Research IndicatorsGraph generated 13 March 2017 using data from PubMed using criteria.
Mouse over the terms for more detail; many indicate links which you can click for dedicated pages about the topic. Tag cloud generated 13 March, 2017 using data from PubMed, MeSH and CancerIndex
Specific Cancers (4)
Data table showing topics related to specific cancers and associated disorders. Scope includes mutations and abnormal protein expression.
Note: list is not exhaustive. Number of papers are based on searches of PubMed (click on topic title for arbitrary criteria used).
OMIM, Johns Hopkin University
Referenced article focusing on the relationship between phenotype and genotype.
International Cancer Genome Consortium.
Summary of gene and mutations by cancer type from ICGC
Cancer Genome Anatomy Project, NCI
COSMIC, Sanger Institute
Somatic mutation information and related details
GEO Profiles, NCBI
Search the gene expression profiles from curated DataSets in the Gene Expression Omnibus (GEO) repository.
Latest Publications: RBX1 (cancer-related)
Wang W, Chen H, Liu Z, et al.Regulator of cullins-1 expression knockdown suppresses the malignant progression of muscle-invasive transitional cell carcinoma by regulating mTOR/DEPTOR pathway.
Br J Cancer. 2016; 114(3):305-13 [PubMed
] Free Access to Full Article Related Publications
BACKGROUND: Regulator of cullins-1 (ROC1) is a key subunit in the cullin-RING ligase (CRL) protein complex. Our previous study indicated that ROC1 was essential for bladder cancer cell survival and that ROC1 knockdown inhibited CRL activity, triggering G2 phase arrest and senescence. However, the role of ROC1 in the malignant progression of bladder cancer remained unknown.
METHODS: ROC1 expression in cancer cells was knocked down by siRNA silencing. The effects of ROC1 silencing were evaluated by in vitro assays for cell migration and by an in vivo mouse metastasis model. Epithelial-mesenchymal transition (EMT) induction was evaluated by immunofluorescence staining and western blotting of EMT-associated proteins. ROC1 expression in human tumours was further evaluated by immunohistochemical analysis.
RESULTS: ROC1 knockdown suppresses bladder cancer cell migration by inhibiting EMT. ROC1 knockdown inhibited EMT by inhibiting mammalian target of rapamycin (mTOR) activity via the accumulation of the mTOR-inhibitory protein DEPTOR, a CRL substrate. DEPTOR knockdown partially rescued ROC1 knockdown-inhibited EMT and the ROC1-induced inhibition of cancer cell migration. Furthermore, in vivo studies using a nude mouse metastasis model confirmed the in vitro data. Finally, tissue microarray analysis of clinical bladder cancer specimens indicated a positive correlation between ROC1 expression and EMT.
CONCLUSIONS: ROC1 has an important role in the malignant progression of bladder cancer via the mTOR/DEPTOR pathway. ROC1 may serve as a novel therapeutic target for the treatment of muscle-invasive transitional cell carcinoma.
Nrf2 (nuclear factor erytheroid-derived-2-like 2) transcriptional programmes are activated by a variety of cellular stress conditions to maintain cellular homoeostasis. Under non-stress conditions, Nrf2 is under tight regulation by the ubiquitin proteasome system (UPS). Detailed mechanistic investigations have shown the Kelch-like ECH-associated protein 1 (Keap1)-cullin3 (Cul3)-ring-box1 (Rbx1) E3-ligase to be the primary Nrf2 regulatory system. Recently, both beta-transducin repeat-containing E3 ubiquitin protein ligase (β-TrCP) and E3 ubiquitin-protein ligase synoviolin (Hrd1) have been identified as novel E3 ubiquitin ligases that negatively regulate Nrf2 through Keap1-independent mechanisms. In addition to UPS-mediated regulation of Nrf2, investigations have revealed a cross-talk between Nrf2 and the autophagic pathway resulting in activation of Nrf2 in a non-canonical manner. In addition to regulation at the protein level, Nrf2 was recently shown to be regulated at the transcriptional level by oncogenic K-rat sarcoma (Ras). A consequence of these differential regulatory mechanisms is the dual role of Nrf2 in cancer: the canonical, protective role and the non-canonical 'dark-side' of Nrf2. Based on the protective role of Nrf2, a vast effort has been dedicated towards identifying novel chemical inducers of Nrf2 for the purpose of chemoprevention. On the other hand, upon malignant transformation, some cancer cells have a constitutively high level of Nrf2 offering a growth advantage, as well as rendering cancer cells resistant to chemotherapeutics. This discovery has led to a new paradigm in cancer treatment; the initially counterintuitive use of Nrf2 inhibitors as adjuvants in chemotherapy. Herein, we will discuss the mechanisms of Nrf2 regulation and how this detailed molecular understanding can be leveraged to develop Nrf2 modulators to prevent diseases, mitigate disease progression or overcome chemoresistance.
Uddin S, Bhat AA, Krishnankutty R, et al.Involvement of F-BOX proteins in progression and development of human malignancies.
Semin Cancer Biol. 2016; 36:18-32 [PubMed
] Related Publications
The Ubiquitin Proteasome System (UPS) is a core regulator with various protein components (ubiquitin-activating E1 enzymes, ubiquitin-conjugating E2 enzymes, ubiquitin-protein E3 ligases, and the 26S proteasome) which work together in a coordinated fashion to ensure the appropriate and efficient proteolysis of target substrates. E3 ubiquitin ligases are essential components of the UPS machinery, working with E1 and E2 enzymes to bind substrates and assist the transport of ubiquitin molecules onto the target protein. As the UPS controls the degradation of several oncogenes and tumor suppressors, dysregulation of this pathway leads to several human malignancies. A major category of E3 Ub ligases, the SCF (Skp-Cullin-F-box) complex, is composed of four principal components: Skp1, Cul1/Cdc53, Roc1/Rbx1/Hrt1, and an F-box protein (FBP). FBPs are the substrate recognition components of SCF complexes and function as adaptors that bring substrates into physical proximity with the rest of the SCF. Besides acting as a component of SCF complexes, FBPs are involved in DNA replication, transcription, cell differentiation and cell death. This review will highlight the recent literature on three well characterized FBPs SKP2, Fbw7, and beta-TRCP. In particular, we will focus on the involvement of these deregulated FBPs in the progression and development of various human cancers. We will also highlight some novel substrates recently identified for these FBPs.
