Yibin Ren

p28GANK overexpression accelerates hepatocellular carcinoma invasiveness and metastasis via phosphoinositol 3-kinase/AKT/hypoxia-inducible factor-1α pathways†

The overall survival of patients with hepatocellular carcinoma (HCC) remains poor, and the molecular mechanisms underlying HCC progression and aggressiveness are unclear. Here, we report that increased expression of p28GANK (Gankyrin, PSMD10, or p28) in human HCC predicts poor survival and disease recurrence after surgery. Patients with HCC who have large tumors, with vascular invasion and intrahepatic or distant metastasis, expressed high levels of p28GANK. Invasive tumors overexpressing p28GANK were featured by active epithelial‐mesenchymal transition (EMT), and exhibited increased angiogenesis associated with vascular endothelial growth factor overexpression, whereas silencing p28GANK expression attenuated EMT and motility/invasion of tumor cells. The p28GANK activates phosphoinositide 3‐kinase (PI3K)–V‐akt Murine Thymoma Viral Oncogene Homolog (AKT)–hypoxia‐inducible factor 1α (HIF‐1α) signaling to promote TWIST1, vascular endothelial growth factor, and metalloproteinase 2 expression. Suppression of the PI3K–AKT–HIF‐1α pathway interfered with p28GANK‐mediated EMT and invasion. Consistently, we detected a significant correlation between p28GANK expression and p‐AKT levels in a cohort of HCC biopsies, and the combination of these two parameters is a more powerful predictor of poor prognosis. Conclusion: These results present novel mechanistic insight into a critical role of p28GANK in HCC progression and metastasis. (HEPATOLOGY 2011)

The oncoprotein p28GANK establishes a positive feedback loop in β-catenin signaling.

p28GANK (also known as PSMD10 or gankyrin) is a novel oncoprotein that is highly expressed in hepatocellular carcinoma (HCC). Through its interaction with various proteins, p28GANK mediates the degradation of the tumor suppressor proteins Rb and p53. Although p53 was reported to downregulate β-catenin, whether p28GANK is involved in the regulation of β-catenin remains uncertain. Here we report that both growth factors and Ras upregulate p28GANK expression through the activation of the phosphoinositide 3-kinase-AKT pathway. Upregulation of p28GANK expression subsequently enhanced the transcription activity of β-catenin. This effect was observed in p53-deficient cells, suggesting a p53-independent mechanism for the p28GANK-mediated activation of β-catenin. p28GANK overexpression also reduced E-cadherin protein levels, leading to increased release of free β-catenin into the cytoplasm from the cadherin-bound pool. Interestingly, exogenous expression of p28GANK resulted in elevated expression of the endogenous protein. We also observed that both β-catenin and c-Myc were transcriptional activators of p28GANK, and a correlation between p28GANK overexpression and c-Myc, cyclin D1 and β-catenin activation in primary human HCC. Together, these results suggest that p28GANK expression is regulated by a positive feedback loop involving β-catenin, which may play a critical role in tumorigenesis and the progression of HCC.

Interleukin-1β/Iinterleukin-1 receptor-associated kinase 1 inflammatory signaling contributes to persistent Gankyrin activation during hepatocarcinogenesis

Hepatocellular carcinoma (HCC) is a prototype of inflammation‐associated cancer. Oncoprotein Gankyrin, which mostly increases in HCC, plays a critical role in HCC development and metastasis. However, the exact mechanism of Gankyrin up‐regulation in HCC remains unclear. A Gankyrin luciferase reporter was developed to screen a potential regulator for Gankyrin from a list of proinflammatory cytokines, and interleukin (IL)‐1β was found as one of its activators. In clinical premalignant and malignant liver disease samples, enhanced IL‐1β/interleukin‐1 receptor‐associated kinase 1 (IRAK‐1) signaling accompanied by increased Gankyrin was observed. Lower expression of Gankyrin and phospho‐IRAK‐1 are favorable prognostic markers for HCC. A similar correlation was observed in the diethylnitrosamine (DEN) model of rat hepatocarcinogenesis. The results from Gankyrin reporter activity, real‐time polymerase chain reaction, or immunoblotting further confirmed the up‐regulation of Gankyrin by IL‐1β/IRAK‐1 inflammatory signaling. Moreover, a series of Gankyrins truncated reporters were constructed, and electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP) were performed to analyze the properties of Gankyrin promoter. Mechanistically, the core promoter of Gankyrin contains the binding site of nuclear factor Y (NF‐Y) family members, which can recruit histone acetyltransferase coactivator E1A‐binding protein p300 (p300) or CREB‐binding protein (CBP) to promote Gankyrin transcription. Conversely, knockdown of NF‐Y, p300, or CBP inhibits Gankyrin expression. IL‐1β stimulation causes sequential phosphorylation of IRAK‐1, c‐Jun N‐terminal kinase (JNK), and p300 and enhances recruitment of the p300/CBP/NF‐Y complex to Gankyrin promoter. Inhibition of phospho‐JNK impairs IL‐1β/IRAK‐1 signaling‐mediated up‐regulation of Gankyrin. Conclusion: The finding of IL‐1β/IRAK‐1 signaling promoting Gankyrin expression through JNK and NF‐Y/p300/CBP complex provides a fresh view on inflammation‐enhanced hepatocarcinogenesis. (Hepatology 2015;61:585‐597)

p28GANK inhibits endoplasmic reticulum stress-induced cell death via enhancement of the endoplasmic reticulum adaptive capacity

