Organization and Promoter Analysis of the Zebrafish (Danio rerio) Interferon Gene

Abstract

Interferon plays important roles in confronting viral infections as the first line of defense. For the purpose of understanding the molecular mechanism which controls transcription of the interferon gene, we cloned and sequenced the interferon promoter region of the zebrafish interferon gene and characterized its activity using firefly luciferase transient transfection expression assays. Different fragments of the zebrafish interferon 5'-flanking region were transfected into ZFL cells. In these cell lines, maximum promoter activity was located in 2.2 kb of the zebrafish interferon 5' flanking region of the ZFL cell line. In this study, we investigated whether the viral replicative intermediate double-stranded RNA (herein we used synthetic polyinosinic-polycytidylic acid [poly(I):poly(C)] modifies the effects of interferon on gene expression. For this purpose, all zebrafish interferon promoter fragments were treated with either 1, 10, or 100 microg/ml poly(I):poly(C). The results showed that after treatment with 10 microg/ml poly(I):poly(C), high promoter activity was observed in the -2.2-kb interferon promoter fragment. Several putative transcription factors were shown in the promoter region, including IRF-1, C/EBP, NFkappaB, and GATA-1. Further study of the in vivo expression of the interferon promoter during development was carried out in transgenic zebrafish expressing an interferon promoter-driven green fluorescent protein (GFP) encoding the GFP cDNA transgene. Morphological studies of transgenic zebrafish indicated that the interferon promoter-driven GFP transcripts appeared in the yolk, head, and lymphoid organs. These results indicate that the interferon promoter is active in a tissue-specific manner, and suggest that the interferon promoter plays an important role in virus resistance during teleost growth.