Aromatase inhibitors are effective drugs that reduce or eliminate hormone-sensitive breast cancer. However, despite their efficacy, resistance to these drugs can occur in some patients. The INrf2 (Keap1):Nrf2 complex serves as a sensor of drug/radiation-induced oxidative/electrophilic stress. INrf2 constitutively suppresses Nrf2 by functioning as an adapter protein for the Cul3/Rbx1-mediated ubiquitination/degradation of Nrf2. Upon stress, Nrf2 dissociates from INrf2, is stabilized, translocates to the nucleus, and coordinately induces a battery of cytoprotective gene expression. Current studies investigated the role of Nrf2 in aromatase inhibitor resistance. RT-PCR and immunoblot assays showed that aromatase inhibitor-resistant breast cancer LTLTCa and AnaR cells express lower INrf2 and higher Nrf2 protein levels, as compared with drug-sensitive MCF-7Ca and AC1 cells, respectively. The increase in Nrf2 was due to lower ubiquitination/degradation of Nrf2 in aromatase inhibitor-resistant cells. Higher Nrf2-mediated levels of biotransformation enzymes, drug transporters, and antiapoptotic proteins contributed to reduced efficacy of drugs and aversion to apoptosis that led to drug resistance. shRNA inhibition of Nrf2 in LTLTCa (LTLTCa-Nrf2KD) cells reduced resistance and sensitized cells to aromatase inhibitor exemestane. Interestingly, LTLTCa-Nrf2KD cells also showed reduced levels of aldehyde dehydrogenase, a marker of tumor-initiating cells and significantly decreased mammosphere formation, as compared with LTLTCa-Vector control cells. The results together suggest that persistent aromatase inhibitor treatment downregulated INrf2 leading to higher expression of Nrf2 and Nrf2-regulated cytoprotective proteins that resulted in increased aromatase inhibitor drug resistance. These findings provide a rationale for the development of Nrf2 inhibitors to overcome resistance and increase efficacy of aromatase inhibitors.
Díaz-Rodríguez E, Pandiella AModulation of cereblon levels by anti-myeloma agents.
Leuk Lymphoma. 2016; 57(1):167-76 [PubMed
] Related Publications
The use of thalidomide derivatives (IMIDs) has improved multiple myeloma prognosis, through an unknown mechanism of action. Recently one molecular target, the cereblon (CRBN) protein, has been identified. CRBN acts by binding to DDB1-CUL4-ROC1 forming a ubiquitin ligase multiprotein complex. We have generated antibodies to different regions of CRBN protein, and analyzed the biological consequences of augmenting or decreasing CRBN levels. CRBN was expressed in all the myeloma cell lines tested, independently of their sensitivity to IMIDs, and the CRBN-DDB1-CUL4 complex was efficiently formed. At the molecular level, long-term treatment with IMIDs induced a slight decrease in CRBN levels and a reduction in the CRBN-DDB1-CUL4 complex. Interestingly, treatment with other anti-myeloma drugs downregulated cellular contents of CRBN, and in a much faster fashion. These results suggest that CRBN is an important mediator of the cellular response to IMIDs, but also critical in the maintenance of cell viability and/or proliferation.
Chen X, Wang Y, Zang W, et al.miR-194 targets RBX1 gene to modulate proliferation and migration of gastric cancer cells.
Tumour Biol. 2015; 36(4):2393-401 [PubMed
] Related Publications
RING box protein1 (RBX1), an essential component of SCF E3 ubiquitin ligases, plays an important role in gastric cancer. In the study, miR-194 and RBX1 expression was evaluated in 76 pairs of gastric tumor and non-tumor tissue samples by qRT-PCR, and clinicopathological characteristics were analyzed. CCK8, transwell assay, wound healing assay, and flow cytometry assay were performed to evaluate the effect of miR-194 on gastric cancer (GC) cellular proliferation, invasion, migration, apoptosis, and cell cycle, respectively. Luciferase reporter assays and Western blotting were used to evaluate whether RBX1 is a direct target of miR-194. The Kaplan-Meier method and log-rank test were used to evaluate the correlation between miR-194 or RBX1 expression and patient survival. Then, we found that miR-194 was significantly downregulated and RBX1 upregulated in GC tissues; both of which showed significant association with tumor size, location, invasion, and tumor node metastasis. Cell proliferation, invasion, and migration were significantly restricted with miR-194 overexpression. miR-194 downregulated RBX1 protein expression, and luciferase assays showed that binding sites in the RBX1 3'UTR were required for miR-194-mediated repression of RBX1, indicating that RBX1 was a direct target of miR-194. Transfection of RBX1 without the 3'UTR restored the miR-194-inhibiting migration function. miR-194 overexpression or RBX1 lowexpression was associated with prolonged survival of GC patients. In conclusion, upregulation of miR-194 can inhibit proliferation, migration, and invasion of GC cells, possibly by targeting RBX1. Aberrant expression of miR-194 and RBX1 is correlated to GC patient survival time.
The activity of cullin-RING type ubiquitination E3 ligases is regulated by neddylation, a process analogous to ubiquitination that culminates in covalent attachment of the ubiquitin-like protein Nedd8 to cullins. As a component of the E3 for neddylation, SCCRO/DCUN1D1 plays a key regulatory role in neddylation and, consequently, cullin-RING ligase activity. The essential contribution of SCCRO to neddylation is to promote nuclear translocation of the cullin-ROC1 complex. The presence of a myristoyl sequence in SCCRO3, one of four SCCRO paralogues present in humans that localizes to the membrane, raises questions about its function in neddylation. We found that although SCCRO3 binds to CAND1, cullins, and ROC1, it does not efficiently bind to Ubc12, promote cullin neddylation, or conform to the reaction processivity paradigms, suggesting that SCCRO3 does not have E3 activity. Expression of SCCRO3 inhibits SCCRO-promoted neddylation by sequestering cullins to the membrane, thereby blocking its nuclear translocation. Moreover, SCCRO3 inhibits SCCRO transforming activity. The inhibitory effects of SCCRO3 on SCCRO-promoted neddylation and transformation require both an intact myristoyl sequence and PONY domain, confirming that membrane localization and binding to cullins are required for in vivo functions. Taken together, our findings suggest that SCCRO3 functions as a tumor suppressor by antagonizing the neddylation activity of SCCRO.
van der Wijst MG, Brown R, Rots MGNrf2, the master redox switch: the Achilles' heel of ovarian cancer?
