E3 ubiquitin ligase RNF187 promotes growth of spermatogonia via lysine 48-linked polyubiquitination-mediated degradation of KRT36/KRT84.

Ubiquitination is the most common post-translational modification and is essential for various cellular regulatory processes. RNF187, which is known as RING domain AP1 coactivator-1, is a member of the RING finger family. RNF187 can promote the proliferation and migration of various tumor cells. However, whether it has a similar role in regulating spermatogonia is not clear. This study explored the role and molecular mechanism of RNF187 in a mouse spermatogonia cell line (GC-1). We found that RNF187 knockdown reduced the proliferation and migration of GC-1 cells and promoted their apoptosis. RNF187 overexpression significantly increased the proliferation and migration of GC-1 cells. In addition, we identified Keratin36/Keratin84 (KRT36/KRT84) as interactors with RNF187 by co-immunoprecipitation and mass spectrometry analyses. RNF187 promoted GC-1 cell growth by degrading KRT36/KRT84 via lysine 48-linked polyubiquitination. Subsequently, we found that KRT36 or KRT84 overexpression significantly attenuated proliferation and migration of RNF187-overexpressing GC-1 cells. In summary, our study explored the involvement of RNF187 in regulating the growth of spermatogonia via lysine 48-linked polyubiquitination-mediated degradation of KRT36/KRT84. This may provide a promising new strategy for treating infertility caused by abnormal spermatogonia development.

Circular RNA hsa_circ_0000915 promotes propranolol resistance of hemangioma stem cells in infantile haemangiomas.

BACKGROUND: Propranolol is a first-line clinical drug for infantile haemangiomas (IH) therapy. Nevertheless, resistance to propranolol is observed in some patients with IH. Circular RNAs (circRNAs) has been increasingly reported to act as a pivotal regulator in tumor progression. However, the underlying mechanism of circRNAs in IH remains unclear. METHODS: Quantitative real-time polymerase chain reaction was performed to detect Circ_0000915, miR-890 and RNF187 expression. Protein levels were determined using western blot. CCK-8 assay was used to measure cell proliferation. Caspase-3 activity assay and flow cytometry were conducted to determine cell apoptosis. Luciferase reporter assay was carried out to assess the interaction between miR-890 and Circ_0000915 or RNF187. Chromatin immunoprecipitation assay was performed to detect the interaction between STAT3 and Circ_0000915 promoter. Biotin pull-down assay was used to detect the direct interaction between miR-890 and Circ_0000915. In vivo experiments were performed to measure tumor formation. RESULTS: Here, we discovered depletion of Circ_0000915 increased propranolol sensitivity of haemangioma derived stem cells (HemSCs) both in vitro and in vivo, whereas forced expression of Circ_0000915 exhibited opposite effects. Mechanistically, Circ_0000915, transcriptionally induced by IL-6/STAT3 pathway, competed with RNF187 for the biding site in miR-890, led to upregulation of RNF187 by acting as a miR-890 "sponge". Furthermore, silence of miR-890 reversed increased propranolol sensitivity of HemSCs due to Circ_0000915 ablation. Moreover, increased Circ_0000915 and RNF187 levels were observed in IH tissues and positively associated with propranolol resistance, miR-890 exhibited an inverse expression pattern. CONCLUSION: We thereby uncover the activation of IL-6/STAT3/Circ_0000915/miR-890/RNF187 axis in propranolol resistance of IH, and provide therapeutic implications for patients of IH with propranolol resistance.

Overexpression of RNF187 induces cell EMT and apoptosis resistance in NSCLC.

