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Hirohito Yamaguchi1 , Momoko Katsumura1 , Kazuhiko Imakawa1 *, Senkiti Sakai1 , Ronald Christenson2 *
Laboratory of Animal Breeding, Faculty of Agriculture, University of Tokyo, Tokyo 1
USDA-ARS, Roman L. Hruska U.S. Meat Animal Research Center, Clay Center, NE 2
To examine regulatory mechanisms of ovine interferon tau (oIFN) gene expression, potential enhancer/silencer elements of the IFN gene were studied using a transient transfection system with oIFN gene-chloramphenicol acetyltransferase reporter (oIFN-CAT) constructs in human choriocarcinoma cells, JEG3. The experiments, with 5-deletion constructs or a heterologous transcriptional system in which the upstream regions of oIFN were inserted in front of the SV40 promoter, revealed that the upstream region from -654 to -555 bases was the enhancer region required for oIFN transactivation. The oIFN-CAT constructs with site-specific mutagenesis revealed that multiple enhancer elements existed between -654 and -555 bases. A subsequent study with oIFN-CAT constructs designed for detailed analysis of silencer elements revealed that the sequences from -700 to -655 bases and from -502 to -453 bases seemed to act as silencer regions. Gel mobility shift assays revealed that AP-1, GATA and GATA related protein could bind to enhancer elements and that several nuclear factors could bind to the silencer regions if extracted from trophoblasts on day 20, but not day 14 of pregnancy. In co-transfection studies, AP-1 expression enhanced oIFN-CAT transactivation; GATA-1, -2 or -3 expression did not. These results suggest that the upstream region between -654 and -555 bases could be considered as the enhancer region and regions between -700 and -655, -502 and -453 as the silencer for oIFN gene transactivation.
This abstract is being presented on Tuesday, August 3 at 8:00 AM to 10:15 AM at CUB 2nd Floor Ballroom.