Biochim Biophys Acta. 2014; 1846(2):494-509 [PubMed
] Related Publications
Ovarian cancer is the most lethal gynecological tumor type in the world due to late stage detection, and resistance to chemotherapy. Therefore, alternative additional therapies are required. The etiology of ovarian cancer remains largely unknown, but risk factors point toward an important role for oxidative stress. Both healthy and tumor cells can cope with oxidative stress by activating the transcription factor Nrf2 (also known as Nfe2l2), the master regulator of antioxidant and cytoprotective genes. Indeed, for most ovarian cancers, aberrant activation of Nrf2 is observed, which is often associated with a copy number loss within the Nrf2-inhibitory complex KEAP1-CUL3-RBX1. A key role for Nrf2 in ovarian carcinogenesis has been validated by siRNA studies. However, to exploit the Nrf2 pathway for therapeutic interventions, potential side-effects should be minimized. In this review, we explore ovarian cancer specific factors with links to aberrant activity of Nrf2, to be exploited in future combination strategies, synergistic with direct Nrf2 inhibitory drugs. Particularly, we propose to stratify patients based on common ovarian cancer mutations (KRAS, BRAF, ERBB2, BRCA1 and its link with estradiol, TP53) for future NRF2 targeting strategies.
The NFE2-related factor 2 (NRF2) pathway is critical to initiate responses to oxidative stress; however, constitutive activation occurs in different cancer types, including serous ovarian carcinomas (OVCA). The KEAP1/CUL3/RBX1 E3-ubiquitin ligase complex is a regulator of NRF2 levels. Hence, we investigated the DNA-level mechanisms affecting these genes in OVCA. DNA copy-number loss (CNL), promoter hypermethylation, mRNA expression, and sequence mutation for KEAP1, CUL3, and RBX1 were assessed in a cohort of 568 OVCA from The Cancer Genome Atlas. Almost 90% of cases exhibited loss-of-function alterations in any components of the NRF2 inhibitory complex. CNL is the most prominent mechanism of component disruption, with RBX1 being the most frequently disrupted component. These alterations were associated with reduced mRNA expression of complex components, and NRF2 target gene expression was positively enriched in 90% of samples harboring altered complex components. Disruption occurs through a unique DNA-level alteration pattern in OVCA. We conclude that a remarkably high frequency of DNA and mRNA alterations affects components of the KEAP1/CUL3/RBX1 complex, through a unique pattern of genetic mechanisms. Together, these results suggest a key role for the KEAP1/CUL3/RBX1 complex and NRF2 pathway deregulation in OVCA.
The aims of this work are to study the antitumor effect of the adeno-associated virus on the xenografted tumors of chick embryo chorioallantoic membrane and predict potential genes and biological pathways which are associated with renal cell carcinoma. The adeno-associated virus NT4-TAT-6 × His-VHLbeta was constructed and identified. Then, chick embryos with xenografted tumor were divided into three groups and respectively inoculated with rAAV/NT4-TAT-6 × His-VHLbeta (group A), empty virus (group B), and phosphate-buffered saline (group C, the control subject). Antitumor effect in each group was investigated by means of immunofluorescence observation. Genes interacted with von Hippel-Lindau were screened by Search Tool for the Retrieval of Interacting Genes/Proteins database, while pathway analysis were performed based on Kyoto Encyclopedia of Genes and Genomes. The growth of xenografted tumors inoculated with recombinant adeno-associated virus was slower than the control subjects. The tumor volumes of group A showed significant difference compared with group B and group C (P < 0.05). Growth of xenografted tumors which administered with the recombinant adeno-associated virus was inhibited. Among the protein-protein interaction network, TCEB2, HIF1A, TCEB1, CUL2, RBX1, and PHF17 were hub genes which might be involved in the development of renal cell carcinoma. The most significant signaling pathway was renal cell carcinoma. In this paper, we constructed and identified the recombinant adeno-associated virus NT4-TAT-6 × His-VHLbeta and studied the antitumor effect of the adeno-associated virus on xenografted tumors of chicken embryo chorioallantoic membrane. In addition, genes in the protein-protein interaction network which are associated with renal cell carcinoma were revealed and the biological pathway of renal cell carcinoma was identified. Our results provide a gene-therapeutic agent for the treatment of human renal cell carcinoma.
Zhang P, Gao K, Tang Y, et al.Destruction of DDIT3/CHOP protein by wild-type SPOP but not prostate cancer-associated mutants.
Hum Mutat. 2014; 35(9):1142-51 [PubMed
] Related Publications
Characterization of the exome and genome of prostate cancers by next-generation sequencing has identified numerous genetic alternations. SPOP (speckle-type POZ protein) was identified as one of the most frequently affected genes by somatic point mutations in prostate cancer, suggesting SPOP is potentially a key driver for prostate cancer development and progression. However, how SPOP mutations contribute to prostate cancer remains to be elucidated. SPOP acts as an adaptor protein of the CUL3-RBX1 E3 ubiquitin ligase complex and selectively recruits substrates for their ubiquitination and subsequent degradation. DDIT3 is an endoplasmic reticulum (ER) stress-responsive transcription factor playing an essential role in apoptotic execution pathways triggered by ER stress. Here, we identified DDIT3/CHOP as a bona fide substrate for the SPOP-CUL3-RBX1 E3 ubiquitin ligase complex. SPOP recognizes a Ser/Thr-rich degron in the transactivation domain of DDIT3 and triggers DDIT3 degradation via the ubiquitin-proteasome pathway. Strikingly, prostate cancer-associated mutants of SPOP are defective in promoting DDIT3 degradation. This study reveals novel molecular events underlying the regulation of DDIT3 protein homeostasis and provides insight in understanding the relationship between SPOP mutations and ER stress dysregulation in prostate cancer.
