A 5' extended IFN-stimulating response element is crucial for IFN-gamma-induced tripartite motif 22 expression via interaction with IFN regulatory factor-1.
Interferon-gamma is crucial for the noncytopathic clearance of hepatitis B virus. In our previous study, we demonstrated that an IFN-gamma-inducible molecule, tripartite motif (TRIM) 22, played an important role in antiviral immunity against hepatitis B virus. However, the molecular mechanism of TRIM22 induction by IFN-gamma is still unclear. In this study, we identified a novel cis-element termed 5' extended IFN-stimulating response element (5' eISRE) that was crucial for IFN-gamma inducibility of TRIM22 through transfection assays with luciferase reporter constructs and EMSAs. The 5' eISRE consists of an ISRE-like motif (ACTTTCGTTTCTC) and a 6-bp sequence (AATTTA) upstream of it, and all three thymine triplets of this cis-element (AATTTAACTTTCGTTTCTC) were revealed to contribute to the IFN-gamma inducibility of TRIM22 by site-directed mutagenesis. Further studies showed that upon IFN-gamma stimulation, the 5' eISRE could be bound by IFN regulatory factor-1 (IRF-1), but not by STAT1, as demonstrated by supershift analysis and an ELISA-based transcription factor assay. Moreover, overexpression of IRF-1 significantly induced TRIM22 expression, whereas silencing of IRF-1 with specific short interference RNA abolished IFN-gamma-induced TRIM22 expression in HepG2 cells, indicating an IRF-1-dependent expression of TRIM22. Taken together, it was demonstrated in this study that a novel cis-element, 5' eISRE, was crucial for the IFN-gamma-induced transcriptional activity of the TRIM22 gene via interaction with IRF-1.
Gao
Bo
B
Institute for Immunobiology, Department of Immunology, Shanghai Medical College of Fudan University, Shanghai, People's Republic of China.
Wang
Yaxin
Y
Xu
Wei
W
Duan
Zhijian
Z
Xiong
Sidong
S
D024506:5' Flanking Region / Q000235:genetics; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D015153:Blotting, Western; D003513:Cycloheximide / Q000494:pharmacology; D024202:Electrophoretic Mobility Shift Assay; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D056945:Hep G2 Cells; D006801:Humans; D050856:Interferon Regulatory Factor-1 / Q000235:genetics / Q000378:metabolism*; D007371:Interferon-gamma / Q000494:pharmacology*; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D015778:Minor Histocompatibility Antigens; D009154:Mutation; D011485:Protein Binding / Q000187:drug effects; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D034622:RNA Interference; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism*; D020218:Response Elements / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection; D000070584:Tripartite Motif Proteins
D050856:Interferon Regulatory Factor-1; D015778:Minor Histocompatibility Antigens; D011500:Protein Synthesis Inhibitors; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins; C094314:TRIM22 protein, human; D000070584:Tripartite Motif Proteins; D007371:Interferon-gamma; D003513:Cycloheximide; D008156:Luciferases