Hepatocyte Nuclear Factor 4α (HNF4α) Is a Transcription Factor of Vertebrate Fatty Acyl Desaturase Gene as Identified in Marine Teleost Siganus canaliculatus

Rabbitfish Siganus canaliculatus was the first marine teleost demonstrated to have the capability of biosynthesizing long-chain polyunsaturated fatty acids (LC-PUFA) from C-18 precursors, and to possess a Delta 4 fatty acyl desaturase (Delta 4 Fad) which was the first report in vertebrates, and is a good model for studying the regulatory mechanisms of LC-PUFA biosynthesis in teleosts. In order to understand regulatory mechanisms of transcription of Delta 4 Fad, the gene promoter was cloned and characterized in the present study. An upstream sequence of 1859 bp from the initiation codon ATG was cloned as the promoter candidate. On the basis of bioinformatic analysis, several binding sites of transcription factors (TF) including GATA binding protein 2 (GATA-2), CCAAT enhancer binding protein (C/EBP), nuclear factor 1 (NF-1), nuclear factor Y (NF-Y), hepatocyte nuclear factor 4 alpha (HNF4 alpha) and sterol regulatory element (SRE), were identified in the promoter by site-directed mutation and functional assays. HNF4 alpha and NF-1 were confirmed to interact with the core promoter of Delta 4 Fad by gel shift assay and mass spectrometry. Moreover, over-expression of HNF4 alpha increased promoter activity in HEK 293T cells and mRNA level of Delta 4 Fad in rabbitfish primary hepatocytes, respectively. The results indicated that HNF4 alpha is a TF of rabbitfish Delta 4 Fad. To our knowledge, this is the first report on promoter structure of a Delta 4 Fad, and also the first demonstration of HNF4 alpha as a TF of vertebrate Fad gene involved in transcription regulation of LC-PUFA biosynthesis.