Overexpression of RING finger protein 187 (RNF187) was recently revealed to be a driver of several cancers. However, the expression and function of RNF187 in non-small-cell lung cancer (NSCLC) are still unknown. Here, we uncovered that RNF187 expression was significantly higher in NSCLC samples than in matched normal lung samples at both the messenger RNA (3.55 ± 0.79 vs. 1.74 ± 0.63) and protein (2.85 ± 0.14 vs. 1.24 ± 0.02) levels. By downregulating or upregulating RNF187 expression in NSCLC cells, we showed that elevated RNF187 expression distinctly enhanced the migration, invasion, and colony formation of NSCLC cells. Moreover, we revealed that high level of RNF187 promoted NSCLC progression by inducing cell epithelial to mesenchymal transition (EMT) and apoptosis resistance mainly via activating the mitogen-activated protein kinase and PI3K signaling. Clinically, we demonstrated that RNF187 expression was positively associated with advanced TNM stage (p = 1.29 × 10 -6 ), lymph node metastasis ( p = 2.69 × 10 -9 ), and large tumor size ( p = 0.002). Importantly, NSCLC patients with elevated RNF187 expression related to the short overall survival rate( p = 1.29, E-7) and could serve as an independent prognostic factor in NSCLC patients. Thus, elevated RNF187 expression promotes NSCLC development by inducing cell EMT and apoptosis resistance, and RNF187 may be a novel prognostic indicator for NSCLC patients after curative resection.

High level of RNF187 contributes to the progression and drug resistance of osteosarcoma.

Objectives: Ring finger protein 187 (RNF187) was recently demonstrated to be up-regulation and function as a promoter in multiple cancers. However, the roles of RNF187 in osteosarcoma (OS) are unclear. Here, we tried to reveal the clinicopathological and biological roles of RNF187 in OS. Materials and Methods: We employed the quantitative real-time polymerase chain reaction (qRT-PCR) and immunohistochemistry (IHC) to determine the expression of RNF187 in OS tissues and cells. Migration and invasion capacities were analyzed by wound healing and transwell assays, and colony formation and CCK8 assays were performed to investigate proliferative ability. The functional effects of RNF187 on OS drugs resistance were further determined by CCK8 and western blot assays. Then, the relationship between RNF187 expression and clinical implications was analyzed by tissue microarrays (TMAs) including 51 OS cases. Moreover, the prognostic value was also determined by Kaplan-Meier analysis. Results: We reported that RNF187 mRNA was significantly increased in OS tissues compared to matched nontumorous tissues (3.83 ±0.79 vs. 1.70 ± 0.63), which was in line with the IHC assay in TMAs. By RNA interference and cDNA transfection, we showed high level of RNF187 increased the migration, invasion and proliferation of OS cells. Moreover, we demonstrated that elevated RNF187 expression induced OS cell drugs resistance, activated the ERK1/2 molecular and markedly enhanced the BCL-2 expression. Clinically, OS patients with high level of RNF187 was associated with Histologic differentiation (p=0.001), an advanced Enneking stage (p=0.001), response to chemotherapy (p=0.004), and metastasis (p= 0.001). Clinically, our data displayed that the RNF187 overexpression in OS samples associated with shorten overall survival (p=0.001) and high tumor recurrence (p=0.001) in postoperative OS patients. Conclusions: Our results indicate that Elevated RNF187 expression is a new adverse outcomes marker for OS patients and may be used as a new therapeutic target of OS.

RING finger protein 187 as a novel potential biomarker for predicting the prognosis of ovarian carcinoma in 2 cancer centers.

RING finger protein 187 (RNF187) has been used to predict prognosis of several human carcinomas. However, the clinicopathologic and prognostic implication of RNF187 expression in ovarian carcinomas remains not to be evaluated. The aim of this study was to explore the clinicopathologic and the prognostic significance of RNF187 in patients with ovarian carcinomas. Expression levels of RNF187 protein were investigated by immunohistochemical staining based on tissue-microarray composed of 147 patients with ovarian carcinomas. Receiver operating characteristic curve analysis was used to select the ideal cut-off value of RNF187 expression in ovarian carcinoma, and then analyze the correlation between the status of RNF187 expression and various clinicopathologic variables by chi-square test. Univariate analysis was employed to investigate the association between clinicopathologic variables and prognosis of patients by Kaplan-Meier method. Multivariate analysis was performed to identify the independent prognostic factors by the Cox regression model. Our results demonstrated that high expression of RNF187 was significantly associated with late FIGO stage, high histologic grade and pN1 stage in ovarian carcinoma (P < 0.05). Univariate analysis uncovered patients with the high expression of RNF187 have the worse overall survival and disease-free survival (P < 0.05). More surprisingly, multivariate analysis determined that the RNF187 expression was served as an independent prognostic factor in ovarian carcinoma. The high expression of RNF187 might influence a more aggressive biological behavior in ovarian carcinoma. Therefore, RNF187 expression could be useful to act as a new independent prognostic biomarker for patients with ovarian carcinoma.

