Down-Regulated CUEDC2 Increases GDNF Expression by Stabilizing CREB Through Reducing Its Ubiquitination in Glioma.

Abnormally high expression of glial cell line-derived neurotrophic factor (GDNF) derived from glioma cells has essential impacts on gliomagenesis and development, but the molecular basis underlying increased GDNF expression in glioma cells remain unclear. This work aimed to study the molecular mechanisms that may explain the accumulation of GDNF in glioma. Firstly, we observed that cAMP response element-binding protein (CREB), known as an important transcription factor for binding of GDNF promoter region, was highly expressed with an apparent accumulation into the nucleus of glioma cells, which may contribute to the transcription of GDNF. Secondly, CUE domain-containing protein 2 (CUEDC2), a ubiquitin-regulated protein, could increase the amount of binding between the E3 ligase tripartite motif-containing 21 (TRIM21) and CREB and affect the CREB level. Like our previous study, it showed that there was a significantly down-regulation of CUEDC2 in glioma. Finally, our data suggest that GDNF expression is indirectly regulated by transcription factor ubiquitination. Indeed, down-regulation of CUEDC2, decreased the ubiquitination and degradation of CREB, which was associated to high levels of GDNF. Furthermore, abundant CREB involved in the binding to the GDNF promoter region contributes to GDNF high expression in glioma cells. Collectively, it was verified the GDNF expression was affected by CREB ubiquitination regulated by CUEDC2 level.

CUE domain-containing protein 2 promotes the Warburg effect and tumorigenesis.

Cancer progression depends on cellular metabolic reprogramming as both direct and indirect consequence of oncogenic lesions; however, the underlying mechanisms are still poorly understood. Here, we report that CUEDC2 (CUE domain-containing protein 2) plays a vital role in facilitating aerobic glycolysis, or Warburg effect, in cancer cells. Mechanistically, we show that CUEDC2 upregulates the two key glycolytic proteins GLUT3 and LDHA via interacting with the glucocorticoid receptor (GR) or 14-3-3ζ, respectively. We further demonstrate that enhanced aerobic glycolysis is essential for the role of CUEDC2 to drive cancer progression. Moreover, using tissue microarray analysis, we show a correlation between the aberrant expression of CUEDC2, and GLUT3 and LDHA in clinical HCC samples, further demonstrating a link between CUEDC2 and the Warburg effect during cancer development. Taken together, our findings reveal a previously unappreciated function of CUEDC2 in cancer cell metabolism and tumorigenesis, illustrating how close oncogenic lesions are intertwined with metabolic alterations promoting cancer progression.

Sinomenine inhibits breast cancer cell invasion and migration by suppressing NF-κB activation mediated by IL-4/miR-324-5p/CUEDC2 axis.

Nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) is a vital transcription factor that regulates multiple important biological processes, including the epithelial-mesenchymal transition (EMT) and metastasis of breast cancer. Sinomenine is an isoquinoline well known for its remarkable curative effect on rheumatic and arthritic diseases and can induce apoptosis of several cancer cell types. Recently, sinomenine was reported as a tumor suppressor via inhibiting cell proliferation and inducing apoptosis. However, the role and mechanism of sinomenine in invasion and metastasis of breast cancer are largely unknown. Here, we report that sinomenine suppressed the invasion and migration of MDA-MB-231 and 4T1 breast cancer cells in a dose-dependent manner. We detected binding of NF-κB to the inhibitor of NF-κB (IκB) after the MDA-MB-231 cells were treated with 0.25, 0.5, and 1 mM sinomenine. Co-IP analysis revealed that sinomenine enhanced the binding of NF-κB and IκB in a dose-dependent manner, suggesting that sinomenine had an effect on inactivation of NF-κB. Western blotting and ELISA approaches indicated that the suppression effect was closely associated with the phosphorylation of IκB kinase (IKK) and its negative regulator CUEDC2. Sinomenine treatment decreased miR-324-5p expression, thus increased the level of its target gene CUEDC2, and then blocked the phosphorylation of IKK through altering the upstream axis. Finally, transfection of a miR-324-5p mimic inhibited the suppression of invasion and metastasis of MDA-MB-231 and 4T1 cell by sinomenine, providing evidence that sinomenine treatment suppressed breast cancer cell invasion and metastasis via regulation of the IL4/miR-324-5p/CUEDC2 axis. Our findings reveal a novel mechanism by which sinomenine suppresses cancer cell invasion and metastasis, i.e., blocking NF-κB activation.

