Identification of a functional transcriptional factor AP-1 site in the sheep interferon tau gene that mediates a response to PMA in JEG3 cells.
To examine regulatory mechanisms of sheep interferon tau (oIFNtau) gene expression, potential enhancer/silencer elements of the oIFNtau gene were examined using a transient transfection system with oIFNtau gene-chloramphenicol acetyltransferase (oIFNtau-CAT) reporter constructs in human choriocarcinoma cells, JEG3. Experiments with 5'-deletion constructs revealed that the upstream regions from bases -654 to -607 and from bases -606 to -555 were essential for oIFNtau gene expression. In a heterologous transcriptional system in which the upstream regions of oIFNtau were inserted in front of simian virus 40 (SV40) promoter, the regions between bases -654 and -555 were determined as being the enhancer region required for oIFNtau-SV40-CAT transactivation. A subsequent study with the oIFNtau-CAT constructs lacking the upstream region between bases -542 and -124 revealed that, in addition to the further upstream region between bases -1000 and -654, the sequences from bases -543 to -452 seemed to act as silencer regions. The oIFNtau-CAT constructs with site-specific mutagenesis revealed that multiple enhancer elements existed between bases -654 and -555 of the oIFNtau gene. On the basis of nucleotide sequence analysis, there are numerous sites between bases -654 and -555 to which potential transcriptional factors, AP-1, GATA and GATA-related proteins, could bind. Furthermore, gel mobility-shift assays revealed that AP-1 or other nuclear factors could bind to these elements. In co-transfection studies, the expression of c-Jun plus c-Fos enhanced the transactivation of oIFNtau-CAT but the expression of GATA-1, GATA-2 or GATA-3 did not. Taken together, these results suggest that the upstream region between bases -654 and -555 could be considered as the enhancer region for oIFNtau gene transactivation.
Yamaguchi
H
H
Laboratory of Animal Breeding, Faculty of Agriculture, University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan.
Ikeda
Y
Y
Moreno
J I
JI
Katsumura
M
M
Miyazawa
T
T
Takahashi
E
E
Imakawa
K
K
Sakai
S
S
Christenson
R K
RK
D000818:Animals; D001483:Base Sequence; D001667:Binding, Competitive; D016384:Consensus Sequence / Q000235:genetics; D015342:DNA Probes / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D004742:Enhancer Elements, Genetic / Q000235:genetics; D005786:Gene Expression Regulation / Q000187:drug effects*; D017930:Genes, Reporter; D006801:Humans; D007370:Interferon Type I / Q000235:genetics*; D016297:Mutagenesis, Site-Directed; D009687:Nuclear Proteins / Q000378:metabolism; D011257:Pregnancy Proteins / Q000235:genetics*; D011401:Promoter Regions, Genetic / Q000235:genetics; D020218:Response Elements / Q000235:genetics*; D017384:Sequence Deletion; D012756:Sheep / Q000235:genetics*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D018808:Transcription Factor AP-1 / Q000378:metabolism*; D014157:Transcription Factors / Q000235:genetics / Q000502:physiology; D014162:Transfection; D014407:Tumor Cells, Cultured
D015342:DNA Probes; D004268:DNA-Binding Proteins; D007370:Interferon Type I; D009687:Nuclear Proteins; D011257:Pregnancy Proteins; D018808:Transcription Factor AP-1; D014157:Transcription Factors; C020947:interferon tau; D013755:Tetradecanoylphorbol Acetate