Although it has been suggested that kinesin family member 14 (KIF14) has oncogenic potential in various cancers, including hepatocellular carcinoma (HCC), the molecular mechanism of this potential remains unknown. We aimed to elucidate the role of KIF14 in hepatocarcinogenesis by knocking down KIF14 in HCC cells that overexpressed KIF14. After KIF14 knockdown, changes in tumor cell growth, cell cycle and cytokinesis were examined. We also examined cell cycle regulatory molecules and upstream Skp1/Cul1/F-box (SCF) complex molecules. Knockdown of KIF14 resulted in suppression of cell proliferation and failure of cytokinesis, whereas KIF14 overexpression increased cell proliferation. In KIF14-silenced cells, the levels of cyclins E1, D1 and B1 were profoundly decreased compared with control cells. Of the cyclin-dependent kinase inhibitors, the p27(Kip1) protein level specifically increased after KIF14 knockdown. The increase in p27(Kip1) was not due to elevation of its mRNA level, but was due to inhibition of the proteasome-dependent degradation pathway. To explore the pathway upstream of this event, we measured the levels of SCF complex molecules, including Skp1, Skp2, Cul1, Roc1 and Cks1. The levels of Skp2 and its cofactor Cks1 decreased in the KIF14 knockdown cells where p27(Kip1) accumulated. Overexpression of Skp2 in the KIF14 knockdown cells attenuated the failure of cytokinesis. On the basis of these results, we postulate that KIF14 knockdown downregulates the expression of Skp2 and Cks1, which target p27(Kip1) for degradation by the 26S proteasome, leading to accumulation of p27(Kip1). The downregulation of Skp2 and Cks1 also resulted in cytokinesis failure, which may inhibit tumor growth. To the best of our knowledge, this is the first report that has identified the molecular target and oncogenic effect of KIF14 in HCC.
Martinez VD, Vucic EA, Thu KL, et al.Disruption of KEAP1/CUL3/RBX1 E3-ubiquitin ligase complex components by multiple genetic mechanisms: Association with poor prognosis in head and neck cancer.
Head Neck. 2015; 37(5):727-34 [PubMed
] Related Publications
BACKGROUND: The NRF2 pathway has multiple pro-tumorigenic functions, and Nrf2 levels are increased in head and neck squamous cell carcinoma (HNSCC). The KEAP1/CUL3/RBX1 E3-ubiquitin ligase complex is a negative regulator of NRF2. In this study, we investigated mechanisms of disruption of individual complex components.
METHODS: Clinical and genomic profiles for 302 patients with HNSCC were obtained from The Cancer Genome Atlas. Combined pattern of epi/genetic alterations for individual components revealed frequent of complex disruption. Gene-set enrichment analysis was performed on expression data to identify affected pathways.
RESULTS: DNA loss is the main mechanism of alteration for all component genes, whereas hypermethylation largely affects only KEAP1. Combined analysis revealed that 64% of patients with HNSCC have disruption in this protein complex. Concordantly, NRF2-associated gene signature is enriched in HNSCC. Survival was significantly diminished among patients with one or more disrupted components.
CONCLUSION: The KEAP1/CUL3/RBX1 E3-ubiquitin ligase complex is frequently disrupted in HNSCC by multiple mechanisms. NRF2-based prognostics will benefit from integrated analysis of component genes.
Wu HC, Lin YC, Liu CH, et al.USP11 regulates PML stability to control Notch-induced malignancy in brain tumours.
Nat Commun. 2014; 5:3214 [PubMed
] Related Publications
The promyelocytic leukaemia (PML) protein controls multiple tumour suppressive functions and is downregulated in diverse types of human cancers through incompletely characterized post-translational mechanisms. Here we identify USP11 as a PML regulator by RNAi screening. USP11 deubiquitinates and stabilizes PML, thereby counteracting the functions of PML ubiquitin ligases RNF4 and the KLHL20-Cul3 (Cullin 3)-Roc1 complex. We find that USP11 is transcriptionally repressed through a Notch/Hey1-dependent mechanism, leading to PML destabilization. In human glioma, Hey1 upregulation correlates with USP11 and PML downregulation and with high-grade malignancy. The Notch/Hey1-induced downregulation of USP11 and PML not only confers multiple malignant characteristics of aggressive glioma, including proliferation, invasiveness and tumour growth in an orthotopic mouse model, but also potentiates self-renewal, tumour-forming capacity and therapeutic resistance of patient-derived glioma-initiating cells. Our study uncovers a PML degradation mechanism through Notch/Hey1-induced repression of the PML deubiquitinase USP11 and suggests an important role for this pathway in brain tumour pathogenesis.
Cullin-RING ligases (CRLs) are a family of E3 ubiquitin ligase complexes that rely on either RING-box 1 (RBX1) or sensitive to apoptosis gene (SAG), also known as RBX2, for activity. RBX1 and SAG are both overexpressed in human lung cancer; however, their contribution to patient survival and lung tumorigenesis is unknown. Here, we report that overexpression of SAG, but not RBX1, correlates with poor patient prognosis and more advanced disease. We found that SAG is overexpressed in murine KrasG12D-driven lung tumors and that Sag deletion suppressed lung tumorigenesis and extended murine life span. Using cultured lung cancer cells, we showed that SAG knockdown suppressed growth and survival, inactivated both NF-κB and mTOR pathways, and resulted in accumulation of tumor suppressor substrates, including p21, p27, NOXA, and BIM. Importantly, growth suppression by SAG knockdown was partially rescued by simultaneous knockdown of p21 or the mTOR inhibitor DEPTOR. Treatment with MLN4924, a small molecule inhibitor of CRL E3s, also inhibited the formation of KrasG12D-induced lung tumors through a similar mechanism involving inactivation of NF-κB and mTOR and accumulation of tumor suppressor substrates. Together, our results demonstrate that Sag is a Kras-cooperating oncogene that promotes lung tumorigenesis and suggest that targeting SAG-CRL E3 ligases may be an effective therapeutic approach for Kras-driven lung cancers.