An essential role of RNF187 in Notch1 mediated metastasis of hepatocellular carcinoma.

BACKGROUND: Aberrant activation of Notch signaling has been causally linked to the metastasis of hepatocellular carcinoma (HCC), however the underlying molecular mechanisms are still poorly understood. RING finger protein 187 (RNF187) was recently revealed to be a driver of several cancers, but its expression pattern and biological function in HCC are unknown. METHODS: The expression levels of Notch1 and RNF187 were assessed in two independent cohorts of HCC tissues, and modulation of Notch1 in HCC cells was performed to explore the regulatory role of Notch1 in HCC metastasis. RNA-sequencing (RNA-seq), bioinformatics analysis, luciferase reporter analysis, and chromatin immunoprecipitation assay (ChIP) were used to clarify the relationship between Notch1 signaling and its potential target Ring finger protein 187 (RNF187). Gain- and loss-of-function studies were used to dissect the role of Notch1-RNF187 signaling in promoting HCC metastasis. The impact of Notch1-RNF187 activity in determining clinical prognosis for HCC patients was evaluated by multivariate Cox regression. RESULTS: By RNA-seq, luciferase reporter analysis, and ChIP assay, RNF187 was confirmed to be a direct transcriptional target of Notch1, as Notch1 could activate RNF187 promoter whereas the pro-migratory and pro-invasive effects of Notch1 were significantly attenuated by RNF187 knockdown. Meanwhile, RNF187 silencing could attenuate the Notch1-dependent epithelial-mesenchymal transition (EMT). Moreover, overexpression of RNF187 counteracted the inhibitory effect of Notch1 knockdown on cancer progression. Importantly, HCC patients with high level of hepatic Notch1 expression had shorter disease-free survival (DFS) than those with low level of hepatic Notch1 expression. Furthermore, patients with high level of Notch1 and RNF187 co-expression showed the shortest DFS. The expression level of Notch1 and RNF187 was an independent prognostic factor for HCC. CONCLUSIONS: For the first time we identified that RNF187 is an essential factor for Notch1 to promote invasion and metastasis of HCC. Of highly clinical relevance, we found that activation of Notch1-RNF187 correlates with a worse prognosis of HCC patients. These findings provide a solid foundation for developing novel strategies to tackle HCC metastasis.

Up-regulation of RNF187 induces hepatocellular carcinoma cell epithelial to mesenchymal transitions.

Ring finger protein 187 (RNF187) has been identified to be a co-activator linking c Jun to Ras signaling. However, the expression and function of RNF187 in hepatocellular carcinomas (HCC) remains unclear. Here, we tried to determine the expression and roles of RNF187 in hepatocellular carcinomas (HCC).The expression of RNF187 was determined in HCC tissues and cell lines, and we found that RNF187 expressed highly in HCC tissues compared with the corresponding adjacent liver tissues both in mRNA and protein level, which was consistent with the result of immunohistochemistry on HCC tissue microarrays. In HCC cell lines, the level of RNF187 was positively associated with the HCC cells metastatic potential. By the RNF187 interference and cDNA transfection, we showed that the high level of RNF187 induced the HCC cells invasion and metastasis both in vitro and in vivo, as well as the high ability of colony formation.Mechanistically, we detected the high level of RNF187 promoted cell scatter by inducing epithelial-mesenchymal transition (EMT). Clinically, the high level of RNA187 was significantly correlated with a malignant phenotype, including larger tumor size, multiple tumors, and microvascular invasion. Importantly, high level of RNF187 correlated with HCC patients' shorter OS and lower disease free survival rates than those with low level of RNF187. Our results revealed that elevated expression of RNF187 induced hepatocellular carcinoma cell epithelial to mesenchymal transitions, and represented a novel marker for predicting the poor prognosis of HCC.