E2F1 may be required for efficient EBV lytic replication, since at least one critical viral replication gene (the viral DNA polymerase) is activated by E2F (C. Liu, N. D. Sista, and J. S. Pagano, J. Virol. 70:2545–2555, 1996).

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Human Cytomegalovirus pp71 Stimulates Cell Cycle Progression by Inducing the Proteasome-Dependent Degradation of the Retinoblastoma Family of Tumor Suppressors

Molecular and Cellular Biology ◽

10.1128/mcb.23.6.1885-1895.2003 ◽

2003 ◽

Vol 23 (6) ◽

pp. 1885-1895 ◽

Cited By ~ 126

Author(s):

Robert F. Kalejta ◽

Jill T. Bechtel ◽

Thomas Shenk

Keyword(s):

Cell Cycle ◽

Human Cytomegalovirus ◽

Tumor Suppressors ◽

Family Members ◽

Simian Virus 40 ◽

Cellular Transformation ◽

Simian Virus ◽

T Antigen ◽

Cellular Growth ◽

Dependent Manner

ABSTRACT The oncoproteins of the DNA tumor viruses, adenovirus E1A, simian virus 40 T antigen, and papillomavirus E7, each interact with the retinoblastoma family of tumor suppressors, leading to cell cycle stimulation, apoptosis induction, and cellular transformation. These proteins utilize a conserved LXCXE motif, which is also found in cellular proteins, to target the retinoblastoma family. Here, we describe a herpesvirus protein that shares a subset of the properties of the DNA tumor virus oncoproteins but maintains important differences as well. The human cytomegalovirus pp71 protein employs an LXCXD motif to attack the retinoblastoma family members and induce DNA synthesis in quiescent cells. pp71 binds to and induces the degradation of the hypophosphorylated forms of the retinoblastoma protein and its family members p107 and p130 in a proteasome-dependent manner. However, pp71 does not induce apoptosis and fails to transform cells. Thus, the similarities and differences in comparison to E1A, T antigen, and E7 make pp71 an interesting new tool with which to further dissect the role of the retinoblastoma/E2F pathway in cellular growth control and carcinogenesis.

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Characterization of a novel separase-interacting protein and candidate new securin, Eip1p, in the fungal pathogen Candida albicans

Molecular Biology of the Cell ◽

10.1091/mbc.e18-11-0696 ◽

2019 ◽

Vol 30 (19) ◽

pp. 2469-2489 ◽

Cited By ~ 1

Author(s):

Samantha Sparapani ◽

Catherine Bachewich

Keyword(s):

Candida Albicans ◽

Chromosome Segregation ◽

Affinity Purification ◽

Sequence Divergence ◽

Dependent Manner ◽

Uncharacterized Protein ◽

Interacting Protein ◽

Functional Analyses ◽

Anti Fungal ◽

Mitotic Regulation

Proper chromosome segregation is crucial for maintaining genomic stability and dependent on separase, a conserved and essential cohesin protease. Securins are key regulators of separases, but remain elusive in many organisms due to sequence divergence. Here, we demonstrate that the separase homologue Esp1p in the ascomycete Candida albicans, an important pathogen of humans, is essential for chromosome segregation . However, C. albicans lacks a sequence homologue of securins found in model ascomycetes. We sought a functional homologue through identifying Esp1p interacting factors. Affinity purification of Esp1p and mass spectrometry revealed Esp1p-Interacting Protein1 (Eip1p)/Orf19.955p, an uncharacterized protein specific to Candida species. Functional analyses demonstrated that Eip1p is important for chromosome segregation but not essential, and modulated in an APCCdc20-dependent manner, similar to securins. Eip1p is strongly enriched in response to methyl methanesulfate (MMS) or hydroxyurea (HU) treatment, and its depletion partially suppresses an MMS or HU-induced metaphase block. Further, Eip1p depletion reduces Mcd1p/Scc1p, a cohesin subunit and separase target. Thus, Eip1p may function as a securin. However, other defects in Eip1p-depleted cells suggest additional roles. Overall, the results introduce a candidate new securin, provide an approach for identifying these divergent proteins, reveal a putative anti-fungal therapeutic target, and highlight variations in mitotic regulation in eukaryotes.