Migita K, Takayama T, Matsumoto S, et al.Prognostic impact of RING box protein-1 (RBX1) expression in gastric cancer.
Gastric Cancer. 2014; 17(4):601-9 [PubMed
] Related Publications
BACKGROUND: RING box protein-1 (RBX1) is an essential component of the E3 ubiquitin ligase Skp1/Cullin/RBX1/F-box protein complex. Although an altered expression of RBX1 has been reported in several human cancers, the role of RBX1 in gastric cancer remains unknown.
METHODS: We investigated the RBX1 expression in primary gastric cancer tissues from 145 patients by immunohistochemistry, and explored its clinical relevance and prognostic value. Furthermore, the effect of RBX1 expression on cancer cell proliferation was analyzed in vitro using a siRNA silencing technique.
RESULTS: The RBX1 expression was abundant in gastric cancer tissues. There was a significant difference in the expression level of RBX1 in terms of the tumor depth (P = 0.008), presence of distant metastasis (P = 0.016) and venous invasion (P = 0.005). The postoperative overall (P < 0.001) and relapse-free survival (P < 0.001) rates were significantly poorer in patients with RBX1-high tumors than in patients with RBX1-low tumors. There was a significant correlation of the RBX1 status with postoperative hematogenous recurrence (P = 0.013). Importantly, the RBX1 status was identified as an independent prognostic factor for gastric cancer (P = 0.002). Furthermore, RBX1 gene silencing significantly inhibited the proliferation of gastric cancer cells in vitro.
CONCLUSIONS: The RBX1 expression has a significant prognostic value in gastric cancer. RBX1 might play an important role in regulating the proliferation of gastric cancer cells and promoting the development of postoperative recurrence. Our data provide a rationale for developing a novel therapy targeting RBX1 for gastric cancer.
Martinez VD, Vucic EA, Pikor LA, et al.Frequent concerted genetic mechanisms disrupt multiple components of the NRF2 inhibitor KEAP1/CUL3/RBX1 E3-ubiquitin ligase complex in thyroid cancer.
Mol Cancer. 2013; 12(1):124 [PubMed
] Free Access to Full Article Related Publications
BACKGROUND: Reactive oxygen species contribute to normal thyroid function. The NRF2 oxidative response pathway is frequently and constitutively activated in multiple tumor types, including papillary thyroid carcinoma (PTC). Genetic mechanisms underlying NRF2 pathway activation in PTC are not fully understood. Thus, we aimed to determine whether inactivating patterns of DNA-level alterations affect genes encoding for individual NRF2 inhibitor complex components (CUL3/KEAP1/RBX1) occur in PTC.
FINDINGS: Combined patterns of epi/genetic alterations for KEAP1/CUL3/RBX1 E3 ubiquitin-ligase complex components were simultaneously interrogated for a panel of 310 PTC cases and 40 adjacent non-malignant tissues. Data were obtained from The Cancer Genome Atlas project. Enrichment of NRF2 pathway activation was assessed by gene-set enrichment analysis using transcriptome data. Our analyses revealed that PTC sustain a strikingly high frequency (80.6%) of disruption to multiple component genes of the NRF2 inhibitor complex. Hypermethylation is the predominant inactivating mechanism primarily affecting KEAP1 (70.6%) and CUL3 (20%), while copy number loss mostly affects RBX1 (16.8%). Concordantly, NRF2-associated gene expression signatures are positively and significantly enriched in PTC.
CONCLUSIONS: The KEAP1/CUL3/RBX1 E3-ubiquitin ligase complex is almost ubiquitously affected by multiple DNA-level mechanisms and downstream NRF2 pathway targets are activated in PTC. Given the importance of this pathway to normal thyroid function as well as to cancer; targeted inhibition of NRF2 regulators may impact strategies for therapeutic intervention involving this pathway.
Achiwa Y, Hasegawa K, Udagawa YEffect of ursolic acid on MAPK in cyclin D1 signaling and RING-type E3 ligase (SCF E3s) in two endometrial cancer cell lines.
Nutr Cancer. 2013; 65(7):1026-33 [PubMed
] Related Publications
Cyclin D1 regulates G1 progression and is important in the development and proliferation of various human cancers. Cyclin D1 gene expression is activated by the Ras kinase cascade. Nuclear cyclin D1 levels are dependent on cytoplasmic degradation of cyclin D1 via ubiquitin-mediated proteolysis. We sought to determine whether the important MAPK signaling pathway, in the cyclin D1 cascade, including FBXW8, Cullin1, and the ubiquitination pathway mediated these effects. Ursolic acid (UA) treatment of SNG-2 cells, an endometrial cancer cell line, decreased cyclin D1, pERK1/2, FBXW8, and Cullin1 levels in a dose- and time-dependent manner. RING-type E3 ligase consists of CulIin1, Rbx, Skp1, and a member of the F-box protein family. In SNG-2, both dose- and time-dependent inhibition of Rbx 1 were observed following treatment with UA. Moreover, in HEC108 cells, another endometrial cancer cell line, UA treatment decreased cyclin D1, pERK1/2, and Cullin1 levels in a dose- and time-dependent manner and UA markedly inhibited FBXW8. Treatment of HEC108 cells moderately decreased Rbx1 in a dose- and-time-dependent fashion. In contrast, UA treatment increased ubiquitinated proteins in a dose- and time-dependent manner in both cell lines. RING-type E3 ligase accumulated in the cytoplasm following UA treatment of SNG-2cells. That in turn prevented cytoplasmic degradation of cyclin D1 via RING-type E3 (SCF E3s) ligase. In conclusion, our study found inhibition of the MAPK- cyclin D1 pathway and RING type E3 ligase (SCF E3s) in both endometrial cancer cell lines. Furthermore, CD36 was noted as a cell surface receptor for UA.
