The NUCKS1-SKP2-p21/p27 axis controls S phase entry.
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ABSTRACT:
SUBMITTER: Hume S
PROVIDER: S-EPMC8630071 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Poorly differentiated colorectal cancers (CRCs) are more aggressive and lack targeted therapies. We and others previously reported the predominant role of tumor-suppressor NDRG2 in promoting CRC differentiation, but the underlying mechanism is largely unknown. Herein, we demonstrate that NDRG2 induction of CRC cell differentiation is dependent on the repression of E3 ligase Skp2 activity. In patients and Ndrg2 knockout mice, NDRG2 and Skp2 are negatively correlated and associated with cell differentiation stage. Further, NDRG2 suppression of Skp2 contributes to the inductions and stabilizations of p21 and p27, which are Skp2 target proteins for degradation. The reduction of either p21 or p27 levels by shRNA can decrease NDRG2-induced AKP activity and resume cell growth inhibition, thus both p21 and p27 are required for NDRG2 effect on the promotion of cell differentiation in CRCs. The mechanistic study shows that NDRG2 suppresses ?-catenin nuclear translocation and decreases the occupancy of ?-catenin/TCF complex on Skp2 promoter, potentially through dephosphorylating GSK-3?. By subjecting a series of NDRG2 deletion mutants to Skp2 expression, the loss of NH2-terminal domain can completely abolish NDRG2-dependent differentiation induction. Supporting the biological significance of the reciprocal relationship between NDRG2 and Skp2, an NDRG2low/Skp2high gene expression signature correlates with poor CRC patient outcome and could be considered as a diagnostic marker of CRCs.
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Project description:Grade IV astrocytoma or glioblastoma multiforme (GBM) is one of the most aggressive and lethal tumors affecting humans. ADAR2-mediated A-to-I RNA editing, an essential post-transcriptional modification event in brain, is impaired in GBMs and astrocytoma cell lines. However, the role of ADAR2 editing in astrocytomas remains to be defined. Here, we show that ADAR2 editing rescue in astrocytomas prevents tumor growth in vivo and modulates an important cell cycle pathway involving the Skp2/p21/p27 proteins, often altered in glioblastoma. We demonstrate that ADAR2 deaminase activity is essential to inhibit tumor growth. Indeed, we identify the phosphatase CDC14B, which acts upstream of the Skp2/p21/p27 pathway, as a novel and critical ADAR2 target gene involved in glioblastoma growth. Specifically, ADAR2-mediated editing on CDC14B pre-mRNA increases its expression with a consequent reduction of the Skp2 target protein, as shown both in vitro and in vivo. We found that, compared to normal brain, both CDC14B editing and expression are progressively impaired in astrocytomas from grade I to IV, being very low in GBMs. These findings (1) demonstrate that post-transcriptional A-to-I RNA editing might be crucial for glioblastoma pathogenesis, (2) identify ADAR2-editing enzyme as a novel candidate tumor suppressor gene and (3) provide proof of principle that ADAR2 or its substrates may represent a suitable target(s) for possible novel, more effective and less toxic approaches to the treatment of GBMs.
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Project description:MYC is a pleiotropic transcription factor that controls a number of fundamental cellular processes required for the proliferation and survival of normal and malignant cells, including the cell cycle. MYC interacts with several central cell cycle regulators that control the balance between cell cycle progression and temporary or permanent cell cycle arrest (cellular senescence). Among these are the cyclin E/A/cyclin-dependent kinase 2 (CDK2) complexes, the CDK inhibitor p27KIP1 (p27) and the E3 ubiquitin ligase component S-phase kinase-associated protein 2 (SKP2), which control each other by forming a triangular network. MYC is engaged in bidirectional crosstalk with each of these players; while MYC regulates their expression and/or activity, these factors in turn modulate MYC through protein interactions and post-translational modifications including phosphorylation and ubiquitylation, impacting on MYC's transcriptional output on genes involved in cell cycle progression and senescence. Here we elaborate on these network interactions with MYC and their impact on transcription, cell cycle, replication and stress signaling, and on the role of other players interconnected to this network, such as CDK1, the retinoblastoma protein (pRB), protein phosphatase 2A (PP2A), the F-box proteins FBXW7 and FBXO28, the RAS oncoprotein and the ubiquitin/proteasome system. Finally, we describe how the MYC/CDK2/p27/SKP2 axis impacts on tumor development and discuss possible ways to interfere therapeutically with this system to improve cancer treatment.
