Identification of nuclear proteins interacting with the liver-specific enhancer B element of the antifreeze protein gene in winter flounder.

The intron of the winter flounder antifreeze protein (AFP) gene contains a liver-specific enhancer element B as demonstrated by transient expression in mammalian cells. Element B interacts with rat C/EBPalpha and a novel protein, tentatively designated as the antifreeze enhancer-binding protein (AEP). Present studies revealed that nuclear proteins from the winter flounder liver interact similarly and specifically to element B as shown by footprinting analysis and gel retardation assays. The presence of C/EBP in the flounder liver was confirmed by Western blot analysis. In vitro transcription assays in its homologous system further demonstrated the transactivation activity of the AFP gene intron. The present findings suggest that the mechanisms for regulating liver-specific transcription are evolutionarily conserved.