Cullin-RING ubiquitin ligases (CRLs) are the largest family of E3 ligases and require cullin neddylation for their activation. The NEDD8-activating enzyme inhibitor MLN4924 reportedly blocked cullin neddylation and inactivated CRLs, which resulted in apoptosis induction and tumor suppression. However, CRL roles in ovarian cancer cell survival and the ovarian tumor repressing effects of MLN4924 are unknown. We show here that CRL4 components are highly expressed in human epithelial ovarian cancer tissues. MLN4924-induced DNA damage, cell cycle arrest, and apoptosis in ovarian cancer cells in a time- and dose-dependent manner. In addition, MLN4924 sensitized ovarian cancer cells to other chemotherapeutic drug treatments. Depletion of CRL4 components Roc1/2, Cul4a, and DDB1 had inhibitory effects on ovarian cancer cells similar to MLN4924 treatment, which suggested that CRL4 inhibition contributed to the chemotherapeutic effect of MLN4924 in ovarian cancers. We also investigated for key CRL4 substrate adaptors required for ovarian cancer cells. Depleting Vprbp/Dcaf1 did not significantly affect ovarian cancer cell growth, even though it was expressed by ovarian cancer tissues. However, depleting Cdt2/Dcaf2 mimicked the pharmacological effects of MLN4924 and caused the accumulation of its substrate, CDT1, both in vitro and in vivo. MLN4924-induced DNA damage and apoptosis were partially rescued by Cdt1 depletion, suggesting that CRL4(CDT2) repression and CDT1 accumulation were key biochemical events contributing to the genotoxic effects of MLN4924 in ovarian cancer cells. Taken together, these results indicate that CRL4(CDT2) is a potential drug target in ovarian cancers and that MLN4924 may be an effective anticancer agent for targeted ovarian cancer therapy.
Regulator of Cullins-1 (ROC1) is a key subunit in the Cullin-RING ligase (CRL) protein complex. Overexpression of ROC1 protein is associated with tumor progression and poor prognosis of non-muscle invasive bladder transitional cell carcinoma (NMIBC). This study was designed to assess the effects of ROC1 knockdown in bladder cancer cells and to determine the potential mechanisms involved. A total of 112 bladder cancer tissue specimens were recruited for immunohistochemical analyses of ROC1 overexpression. Bladder cancer cell lines were used to knockdown ROC1 expression using ROC1 siRNA. Our data showed that ROC1 knockdown remarkably inhibited bladder cancer cell growth, arrested cells at the G2 phase of the cell cycle, and induced the p53-dependent cell senescence. Molecularly, G2 arrest was associated with upregulation of p21, p27, cyclin B1, and Cdc2 proteins. ROC1 knockdown induced-senescence functioned through p53/p21 pathway. Knockdown of p21 expression partially rescued ROC1 knockdown-induced growth inhibition in cancer cells. Furthermore, nude mouse xenograft analyses confirmed these in vitro data. In conclusion, data from the current study indicate that ROC1 plays an essential role in bladder cancer progression and could serve as a novel anticancer target for bladder transitional cell carcinoma (BTCC).
Lockwood WW, Chandel SK, Stewart GL, et al.The novel ubiquitin ligase complex, SCF(Fbxw4), interacts with the COP9 signalosome in an F-box dependent manner, is mutated, lost and under-expressed in human cancers.
PLoS One. 2013; 8(5):e63610 [PubMed
] Free Access to Full Article Related Publications
Identification of novel proteins that can potentially contribute to carcinogenesis is a requisite venture. Herein, we report the first biochemical characterization of the novel F-box and WD40 containing protein, FBXW4. We have identified interacting protein partners and demonstrated that FBXW4 is part of a ubiquitin ligase complex. Furthermore, the Fbxw4 locus is a common site of proviral insertion in a variety of retroviral insertional mutagenesis murine cancer models and Fbxw4 mRNA is highly expressed in the involuting murine mammary gland. To begin to characterize the biochemical function of Fbxw4, we used proteomic analysis to demonstrate that Fbxw4 interacts with Skp1 (SKP1), Cullin1 (CUL1), Ring-box1 (RBX1) and all components of the COP9 signalosome. All of these interactions are dependent on an intact F-box domain of Fbxw4. Furthermore, Fbxw4 is capable of interacting with ubiquitinated proteins within cells in an F-box dependent manner. Finally, we demonstrate that FBXW4 is mutated, lost and under-expressed in a variety of human cancer cell lines and clinical patient samples. Importantly, expression of FBXW4 correlates with survival of patients with non-small cell lung cancer. Taken together, we suggest that FBXW4 may be a novel tumor suppressor that regulates important cellular processes.
Wang W, Qiu J, Liu Z, et al.Overexpression of RING box protein-1 (RBX1) associated with poor prognosis of non-muscle-invasive bladder transitional cell carcinoma.
J Surg Oncol. 2013; 107(7):758-61 [PubMed
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BACKGROUND AND OBJECTIVE: RING box protein-1 (RBX1) is a key subunit of the ubiquitin E3 ligase Skp1/Cullin1/Rbx1/F-box protein complex. Altered expression RBX1 is shown to associate with tumorigenesis and tumor progression. This study detected RBX1 expression for association with clinical significance (such as clinicopathological data and survival of the patients) in non-muscle-invasive bladder transitional cell carcinoma (NMIBC).
METHODS: A total of 70 primary NMIBC tissue specimens and 24 normal tissue specimens were recruited and analyzed immunohistochemically for expression of RBX1 protein and associated with clinicopathological data and survival of the patients.
RESULTS: RBX1 was highly expressed in NMIBC, but was lowly expressed in the normal tissue. RBX1 expression was associated with high tumor grade and advanced clinical stage (P < 0.01 and P < 0.05, respectively). Moreover, patients with high RBX1 expression had shorter recurrence-free survival and progression-free survival rates (P < 0.001 and P < 0.01, respectively). Multivariate analysis demonstrated that RBX1 expression is an independent prognostic factor for tumor recurrence and progression of NMIBC (P < 0.05).
CONCLUSIONS: Overexpression of RBX1 protein contributes to tumor progression and poor prognosis of NMIBC.
