Functional analysis and temporal expression of promoter regions from fish antifreeze protein genes in transgenic Japanese medaka embryos.

Several series of sequences that are upstream of the transcriptional start site of different types of fish AFP genes were fused to the bacterial CAT gene, and their transcriptional role was examined in a transient expression assay after microinjection into Japanese medaka (Oryzias latipes) embryos at the 1-4 cell stage. Our studies demonstrated that the AFP genes have functional promoter regions containing positive as well as negative regulatory regions, indicating that these genes could be regulated at multiple sites. We also observed a promoter-specific pattern of temporal expression. Typically, the CAT expression was low in the first 4 days of embryonic development or before the stage of body pigmentation, followed by a sharp increase. The high level was maintained until hatching (11-13 days after fertilization), by which time the activity decreased to a very low level.