CUEDC2 Protects Against Experimental Colitis and Suppresses Excessive Proliferation of Intestinal Mucosa.

BACKGROUND: CUEDC2, a CUE domain-containing protein, is highly expressed in many tumors, which also may be associated with inflammation. AIMS: In this study, we studied whether CUEDC2 plays a role in the progress of inflammatory bowel disease using CUEDC2 knockout (KO) mice and discussed the effects of CUEDC2 on cell proliferation in colonic mucosa. METHODS: CUEDC2 KO mice were administered with drinking dextran sodium sulfate (DSS) to establish colitis mice model. At different time points after DSS administration, body weight and stool consistency of mice were graded. Cytokines in colon tissue such as IL-6 were measured by RT-PCR. NF-κB and STAT3 signaling pathways in colon tissue were assessed by western blotting. Besides, cell proliferation of intestinal mucosa was analyzed by immunohistochemical staining. RESULTS: CUEDC2 alleviated the colonic inflammation, showing elevated body weight loss, worse diarrhea, and more severe colonic mucosal injury in CUEDC2 KO mice than WT mice. Moreover, pro-inflammatory cytokines such as IL-6, TNFα, COX2, and MIP2 were significantly elevated. In CUEDC2 KO mice, the NF-κB and STAT3 signaling pathways were increasingly activated in different stages of progression of the colonic inflammation, and the percentage of proliferating cells as indicated by Ki67, CyclinD1, and BrdU in the inflammatory tissues was significantly increased. CONCLUSIONS: Our findings demonstrate that CUEDC2 plays an important role in protection from colonic inflammation, primarily by inhibiting the NF-κB and STAT3 signaling pathways and preventing excessive proliferation of the inflammatory epithelial cell.

CUEDC2 interacts with heat shock protein 70 and negatively regulates its chaperone activity.

Recently studies have revealed that CUEDC2, a CUE domain-containing protein, plays critical roles in many biological processes, such as cell cycle, inflammation and tumorigenesis. In this study, to further explore the function of CUEDC2, we performed affinity purification combined with mass spectrometry analysis to identify its interaction proteins, which led to the identification of heat shock protein 70 (HSP70). We confirmed the interaction between CUEDC2 and HSP70 in vivo by co-immunoprecipitation assays. Mapping experiments revealed that CUE domain was required for their binding, while the PBD and CT domains of HSP70, mediated the interaction with CUEDC2. The intracellular Luciferase refolding assay indicated that CUEDC2 could inhibit the chaperone activity of HSP70. Together, our results identify HSP70 as a novel CUEDC2 interaction protein and suggest that CUEDC2 might play important roles in regulating HSP70 mediated stress responses.

Long Noncoding RNA AROD Inhibits Host Antiviral Innate Immunity via the miR-324-5p-CUEDC2 Axis.

Long noncoding RNAs (lncRNAs) are a class of noncoding RNAs that are involved in multiple biological processes. Here, we report a mechanism through which the lnc-AROD-miR-324-5p-CUEDC2 axis regulates the host innate immune response, using influenza A virus (IAV) as a model. We identified that host lnc-AROD without protein-coding capability is composed of 975 nucleotides. Moreover, lnc-AROD inhibited interferon-ß expression, as well as interferon-stimulated genes ISG15 and MxA. Furthermore, in vivo assays confirmed that lnc-AROD overexpression increased flu virus pathogenicity and mortality in mice. Mechanistically, lnc-AROD interacted with miR-324-5p, leading to decreased binding of miR-324-5p to CUEDC2. Collectively, our findings demonstrated that lnc-AROD is a critical regulator of the host antiviral response via the miR-324-5p-CUEDC2 axis, and lnc-AROD functions as competing endogenous RNA. Our results also provided evidence that lnc-AROD serves as an inhibitor of the antiviral immune response and may represent a potential drug target. IMPORTANCE lnc-AROD is a potential diagnostic and discriminative biomarker for different cancers. However, so far the mechanisms of lnc-AROD regulating virus replication are not well understood. In this study, we identified that lnc-AROD is downregulated during RNA virus infection. We demonstrated that lnc-AROD enhanced CUEDC2 expression, which in turn inhibited innate immunity and favored IAV replication. Our studies indicated that lnc-AROD functions as a competing endogenous RNA that binds miR-324-5p and reduces its inhibitory effect on CUEDC2. Taken together, our findings reveal that lnc-AROD plays an important role during the host antiviral immune response.

