Induction of Huh-7 cell apoptosis by HCV core proteins via CK1α-p53-Bid signaling pathway

Hepatitis C virus (HCV)-infected liver cells sensitize host cells to tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL)-induced cell apoptosis; however, the precise mechanisms are unknown. In the present study, flow cytometry demonstrated that the Annexin V-positive Huh-7 cell number was higher in groups transfected with core proteins when compared with the pcDNA3.1 group. The mRNA and protein expression levels of B-cell lymphoma 2 (Bcl-2) were negatively associated, while Bcl-2-associated X protein (Bax) were positively correlated, with cell apoptotic rate, which, were verified by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and western blotting. There were no significant differences in the expressions of casein kinase 1 (CK1)-ε, CK1γ or CK1δ; however, the mRNA and protein levels of CK1α were markedly higher in groups transfected with the T (those derived from the HCV-J6 strain), NT (those derived from non-tumor tissues) and C191 (those derived from tumor tissues) HCV core proteins than in mock group. When compared with the Mock and Negative Control (control known-down) groups, the mRNA and protein levels of CK1α were lower in the CK1α known-down group, and there were no marked Huh-7 cell morphological changes among the 3 groups. There was more sensitivity to cell apoptosis in CK1α-silenced, however, not in non-CK1α-silenced, Huh-7 cells. BH3 interacting-domain death agonist (Bid) protein levels in CK1α-silenced Huh-7 cells were higher when compared with non-CK1α-silenced Huh-7 cells, and the level of p53 that translocated to the nucleus increased. Chromatin immunoprecipitation-PCR demonstrated that p53 bound to human Bid gene promoter. The level of the Bid promoter in CK1α-silenced Huh-7 cells was significantly higher than in the non-CK1α-silenced Huh-7 cells. Electron microscopy indicated that p53 knockdown decreased HCV core protein and TRAIL-induced cell apoptosis. Bid/caspase-8 protein levels in CK1α-silenced Huh-7 cells that were transfected with p53 siRNA were lower than in the control group. The present study demonstrated that HCV core proteins sensitize host cells to TRAIL-induced cell apoptosis by activating the CK1α-p53-Bid dependent pathway.

[1]  A. Qu,et al.  HCV core protein represses the apoptosis and improves the autophagy of human hepatocytes. , 2015, International journal of clinical and experimental medicine.

[2]  T. McDonnell,et al.  Induction of p53-mediated transcription and apoptosis by exportin-1 (XPO1) inhibition in mantle cell lymphoma , 2014, Cancer science.

[3]  P. Schirmacher,et al.  Hepatocyte expression of TRAIL pathway regulators correlates with histopathological and clinical parameters in chronic HCV infection. , 2014, Pathology, research and practice.

[4]  Lei Zhang,et al.  High Prevalence of HIV, HCV and Tuberculosis and Associated Risk Behaviours among New Entrants of Methadone Maintenance Treatment Clinics in Guangdong Province, China , 2013, PloS one.

[5]  K. Bulak,et al.  P53 protein in proliferation, repair and apoptosis of cells , 2013, Protoplasma.

[6]  G. Song,et al.  p53 facilitates BH3-only BID nuclear export to induce apoptosis in the irrepairable DNA damage response. , 2011, Medical hypotheses.

[7]  Steffen Jung,et al.  CKIα ablation highlights a critical role for p53 in invasiveness control , 2011, Nature.

[8]  Y. Huang,et al.  HCV Subtype Characterization among Injection Drug Users: Implication for a Crucial Role of Zhenjiang in HCV Transmission in China , 2011, PloS one.

[9]  Yan Wang,et al.  MST1 Promotes Apoptosis through Regulating Sirt1-dependent p53 Deacetylation* , 2011, The Journal of Biological Chemistry.

[10]  Jen-Leih Wu,et al.  Thioacetamide accelerates steatohepatitis, cirrhosis and HCC by expressing HCV core protein in transgenic zebrafish Danio rerio. , 2008, Toxicology.

[11]  M. IborraColomino,et al.  Development of hepatocellular carcinoma (HCC) in peginterferon and ribavirin HCV-infected cirrhotic treated patients. , 2006 .

[12]  Xiaobin Long,et al.  [Study on the epidemiology and HCV genotype distribution of HIV/HCV co-infection among HIV infected blood donors in China]. , 2006, Zhonghua gan zang bing za zhi = Zhonghua ganzangbing zazhi = Chinese journal of hepatology.

[13]  Yu Pan,et al.  Regulation of p53-MDMX Interaction by Casein Kinase 1 Alpha , 2005, Molecular and Cellular Biology.

[14]  J. Houghton,et al.  Casein Kinase I Attenuates Tumor Necrosis Factor–Related Apoptosis-Inducing Ligand–Induced Apoptosis by Regulating the Recruitment of Fas-Associated Death Domain and Procaspase-8 to the Death-Inducing Signaling Complex , 2004, Cancer Research.

[15]  Ning Li,et al.  [The influence of HCV core protein and apoptosis on cellular telomerase activities]. , 2004, Zhonghua gan zang bing za zhi = Zhonghua ganzangbing zazhi = Chinese journal of hepatology.

[16]  J. Gueulette,et al.  Caspase 8-mediated cleavage of the pro-apoptotic BCL-2 family member BID in p53-dependent apoptosis. , 2003, Biochemical and biophysical research communications.

[17]  M. Kasuga,et al.  Different apoptotic regulation of TRAIL-caspase pathway in HBV- and HCV-related hepatocellular carcinoma. , 2003, International journal of molecular medicine.

[18]  C. Hahn,et al.  The HCV core protein acts as a positive regulator of fas-mediated apoptosis in a human lymphoblastoid T cell line. , 2000, Virology.

[19]  O. Yokosuka,et al.  HCV-core protein accelerates recovery from the insensitivity of liver cells to Fas-mediated apoptosis induced by an injection of anti-Fas antibody in mice. , 2000, Journal of hepatology.

[20]  S. D. Gross,et al.  The casein kinase Ialpha isoform is both physically positioned and functionally competent to regulate multiple events of mRNA metabolism. , 1999, Journal of cell science.

[21]  W. Fiers,et al.  Casein Kinase-1 Phosphorylates the p75 Tumor Necrosis Factor Receptor and Negatively Regulates Tumor Necrosis Factor Signaling for Apoptosis (*) , 1995, The Journal of Biological Chemistry.

[22]  Y. S. Kim,et al.  Interaction of HCV core protein with 14-3-3epsilon protein releases Bax to activate apoptosis. , 2007, Biochemical and biophysical research communications.

[23]  A. Wiseman.  p53 protein or BID protein select the route to either apoptosis (programmed cell death) or to cell cycle arrest opposing carcinogenesis after DNA damage by ROS. , 2006, Medical hypotheses.