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Project description:We have previously shown that the anti-malarial compound Quinacrine (QC) inhibits ovarian cancer (OC) growth by modulating autophagy. In the present study we extended these studies to identify the molecular pathways regulated by QC to promote apoptosis independent of p53 status in OC. QC exhibited strong anti-cancer properties in OC cell lines in contrast to other anti-malarial autophagy inhibiting drugs. QC treatment selectively upregulated cell cycle inhibitor p21, and downregulated F box protein Skp2 and p62/SQSTM1 expression independent of p53 status. Genetic downregulation of key autophagy protein ATG5 abolished QC-mediated effects on both cell cycle protein p21/Skp2 as well as autophagic cargo protein p62. Furthermore, genetic silencing of p62/SQSTM1 resulted in increased sensitivity to QC-mediated apoptosis, downregulated Skp2 mRNA and increased accumulation of p21 expression. Likewise, genetic knockdown of Skp2 resulted in the upregulation of p21 and p27 and increased sensitivity of OC cells to QC treatment. In contrast, transient overexpression of exogenous p62-HA plasmid rescued the QC-mediated Skp2 downregulation indicating the positive regulation of Skp2 by p62. Collectively, these data indicate that QC-mediated effects on cell cycle proteins p21/Skp2is autophagy-dependent and p53-independent in high grade serious OC cells.
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Project description:ObjectiveNatural extracts, including nobiletin, have been reported to enhance the efficacy and sensitivity of chemotherapeutic drugs. However, whether and how nobiletin affects tumor growth and progression in renal cell carcinoma (RCC) are still unclear.MethodsCell proliferation, cell cycle and apoptosis analyses, colony-formation assays, immunoblotting analysis, and qRT-PCR analysis were performed to investigate how nobiletin affected RCC cell proliferation in vitro. The nude mouse model was used to test the efficacy of nobiletin alone or in combination with palbociclib.ResultsNobiletin inhibited cell proliferation by inducing G1 cell cycle arrest and cell apoptosis in RCC cells. Mechanistically, nobiletin decreased SKP2 protein expression by reducing its transcriptional level. The downregulated SKP2 caused accumulation of its substrates, p27 and p21, which further inhibited the activity of the G1 phase-related protein, CDK2, leading to inhibition of cell proliferation and tumor formation. A higher SKP2 protein level indicated less sensitivity to the CDK4/6 inhibitor, palbociclib. A combination of nobiletin and palbociclib showed a synergistic tumor inhibition in vitro and in an in vivo model.ConclusionsNobiletin downregulated the SKP2-p21/p27-CDK2 axis to inhibit tumor progression and showed synergistic tumor inhibition effects with the CDK4/6 inhibitor, palbociclib, on RCC, which indicates a potential new therapeutic strategy.
