Interferon-alpha enhances TRAIL-mediated apoptosis by up-regulating caspase-8 transcription in human hepatoma cells.
OBJECTIVE
IFNalpha is an approved treatment option for patients chronically infected with the hepatitis B and C viruses. Additionally, there is an indication for tumor therapy. The exact mechanisms underlying the antiviral and antitumor effects of IFNalpha are not completely understood. In this study, we investigated if the pro-apoptotic factor caspase-8 is a target gene of IFNalpha signalling.


METHODS
Huh7 hepatoma cells were used for measuring caspase-8 promoter activity in luciferase reporter assays after IFNalpha stimulation. Caspase-8 expression was monitored by RT-PCR, immunoblotting and measurement of enzymatic activity. Functional caspase-8 promoter elements were identified in gelshift assays and by site directed mutagenesis. Caspase-8 was inhibited using siRNA.


RESULTS
IFNalpha treatment induced caspase-8 promoter activity and mRNA expression. We identified a unique promoter element mediating the IFNalpha-dependent increase in caspase-8 transcription. Up-regulation of caspase-8 expression by IFNalpha had no impact on the rate of apoptosis per se. However, co-stimulation with IFNalpha doubled TRAIL-mediated apoptosis and enzymatic caspase-8 activity. The synergistic effect of TRAIL and IFNalpha could be blocked by inhibiting caspase-8 expression.


CONCLUSIONS
We demonstrate that caspase-8 is a target gene of IFNalpha and provide evidence showing that IFNalpha treatment sensitizes cells for apoptosis via enhanced caspase-8 transcription.
Liedtke
Christian
C
Department of Gastroenterology, Hepatology and Endocrinology, Medizinische Hochschule Hannover, Carl-Neuberg-Strasse 1, D-30625 Hannover, Germany. chrliedtke@aol.com
Gröger
Nadine
N
Manns
Michael P
MP
Trautwein
Christian
C
D000970:Antineoplastic Agents / Q000627:therapeutic use; D017209:Apoptosis; D051017:Apoptosis Regulatory Proteins / Q000378:metabolism*; D015153:Blotting, Western; D006528:Carcinoma, Hepatocellular / Q000188:drug therapy* / Q000378:metabolism; D053181:Caspase 8; D020169:Caspases / Q000187:drug effects / Q000235:genetics*; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D066298:In Vitro Techniques; D016898:Interferon-alpha / Q000627:therapeutic use*; D008113:Liver Neoplasms / Q000188:drug therapy* / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000378:metabolism*; D012333:RNA, Messenger / Q000096:biosynthesis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D053221:TNF-Related Apoptosis-Inducing Ligand; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism*; D015854:Up-Regulation
D000970:Antineoplastic Agents; D051017:Apoptosis Regulatory Proteins; D016898:Interferon-alpha; D008562:Membrane Glycoproteins; D012333:RNA, Messenger; D053221:TNF-Related Apoptosis-Inducing Ligand; C505653:TNFSF10 protein, human; D014409:Tumor Necrosis Factor-alpha; C505530:CASP8 protein, human; D053181:Caspase 8; D020169:Caspases