It has been shown that oncoprotein p28GANK, which is consistently overexpressed in human hepatocellular carcinoma (HCC), plays a critical role in tumorigenesis of HCC. However, the underlying mechanism remains unclear. Here, we demonstrated that p28GANK inhibits apoptosis in HCC cells induced by the endoplasmic reticulum (ER) stress. During ER stress, p28GANK enhances the unfolded protein response, promotes ER recovery from translational repression, and thereby facilitates cells ability to cope with the stress conditions. Furthermore, p28GANK upregulates glucose-regulated protein 78 (GRP78), a key ER chaperone protein, which subsequently enhances the ER folding capacity and promotes recovery from ER stress. We also demonstrated that p28GANK increases p38 mitogen-activated protein kinase and Akt phosphorylation, and inhibits nuclear factor kappa B (NF-κB) activation under ER stress, which in turn contributes to GRP78 upregulation. Taken together, our results indicate that p28GANK inhibits ER stress-induced apoptosis in HCC cells, at least in part, by enhancing the adaptive response and GRP78 expression. We propose that p28GANK has potential implications for HCC progression under the ER stress conditions.

The Tyrosine Kinase c-Met Contributes to the Pro-tumorigenic Function of the p38 Kinase in Human Bile Duct Cholangiocarcinoma Cells

Background: Both p38 and c-Met are implicated in the tumorigenesis of cholangiocarcinoma. Results: The inhibition of p38 inhibits human cholangiocarcinoma cell proliferation and invasion through reducing the basal activity of c-Met. Conclusion: c-Met contributes to the pro-tumorigenic action of p38 in human cholangiocarcinoma cells. Significance: We identify a novel molecular mechanism that contributes to understanding the pro-tumorigenic activity of p38 in human cholangiocarcinoma cells. Pro-tumorigenic function of the p38 kinase plays a critical role in human cholangiocarcinogenesis. However, the underlying mechanism remains incompletely understood. Here, we report that c-Met, the tyrosine kinase receptor for hepatocyte growth factor (HGF), contributes to the pro-tumorigenic ability of p38 in human cholangiocarcinoma cells. Both p38 and c-Met promote the proliferation and invasion of human cholangiocarcinoma cells. Importantly, inhibition or knockdown of p38 decreased the basal activation of c-Met. Tyrosine phosphatase inhibitor studies revealed that p38 promotes the activity of c-Met, at least in part, by inhibiting dephosphorylation of the receptor. Moreover, density enhanced phosphatase-1 (DEP-1) is involved in p38-mediated inhibiting dephosphorylation of c-Met. Furthermore, p38 inhibits the degradation of c-Met. Taken together, these data provide a potential mechanism to explain how p38 promotes human cholangiocarcinoma cell proliferation and invasion. We propose that the link between p38 and c-Met is implicated in the progression of human cholangiocarcinoma.

Disturbance of Ca2+ Homeostasis Converts Pro-Met into Non-canonical Tyrosine Kinase p190MetNC in Response to Endoplasmic Reticulum Stress in MHCC97 Cells

Background: Both ER stress and c-Met are implicated in the tumorigenesis of HCC. Results: ER calcium disturbance-induced p190MetNC expression inhibits ER stress-mediated apoptosis. Conclusion: p190MetNC, but not p190Metαβ, plays a critical role in promoting HCC cells survival under ER stress. Significance: We identify a novel biochemical mechanism by which c-Met ensures HCC cell survival under stress conditions. c-Met, the tyrosine-kinase receptor for hepatocyte growth factor, plays a critical role in the tumorigenesis of hepatocellular carcinoma (HCC). However, the underlying mechanism remains incompletely understood. The mature c-Met protein p190Metαβ (consists of a α subunit and a β subunit) is processed from pro-Met. Here we show that pro-Met is processed into p190MetNC by sarco/endoplasmic reticulum calcium-ATPase (SERCA) inhibitor thapsigargin. p190MetNC compensates for the degradation of p190Metαβ and protects human HCC cells from apoptosis mediated by endoplasmic reticulum (ER) stress. In comparison with p190Metαβ, p190MetNC is not cleaved and is expressed as a single-chain polypeptide. Thapsigargin-initiated p190MetNC expression depends on the disturbance of ER calcium homeostasis. Once induced, p190MetNC is activated independent of hepatocyte growth factor engagement. p190MetNC contributes to sustained high basal activation of c-Met downstream pathways during ER calcium disturbance-mediated ER stress. Both p38 MAPK-promoted glucose-regulated protein 78 (GRP78) expression and sustained high basal activation of PI3K/Akt and MEK/ERK are involved in the cytoprotective function of p190MetNC. Importantly, the expression of p190MetNC is detected in some HCC cases. Taken together, these data provide a potential mechanism to explain how c-Met promotes HCC cells survival in response to ER stress. We propose that context-specific processing of c-Met protein is implicated in HCC progression in stressful microenvironments.