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

Abstract

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 p28 GANK inhibits apoptosis in HCC cells induced by the endoplasmic reticulum (ER) stress. During ER stress, p28 GANK enhances the unfolded protein response, promotes ER recovery from translational repression, and thereby facilitates cell's 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 p28 GANK 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 p28 GANK inhibits ER stress-induced apoptosis in HCC cells, at least in part, by enhancing the adaptive response and GRP78 expression. We propose that p28 GANK has potential implications for HCC progression under the ER stress conditions.

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

“…As ER stress activates non-UPR specific survival pathways such as PI3K/Akt and MEK/ERK in parallel with the UPR pathways, we focused on the role of c-Met in regulating non-UPR specific pathways. Because the PI3K/Akt and MEK/ ERK pathways, two downstream signaling pathways of c-Met (22), play a pivotal role in protecting cells against ER stress (28), it is reasonable to suggest that p190Met NC might inhibit thapsigargin-induced apoptosis through the PI3K/ Akt and MEK/ERK pathways. p190Met NC inhibition or knockdown led to PI3K/Akt and MEK/ERK inactivation in ER-stressed MHCC97-H and MHCC97-L cells.…”

“…Although the UPR is generally activated in various solid tumors, it is unclear how tumor cells adapt to long term ER stress. We have previously shown that HCC cells are relatively resistant to ER stress-induced apoptosis (28), but the detailed mechanisms remain largely unknown. The present work reveals that a non-canonical form of c-Met tyrosine-kinase receptor p190Met NC exerts a potent protective effect against prolonged ER stress mediated by ER Ca 2ϩ disturbance in HCC cells.…”

“…As PI3K/Akt and MEK/ERK have been shown to prevent ER stress-induced cell death (28,33) and the activity of Akt and ERK in thapsigargin-treated MHCC97-H and MHCC97-L cells was p190Met NC -dependent, we suspected that p190Met NC might protect HCC cells against ER stress through the PI3K/ Akt and MEK/ERK pathways. This hypothesis was supported by the data, which demonstrated that PI3K inhibitor LY294002 promoted MHCC97-H and MHCC97-L cell apoptosis in response to thapsigargin (Fig.…”

“…Antibody against IR was purchased from Bioworld Technology Corp. HCC cell lines MHCC-97H, MHCC-97L, SMMC-7721, HepG2, PLC, and Huh-7 were cultured in Dulbecco's modified Eagle's medium supplemented with 10% fetal bovine serum and 1% penicillin/ streptomycin in a humidified incubator containing 5% CO 2 and 95% ambient air at 37°C. The protocol used for c-Met and ATF4 knockdown has been previously described (28).…”

“…Protein concentrations were measured using the BCA assay (Santa Cruz Biotechnology). Protein samples were denatured with 4ϫ SDS-loading buffer (200 mM Tris, pH 6.8, 8% SDS, 400 mM DTT, 0.4% bromphenol blue, 40% glycerol) at 100°C for 5 min and subjected to standard SDS-PAGE and Western blot analysis as previously described (28).…”