CUEDC2 Drives ß-Catenin Nuclear Translocation and Promotes Triple-Negative Breast Cancer Tumorigenesis.

Hyperactivation of Wnt signaling is crucial in tumor formation. Fully elucidating the molecular details of how the cancer-specific Wnt signaling pathway is activated or contributes to tumorigenesis will help in determining future treatment strategies. Here, we aimed to explore the contribution of CUEDC2, a novel CUE-domain-containing protein, to the activation of Wnt signaling and the tumorigenesis of triple-negative breast cancer (TNBC) and to determine the underlying mechanisms. TNBC patient samples and disease-free survival (DFS) data were used to determine the association between CUEDC2 and TNBC progression. The effects of CUEDC2 on TNBC were examined in TNBC cells in vitro and in subcutaneous xenograft tumors in vivo. Gene knockdown, immunoprecipitation plus liquid chromatography-tandem mass spectrometry, pull-down, co-immunoprecipitation, localized surface plasmon resonance, and nuclear translocation analysis were used to uncover the mechanisms of CUEDC2 in regulating Wnt signaling and TNBC development. CUEDC2 is sufficient to maintain the hyperactivation of Wnt signaling required for TNBC tumorigenesis. The contribution of CUEDC2 plays a major role in determining the outcome of oncogenic Wnt signaling both in vitro and in vivo. Mechanistically, the CUE domain in CUEDC2 directly bound to the ARM (7-9) domain in ß-catenin, promoted ß-catenin nuclear translocation and enhanced the expression of ß-catenin targeted genes. More importantly, an 11-amino-acid competitive peptide targeting the CUE domain in CUEDC2 blocked the interactions of CUEDC2 and ß-catenin and abrogated the malignant phenotype of TNBC cells in vitro and in vivo. We observed that TNBC patients who exhibited higher levels of CUEDC2 showed marked hyperactivation of the Wnt signaling pathway and poor clinical outcomes, highlighting the clinical relevance of our findings. CUEDC2 promotes TNBC tumor growth by enhancing Wnt signaling through directly binding to ß-catenin and accelerating its nuclear translocation. Targeting the interactions of CUEDC2 and ß-catenin may be a valuable strategy for combating TNBC.

CUEDC2 ablation enhances the efficacy of mesenchymal stem cells in ameliorating cerebral ischemia/reperfusion insult.

Mesenchymal stem cell (MSC) therapy has been reported to be a promising therapeutic option for cerebral ischemia/reperfusion (I/R) insult. However, the poor survival rate of engrafted MSCs under unfavorable cerebral I/R-induced microenvironment inhibits their efficiency during clinical application. CUE domain-containing 2(CUECD2) exhibits its protective role on cardiomyocytes by mediating the antioxidant capacity. Our study explored the functional role of CUEDC2 in cerebral I/R challenge and determined whether CUECD2-modified MSCs could improve the efficacy of treatment of the insulted neurons. We also evaluated the possible mechanisms involved in cerebral I/R condition. Cerebral I/R stimulation suppressed CUEDC2 levels in brain tissues and neurons. siRNA-CUEDC2 in neurons significantly inhibited cerebral I/R-induced apoptosis and oxidative stress levels invitro. Moreover, siRNA-CUEDC2 in the MSCs group remarkably enhanced the therapeutic efficacies in cerebral I/R-induced neuron injury and brain tissue impairment when compared to the non-genetic MSCs treatment group. At the molecular level, siRNA-CUEDC2 in MSCs markedly enhanced its antioxidant and anti-inflammatory effect in co-cultured neurons by upregulating glutathione peroxidase 1 (GPX1) expression levels while suppressing NF-kB activation. These findings provide a novel strategy for the utilization of MSCs to promote cerebral ischemic stroke outcomes.

Comparison of ADAM19 and CUEDC2 expression in EHCC and their clinicopathological significance.