Malloy MT, McIntosh DJ, Walters TS, et al.Trafficking of the transcription factor Nrf2 to promyelocytic leukemia-nuclear bodies: implications for degradation of NRF2 in the nucleus.
J Biol Chem. 2013; 288(20):14569-83 [PubMed
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Ubiquitylation of Nrf2 by the Keap1-Cullin3/RING box1 (Cul3-Rbx1) E3 ubiquitin ligase complex targets Nrf2 for proteasomal degradation in the cytoplasm and is an extensively studied mechanism for regulating the cellular level of Nrf2. Although mechanistic details are lacking, reports abound that Nrf2 can also be degraded in the nucleus. Here, we demonstrate that Nrf2 is a target for sumoylation by both SUMO-1 and SUMO-2. HepG2 cells treated with As2O3, which enhances attachment of SUMO-2/3 to target proteins, increased SUMO-2/3-modification (polysumoylation) of Nrf2. We show that Nrf2 traffics, in part, to promyelocytic leukemia-nuclear bodies (PML-NBs). Cell fractions harboring key components of PML-NBs did not contain biologically active Keap1 but contained modified Nrf2 as well as RING finger protein 4 (RNF4), a poly-SUMO-specific E3 ubiquitin ligase. Overexpression of wild-type RNF4, but not the catalytically inactive mutant, decreased the steady-state levels of Nrf2, measured in the PML-NB-enriched cell fraction. The proteasome inhibitor MG-132 interfered with this decrease, resulting in elevated levels of polysumoylated Nrf2 that was also ubiquitylated. Wild-type RNF4 accelerated the half-life (t½) of Nrf2, measured in PML-NB-enriched cell fractions. These results suggest that RNF4 mediates polyubiquitylation of polysumoylated Nrf2, leading to its subsequent degradation in PML-NBs. Overall, this work identifies Nrf2 as a target for sumoylation and provides a novel mechanism for its degradation in the nucleus, independent of Keap1.
Many biological processes such as cell proliferation, differentiation, and cell death depend precisely on the timely synthesis and degradation of key regulatory proteins. While protein synthesis can be regulated at multiple levels, protein degradation is mainly controlled by the ubiquitin-proteasome system (UPS), which consists of two distinct steps: (1) ubiquitylation of targeted protein by E1 ubiquitin-activating enzyme, E2 ubiquitin-conjugating enzyme and E3 ubiquitin ligase, and (2) subsequent degradation by the 26S proteasome. Among all E3 ubiquitin ligases, the SCF (SKP1-CUL1-F-box protein) E3 ligases are the largest family and are responsible for the turnover of many key regulatory proteins. Aberrant regulation of SCF E3 ligases is associated with various human diseases, such as cancers, including skin cancer. In this review, we provide a comprehensive overview of all currently published data to define a promoting role of SCF E3 ligases in the development of skin cancer. The future directions in this area of research are also discussed with an ultimate goal to develop small molecule inhibitors of SCF E3 ligases as a novel approach for the treatment of human skin cancer. Furthermore, altered components or substrates of SCF E3 ligases may also be developed as the biomarkers for early diagnosis or predicting prognosis.
Nrf2:INrf2 (Keap1) are cellular sensors of oxidative and electrophilic stress. Nrf2 is a nuclear factor that controls the expression and coordinated induction of a battery of genes that encode detoxifying enzymes, drug transporters, antiapoptotic proteins, and proteasomes. In the basal state, Nrf2 is constantly degraded in the cytoplasm by its inhibitor, INrf2. INrf2 functions as an adapter for Cul3/Rbx1 E3 ubiquitin ligase-mediated degradation of Nrf2. Chemicals, including antioxidants, tocopherols including α-tocopherol (vitamin E), and phytochemicals, and radiation antagonize the Nrf2:INrf2 interaction and lead to the stabilization and activation of Nrf2. The signaling events involve preinduction, induction, and postinduction responses that tightly control Nrf2 activation and repression back to the basal state. Oxidative/electrophilic signals activate unknown tyrosine kinases in a preinduction response that phosphorylates specific residues on Nrf2 negative regulators, INrf2, Fyn, and Bach1, leading to their nuclear export, ubiquitination, and degradation. This prepares nuclei for unhindered import of Nrf2. Oxidative/electrophilic modification of INrf2 cysteine 151 followed by PKC phosphorylation of Nrf2 serine 40 in the induction response results in the escape or release of Nrf2 from INrf2. Nrf2 is thus stabilized and translocates to the nucleus, resulting in a coordinated activation of gene expression. This is followed by a postinduction response that controls the "switching off" of Nrf2-activated gene expression. GSK3β, under the control of AKT and PI3K, phosphorylates Fyn, leading to Fyn nuclear localization. Fyn phosphorylates Nrf2 Y568, resulting in nuclear export and degradation of Nrf2. The activation and repression of Nrf2 provide protection against oxidative/electrophilic stress and associated diseases, including cancer. However, deregulation of INrf2 and Nrf2 due to mutations may lead to nuclear accumulation of Nrf2 that reduces apoptosis and promotes oncogenesis and drug resistance.
Malatesta M, Peschiaroli A, Memmi EM, et al.The Cul4A-DDB1 E3 ubiquitin ligase complex represses p73 transcriptional activity.
Oncogene. 2013; 32(39):4721-6 [PubMed
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The Cullin4A (cul4A)-dependent ligase (CDL4A) E3 has been implicated in a variety of biological processes, including cell cycle progression and DNA damage response. Remarkably, CDL4A exerts its function through both proteolytic and non-proteolytic events. Here, we show that the p53 family member p73 is able to interact with the CDL4A complex through its direct binding to the receptor subunit DNA-binding protein 1 (DDB1). As a result, the CDL4A complex is able to monoubiquitylate p73. Modification of p73 by CDL4A-mediated ubiquitylation does not affect p73 protein stability, but negatively regulates p73-dependent transcriptional activity. Indeed, genetic or RNA interference-mediated depletion of DDB1 induces the expression of several p73 target genes in a p53-independent manner. In addition, by exploiting a bioinformatic approach, we found that elevated expression of Cul4A in human breast carcinomas is associated with repression of p73 target genes. In conclusion, our findings add a novel insight into the regulation of p73 by the CDL4A complex, through the inhibition of its transcriptional function.