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Project description:Checkpoint kinase 1 (CHK1; encoded by CHEK1) is an essential gene that monitors DNA replication fidelity and prevents mitotic entry in the presence of under-replicated DNA or exogenous DNA damage. Cancer cells deficient in p53 tumor suppressor function reportedly develop a strong dependency on CHK1 for proper cell cycle progression and maintenance of genome integrity, sparking interest in developing kinase inhibitors. Pharmacological inhibition of CHK1 triggers B-Cell CLL/Lymphoma 2 (BCL2)-regulated cell death in malignant cells largely independently of p53, and has been suggested to kill p53-deficient cancer cells even more effectively. Next to p53 status, our knowledge about factors predicting cancer cell responsiveness to CHK1 inhibitors is limited. Here, we conducted a genome-wide CRISPR/Cas9-based loss-of-function screen to identify genes defining sensitivity to chemical CHK1 inhibitors. Next to the proapoptotic BCL2 family member, BCL2 Binding Component 3 (BBC3; also known as PUMA), the F-box protein S-phase Kinase-Associated Protein 2 (SKP2) was validated to tune the cellular response to CHK1 inhibition. SKP2 is best known for degradation of the Cyclin-dependent Kinase Inhibitor 1B (CDKN1B; also known as p27), thereby promoting G1-S transition and cell cycle progression in response to mitogens. Loss of SKP2 resulted in the predicted increase in p27 protein levels, coinciding with reduced DNA damage upon CHK1-inhibitor treatment and reduced cell death in S-phase. Conversely, overexpression of SKP2, which consequently results in reduced p27 protein levels, enhanced cell death susceptibility to CHK1 inhibition. We propose that assessing SKP2 and p27 expression levels in human malignancies will help to predict the responsiveness to CHK1-inhibitor treatment.
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Project description:AimsVascular smooth muscle cells (VSMCs) normally exhibit a very low proliferative rate. Vessel injury triggers VSMC proliferation, in part, through focal adhesion kinase (FAK) activation, which increases transcription of cyclin D1, a key activator for cell cycle-dependent kinases (CDKs). At the same time, we also observe that FAK regulates the expression of the CDK inhibitors (CDKIs) p27 and p21. However, the mechanism of how FAK controls CDKIs in cell cycle progression is not fully understood.Methods and resultsWe found that pharmacological and genetic FAK inhibition increased p27 and p21 by reducing stability of S-phase kinase-associated protein 2 (Skp2), which targets theCDKIs for degradation. FAK N-terminal domain interacts with Skp2 and an APC/C E3 ligase activator fizzy-related 1 (Fzr1) in the nucleus, which promote ubiquitination and degradation of both Skp2 and Fzr1. Notably, overexpression of cyclin D1 alone failed to promote proliferation of genetic FAK kinase-dead (KD) VSMCs, suggesting that the FAK-Skp2-CDKI signalling axis is distinct from the FAK-cyclin D1 pathway. However, overexpression of both cyclin D1 and Skp2 enabled proliferation of FAK-KD VSMCs, implicating that FAK ought to control both activating and inhibitory switches for CDKs. In vivo, wire injury activated FAK in the cytosol, which increased Skp2 and decreased p27 and p21 levels.ConclusionBoth pharmacological FAK and genetic FAK inhibition reduced Skp2 expression in VSMCs upon injury, which significantly reduced intimal hyperplasia through elevated expression of p27 and p21. This study revealed that nuclear FAK-Skp2-CDKI signalling negatively regulates CDK activity in VSMC proliferation.
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Project description:Deregulated expression of the transcriptional coactivator with PDZ-binding motif (WWTR1/TAZ) is a common feature of basal-like breast cancer (BLBC). Yet, how oncogenic TAZ regulates cell-cycle progression and proliferation in breast cancer remains poorly understood, and whether TAZ is required for tumor maintenance has not been established. Here, using an integrative oncogenomic approach, TAZ-dependent cellular programs essential for tumor growth and progression were identified. Significantly, TAZ-driven tumor cells required sustained TAZ expression, given that its withdrawal impaired both genesis and maintenance of solid tumors. Moreover, temporal inhibition of TAZ diminished the metastatic burden in established macroscopic pulmonary metastases. Mechanistic investigation revealed that TAZ controls distinct gene profiles that determine cancer cell fate through cell-cycle networks, including a specific, causal role for S-phase kinase-associated protein 2 (SKP2) in mediating the neoplastic state. Together, this study elucidates the molecular events that underpin the role of TAZ in BLBC and link to SKP2, a convergent communication node for multiple cancer signaling pathways, as a key downstream effector molecule. IMPLICATIONS: Understanding the molecular role of TAZ and its link to SKP2, a signaling convergent point and key regulator in BLBC, represents an important step toward the identification of novel therapeutic targets for TAZ-dependent breast cancer.