Background: To evaluate the expression and clinicopathological significance of a disintegrin and metalloproteinases 19 (ADAM19) CUE domain containing protein 2 (CUEDC2) in extrahepatic cholangiocarcinoma (EHCC). Materials & methods: Immunostaining of ADAM19 and CUEDC2 was performed by EnVision immunohistochemistry in benign and malignant biliary tract tissues. Result: The expression of ADAM19 and CUEDC2 were significantly higher in EHCC (p < 0.05). ADAM19 expression was positive correlated with CUEDC2 expression in EHCC (p < 0.05). The overall survival time of those with positive expression of ADAM19 and CUEDC2 was lower (p < 0.001). Both positive expression of ADAM19 and CUEDC2 were independent prognostic factors in EHCC. Conclusion: ADAM19 and CUEDC2 have a positive correlation to the pathogenesis and dismal prognosis in EHCC.

CUEDC2 Contributes to Cisplatin-Based Chemotherapy Resistance in Ovarian Serious Carcinoma by Regulating p38 MAPK Signaling.

Chemoresistance remains an obstacle to the successful treatment of ovarian carcinoma. CUE domain-containing 2 (CUEDC2) plays critical roles in tumor genesis and overexpresses in many solid cancers, including ovarian serous carcinoma. In previous study, we found that overexpression of CUEDC2 might be a promising biomarker to evaluate the progression and to predict likely relapse of serous ovarian carcinoma. In present study, we found that higher expression of CUEDC2 was associated with higher resistance to cisplatin. The overall survival (OS) and disease-free survival time (DFS) of patients with cisplatin resistant was shorter than that of those with cisplatin sensitive, respectively, and the cisplatin sensitivity was independent predictor of a shorter OS time and DFS time. Knockdown of CUEDC2 by small interfering RNA enhanced the cisplatin sensitivity of serous ovarian carcinoma cells in SKOV3 cell lines. Furthermore, the phosphorylation of p38 MAPK were obviously increased after CUEDC2 knockdown, while p38 MAPK signaling contributes to cell growth and cell apoptosis. Our data suggest that CUEDC2 takes part in cisplatin-based chemotherapy resistance by regulating p38 MAPK signaling. And CUEDC2 is a promising biomarker and therapeutic target of cisplatin resistance in ovarian serous carcinoma.

Expression of CUEDC2 in colorectal cancer with different invasion and migration abilities.

OBJECTIVE: To investigate CUE domain containing 2 ( CUEDC2) expression in colorectal cancer with different invasion and migration abilities. METHODS: Fresh colon cancer tissues, obtained from patients with or without lymph node metastasis who were treated at the Department of General Surgery, Chinese People's Liberation Army General Hospital, and SW620 and HT29 colorectal cancer cell lines, were analysed for CUEDC2 expression. RESULTS: Real-time polymerase chain reaction showed significantly higher CUEDC2 mRNA levels in colon cancer tissue from patients with ( n = 8) versus without ( n = 8) lymph node metastasis, and in SW620 versus HT29 cells. Western blots revealed significantly higher CUEDC2 protein levels in colon cancer tissues from patients with versus without lymph node metastasis, and in SW620 versus HT29 cells. Colorectal cancer tissues from patients with lymph node metastasis showed more intense immunohistochemical staining and moderate staining of cell nuclei and cytoplasm versus less intense/weak staining in tissues from patients without lymph node metastasis. CONCLUSIONS: CUEDC2 is highly expressed in colorectal cancer tissues and colorectal cancer cell lines with high invasion and migration ability. CUEDC2 may be involved in promoting invasion and metastasis in colorectal cancer.

CUEDC2 modulates cardiomyocyte oxidative capacity by regulating GPX1 stability.