INTRODUCTION: Inhibitor of kappa light polypeptide gene enhancer in B-cells, kinase beta (IKBKB) (IKK-β/IKK-2), which activates NF-κB, is a substrate of the KEAP1-CUL3-RBX1 E3-ubiquitin ligase complex, implicating this complex in NF-κB pathway regulation. We investigated complex component gene disruption as a novel genetic mechanism of NF-κB activation in non-small cell lung cancer.
METHODS: A total of 644 tumor- and 90 cell-line genomes were analyzed for gene dosage status of the individual complex components and IKBKB. Gene expression of these genes and NF-κB target genes were analyzed in 48 tumors. IKBKB protein levels were assessed in tumors with and without complex or IKBKB genetic disruption. Complex component knockdown was performed to assess effects of the E3-ligase complex on IKBKB and NF-κB levels, and phenotypic importance of IKBKB expression was measured by pharmacological inhibition.
RESULTS: We observed strikingly frequent genetic disruption (42%) and aberrant expression (63%) of the E3-ligase complex and IKBKB in the samples examined. Although both adenocarcinomas and squamous cell carcinomas showed complex disruption, the patterns of gene disruption differed. IKBKB levels were elevated with complex disruption, knockdown of complex components increased activated forms of IKBKB and NF-κB proteins, and IKBKB inhibition detriments cell viability, highlighting the biological significance of complex disruption. NF-κB target genes were overexpressed in samples with complex disruption, further demonstrating the effect of complex disruption on NF-κB activity.
CONCLUSIONS: Gene dosage alteration is a prominent mechanism that disrupts each component of the KEAP1-CUL3-RBX1 complex and its NF-κB stimulating substrate, IKBKB. Herein, we show that, multiple component disruption of this complex represents a novel mechanism of NF-κB activation in non-small cell lung cancer.
Steroid receptor co-activator-3 (SRC-3/AIB1) is an oncogene that is amplified and overexpressed in many human cancers. However, the molecular mechanisms that regulate 'activated SRC-3 oncoprotein' turnover during tumorigenesis remain to be elucidated. Here, we report that speckle-type POZ protein (SPOP), a cullin 3 (CUL3)-based ubiquitin ligase, is responsible for SRC-3 ubiquitination and proteolysis. SPOP interacts directly with an SRC-3 phospho-degron in a phosphorylation-dependent manner. Casein kinase Iɛ phosphorylates the S102 in this degron and promotes SPOP-dependent turnover of SRC-3. Short hairpin RNA knockdown and overexpression experiments substantiated that the SPOP/CUL3/Rbx1 ubiquitin ligase complex promotes SRC-3 turnover. A systematic analysis of the SPOP genomic locus revealed that a high percentage of genomic loss or loss of heterozygosity occurs at this locus in breast cancers. Furthermore, we demonstrate that restoration of SPOP expression inhibited SRC-3-mediated oncogenic signaling and tumorigenesis, thus positioning SPOP as a tumor suppressor.
Nai G, Marques MRole of ROC1 protein in the control of cyclin D1 protein expression in skin melanomas.
Pathol Res Pract. 2011; 207(3):174-81 [PubMed
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A decrease in the level of the ROC1 protein, which is involved in cyclin D1 degradation, might explain an increase in cyclin D1 protein in the absence of gene overexpression. This study aimed to investigate the relationship between ROC1 and cyclin D1 expression in skin melanomas. A total of 62 cases of primary skin melanomas and 58 cases of compound melanocytic nevi were assessed. Immunohistochemistry was performed using cyclin D1 and ROC1 antibodies, and fluorescent in situ hybridization was used to assess the amplification of the CCND1 gene. ROC1 was expressed in >50% of cells in 87.9% of the melanocytic nevus cases and in 45.2% of the melanoma cases (p=0.0014). There was a significant negative correlation between ROC1 and cyclin D1 expression in all cases (p=0.0008985). In comparison with cyclin D1, ROC1 expression was increased in 86.2% of the melanocytic nevi and in 45.2% of the melanomas (p<0.001). Among the non-amplified melanomas, 50% expressed cyclin D1 in >50% of the cells and expressed ROC1 in <25%. ROC1 expression is negatively correlated with cyclin D1 expression, demonstrating its importance in the degradation of cyclin D1 in melanomas.
Bai J, Zhou Y, Chen G, et al.Overexpression of Cullin1 is associated with poor prognosis of patients with gastric cancer.
Hum Pathol. 2011; 42(3):375-83 [PubMed
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Cullin1 (Cul1) is a scaffold protein of the ubiquitin E3 ligase Skp1/Cullin1/Rbx1/F-box protein complex, which ubiquitinates a broad range of proteins involved in cell-cycle progression, signal transduction, and transcription. To investigate the role of Cullin1 in the development of gastric cancer, we examined the expression of Cullin1 in primary gastric cancer tissues and analyzed the correlation between Cullin1 expression and clinicopathologic variables and patients survival. We constructed a tissue microarray that includes 792 primary gastric cancer tissues and evaluated the Cullin1 expression by immunohistochemistry in the tumor biopsies. We also studied the role of Cullin1 in gastric cancer cell proliferation and adhesion by performing sulforhodamine B cell proliferation assay and cell attachment assay. The Cullin1 overexpression was significantly correlated with gastric cancer TNM stage (P = .011), depth of invasion (P = .035, comparing T1-T3 versus T4), and lymph node metastasis (P = .036). Furthermore, we showed a strong correlation between high Cullin1 expression and worse overall and 3-year survival rates in gastric cancer patients (P = .042 and P = .026, respectively). Cox regression analysis revealed that Cullin1 expression was an independent prognostic factor to predict 3-year patient outcome in gastric cancer (P = .028). Finally, we found that Cullin1 knockdown inhibits cell growth by up-regulating p27 expression and decreases cell adhesion ability by suppressing the expression of Src family kinases and focal adhesion kinase. Our data indicated that Cullin1 may be an important marker for human gastric cancer lymph node metastasis and prognosis.