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Pisosterol Induces G2/M Cell Cycle Arrest and Apoptosis via the ATM/ATR Signaling Pathway in Human Glioma Cells

Anti-Cancer Agents in Medicinal Chemistry ◽

10.2174/1871520620666200203160117 ◽

2020 ◽

Vol 20 (6) ◽

pp. 734-750

Author(s):

Wallax A.S. Ferreira ◽

Rommel R. Burbano ◽

Claudia do Ó. Pessoa ◽

Maria L. Harada ◽

Bárbara do Nascimento Borges ◽

...

Keyword(s):

Cell Cycle ◽

Cell Cycle Arrest ◽

Signaling Pathway ◽

Glioma Cell ◽

Molecular Mechanisms ◽

Glioma Cells ◽

Dependent Manner ◽

M Cell ◽

Trypan Blue Exclusion ◽

Cycle Arrest

Background: Pisosterol, a triterpene derived from Pisolithus tinctorius, exhibits potential antitumor activity in various malignancies. However, the molecular mechanisms that mediate the pisosterol-specific effects on glioma cells remain unknown. Objective: This study aimed to evaluate the antitumoral effects of pisosterol on glioma cell lines. Methods: The 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide (MTT) and trypan blue exclusion assays were used to evaluate the effect of pisosterol on cell proliferation and viability in glioma cells. The effect of pisosterol on the distribution of the cells in the cell cycle was performed by flow cytometry. The expression and methylation pattern of the promoter region of MYC, ATM, BCL2, BMI1, CASP3, CDK1, CDKN1A, CDKN2A, CDKN2B, CHEK1, MDM2, p14ARF and TP53 was analyzed by RT-qPCR, western blotting and bisulfite sequencing PCR (BSP-PCR). Results: Here, it has been reported that pisosterol markedly induced G2/M arrest and apoptosis and decreased the cell viability and proliferation potential of glioma cells in a dose-dependent manner by increasing the expression of ATM, CASP3, CDK1, CDKN1A, CDKN2A, CDKN2B, CHEK1, p14ARF and TP53 and decreasing the expression of MYC, BCL2, BMI1 and MDM2. Pisosterol also triggered both caspase-independent and caspase-dependent apoptotic pathways by regulating the expression of Bcl-2 and activating caspase-3 and p53. Conclusions: It has been, for the first time, confirmed that the ATM/ATR signaling pathway is a critical mechanism for G2/M arrest in pisosterol-induced glioma cell cycle arrest and suggests that this compound might be a promising anticancer candidate for further investigation.

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A Cytotoxic Natural Product from Patrinia villosa Juss

Anti-Cancer Agents in Medicinal Chemistry ◽

10.2174/1871520619666190416101014 ◽

2019 ◽

Vol 19 (11) ◽

pp. 1399-1404 ◽

Cited By ~ 1

Author(s):

Yangcheng Liu ◽

Wei Liu ◽

Changlan Chen ◽

Zheng Xiang ◽

Hongwei Liu

Keyword(s):

Cell Cycle ◽

Cell Lines ◽

Silica Gel ◽

Column Chromatography ◽

Bioactive Compound ◽

Antitumor Agent ◽

2D Nmr ◽

Preparative Hplc ◽

Dependent Manner ◽

2D Nmr Spectra

Background and Purpose:: Patrinia villosa Juss is an important Chinese herbal medicine widely used for thousands of years, but few reports on the ingredients of the herb have been presented. In this study, we aim to isolate the bioactive compound from the plant. Material and Methods:: The air-dried leaves of P. villosa (15kg) were extracted three times with 70% EtOH under reflux. The condensed extract was suspended in H2O and partitioned with light petroleum, dichloromethane and n-BuOH. The dichloromethane portion was then subjected to normal-phase silica gel column chromatography, ODS silica gel column chromatography and semi-preparative HPLC to yield compound 1. Cytotoxicities of 1 were assayed on HepG2, A549 and A2780 cell lines. The mechanism of apoptosis and cell cycle on A549 was confirmed subsequently. Results: A new impecylone (Impecylone A) was isolated from the leaves of Patrinia villosa Juss, and its structures were established using 1D, 2D-NMR spectra and HR-ESI-MS. Impecylone A could selectivity inhibit HepG2 and A549 cell lines. The compound could induce apoptosis of A549 and arrest the cell cycle at G2/M phase in a dose-dependent manner. Conclusion: Impecylone A is a novel compound from Patrinia villosa Juss and could be a potential antitumor agent especially in the cell lines of A549.

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