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Project description:Hepatocellular carcinoma (HCC) is characterized by a poor prognosis and is one of the leading causes of cancer-related death worldwide. Simvastatin, an HMG-CoA reductase inhibitor, which decreases cholesterol synthesis by inhibiting mevalonate pathways and is widely used to treat cardiovascular diseases. Simvastatin exhibits anticancer effects against several malignancies. However, the molecular mechanisms underlying the anticancer effects of simvastatin on HCC are still not well understood. In this study, we demonstrated simvastatin-induced G0/G1 arrest by inducing p21 and p27 accumulation in HepG2 and Hep3B cells. Simvastatin also promoted AMP-activated protein kinase (AMPK) activation, which induced p21 upregulation by increasing its transcription. Consistent with this finding, we found genetic silencing of AMPK reduced p21 expression; however, AMPK silencing had no effect on p27 expression in HCC cells. Simvastatin decreased Skp2 expression at the transcriptional level, which resulted in p27 accumulation by preventing proteasomal degradation, an effect mediated by signal transducer and activator of transcription 3 (STAT3) inhibition. Constitutive STAT3 activation maintained high-level Skp2 expression and lower level p27 expression and significantly prevented G0/G1 arrest in simvastatin-treated HCC cells. Mevalonate decreased simvastatin-induced AMPK activation and rescued phospho-STAT3 and Skp2 expression in HCC cells, which resulted in the prevention of G0/G1 arrest through inhibition of p21 and p27 accumulation. Moreover, simvastatin significantly decreased tumor growth in HepG2 xenograft mice. Consistently, we found that simvastatin also increased p21 and p27 expression in tumor sections by reducing Skp2 expression and inducing AMPK activation and STAT3 suppression in the same tumor tissues. Taken together, these findings are demonstrative of the existence of a novel pathway in which simvastatin induces G0/G1 arrest by upregulating p21 and p27 by activating AMPK and inhibiting the STAT3-Skp2 axis, respectively. The results identify novel targets that explain the beneficial anticancer effects of simvastatin treatment on HCC in vitro and in vivo.
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Project description:Glucocorticoid therapy is an important treatment modality of hematological malignancies, especially T-cell acute lymphoblastic leukemia (T-ALL). Glucocorticoids are known to induce a cell cycle arrest and apoptosis in T-lymphoma cells. We could demonstrate that the cell cycle arrest induced by the synthetic glucocorticoid dexamethasone (Dex) clearly precedes apoptosis in human CEM T-ALL and murine S49.1 T-lymphoma cells. Cyclin D3 is strongly downregulated, whereas the CDK inhibitor p27 (Kip1) (p27) is strongly upregulated in response to dexamethasone in these cells. RNAi-mediated knockdown of p27 as well as overexpression of its negative regulator Skp2 revealed the critical function of p27 in the Dex-induced G 1 arrest of CEM cells. Our studies indicate that several mechanisms contribute to the increase of p27 protein in our T-lymphoma cell lines. We found a significant upregulation of p27 mRNA in S49.1 and CEM cells. In addition, Dex treatment activated the mouse p27 promotor in reporter gene experiments, indicating a transcriptional regulation. However, the relatively moderate induction of p27 mRNA levels by Dex did not explain the strong increase of p27 protein in CEM and S49.1 cells. We found clear evidence for a posttranslational mechanism responsible for the robust increase in p27 protein. Dex treatment of S49.1 and CEM cells increases the half-life of p27 protein, which indicates that decreased protein degradation is the primary mechanism of p27 induction by glucocorticoids. Interestingly, we found that Dex treatment decreased the protein and mRNA levels of the negative regulator of p27 protein and E3 ubiquitin ligase subunit Skp2. We conclude that the cell cycle inhibitor p27 and its negative regulator Skp2 are key players in the glucocorticoid-induced growth suppression of T-lymphoma cells and should be considered as potential drug targets to improve therapies of T-cell malignancies.
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