The irreversible loss of cardiomyocytes due to oxidative stress is the main cause of heart dysfunction following ischemia/reperfusion (I/R) injury and ageing-induced cardiomyopathy. Here, we report that CUEDC2, a CUE domain-containing protein, plays a critical role in oxidative stress-induced cardiac injury. Cuedc2(-/-) cardiomyocytes exhibited a greater resistance to oxidative stress-induced cell death. Loss of CUEDC2 enhanced the antioxidant capacity of cardiomyocytes, promoted reactive oxygen species (ROS) scavenging, and subsequently inhibited the redox-dependent activation of signaling pathways. Notably, CUEDC2 promoted E3 ubiquitin ligases tripartite motif-containing 33 (TRIM33)-mediated the antioxidant enzyme, glutathione peroxidase 1 (GPX1) ubiquitination, and proteasome-dependent degradation. Ablation of CUEDC2 upregulated the protein level of GPX1 in the heart significantly. Strikingly, in vivo, the infarct size of Cuedc2(-/-) heart was significantly decreased after I/R injury, and aged Cuedc2(-/-) mice preserved better heart function as the overall ROS levels in their hearts were significantly lower. Our results demonstrated a novel role of CUEDC2 in cardiomyocyte death regulation. Manipulating CUEDC2 level might be an attractive therapeutic strategy for promoting cardiomyocyte survival following oxidative stress-induced cardiac injury.

Overexpression of CUEDC2 Predicts Poor Prognosis in Ovarian Serous Carcinomas.

CUEDC2, a newly reported protein, plays critical roles in many biological processes, such as cell cycle, inflammation and tumorigenesis, however, its expression in ovarian serious carcinoma is still poorly understood. In this study, we performed an immunohistochemical study on 101 cases of ovarian serous carcinoma tissues to investigate whether CUEDC2 is a useful biomarker to evaluate the progression of ovarian serous carcinomas. The data showed that the overexpression of CUEDC2 was observed in 59.4% of ovarian serous carcinoma tissue samples and correlated with histopathological grade, patient age at diagnosis, FIGO stage and recurrence. To assess the clinical relevance of CUEDC2, we analyzed the survival follow-up information, the results showed that CUEDC2-positive expression was associated with a shorter disease-free survival time, the median disease-free survival time of CUEDC2-positive patients was 36.0 months compared with 53.9 months of CUEDC2-negative ones (Log-rank χ(2)=6.149, P=0.013). Collectively, our results suggested that CUEDC2 may be a promising biomarker to evaluate the progression of serous ovarian carcinoma and to predict likely relapse of ovarian serous carcinoma.

CUEDC2 down-regulation is associated with tumor growth and poor prognosis in lung adenocarcinoma.

CUE domain-containing 2 (CUEDC2) is a multi-functional protein, which regulates cell cycle, growth factor signaling and inflammation. We found that CUEDC2 was low in lung adenocarcinoma cell lines and lung adenocarcinoma tissues at both mRNA and protein levels. Low levels of CUEDC2 were correlated with a shorter survival time in patients with lung adenocarcinoma (p = 0.004). CUEDC2 expression was correlated with tumor T classification (P = 0.001) at clinical stage (P = 0.001) and tumor size (P = 0.033). Multivariate analysis suggested that CUEDC2 expression is an independent prognostic indicator for patients with lung adenocarcinoma. Ectopic expression of CUEDC2 decreased cell proliferation in vitro and inhibited tumor growth in nude mice in vivo. Knockdown of endogenous CUEDC2 by short hairpin RNAs (shRNAs) increased tumor growth. Inhibition of proliferation by CUEDC2 was associated with inactivation of the PI3K/Akt pathway, induction of p21 and down-regulation of cyclin D1. Our results suggest that decreased expression of CUEDC2 contributes to tumor growth in lung adenocarcinoma, leading to a poor clinical outcome.

CUEDC2 sensitizes chronic myeloid leukemic cells to imatinib treatment.

CUEDC2, a newly reported protein, has been found to be ubiquitously expressed in human tissues and repress NF-κB activity. To study the role of CUEDC2 in chronic myeloid leukemia (CML), we explored the function of CUEDC2 in CML cells through using the CML cell line K562 and its imatinib resistant cells K562/G01. K562 cells expressed a relatively higher level of CUEDC2 compared to K562/G01 cells. Knockdown of CUEDC2 in K562 cells resulted in decreased cell apoptosis after imatinib treatment; when CUEDC2 was overexpressed in K562/G01 cells, imatinib induced more cell apoptosis. By analyzing the activity of NF-κB, the results indicated a negative association between the expression of CUEDC2 and NF-κB signaling pathway in these CML cells. Our data suggested that the expression level of CUEDC2 has an inverse correlation with imatinib resistance and activity of NF-κB signaling pathway in CML cells, CUEDC2 could regulate imatinib sensitivity in CML cells at least partially through NF-κB signaling pathway.