Hepatocyte nuclear factor 4α (Hnf4α) is involved in transcriptional regulation of Δ6/Δ5 fatty acyl desaturase (Fad) gene expression in marine teleost Siganus canaliculatus.

Abstract

As the first marine teleost demonstrated to biosynthesize long-chain polyunsaturated fatty acids (LC-PUFAs) from C18 precursors such as linoleic acid (LOA, 18:2n-6) and α-linolenic acid (ALA, 18:3n-3), the rabbitfish (Siganus canaliculatus) contains the complete enzymatic system for LC-PUFA biosynthesis, including Δ6/Δ5 fatty acid desaturase (Fad), Δ4 Fad, and elongase 5 (Elovl5). Previously, our group demonstrated that hepatocyte nuclear factor 4α (Hnf4α) is a transcription factor (TF) for rabbitfish Δ4 fad and elovl5, and interacts with the core promoter of Δ6/Δ5 fad. To fully clarify the role of Hnf4α in the regulation of LC-PUFA biosynthesis, the present study aimed to explore the regulatory role of Hnf4α on Δ6/Δ5 fad gene expression. First, Hnf4α overexpression and agonist assays identified the Hnf4α response region in the Δ6/Δ5 fad core promoter as -456 bp to +51 bp. Bioinformatic analysis predicted four potential Hnf4α binding elements in the core promoter, which were confirmed by site-directed mutation and functional assays in a dual luciferase assay system. Moreover, the mRNA expression levels of hnf4α, Δ6/Δ5 fad, and Δ4 fad were significantly increased in the S. canaliculatus hepatocyte line (SCHL) cells after treatment with Hnf4α agonists (Alverine and Benfluorex) or its mRNA overexpression. By contrast, the expression levels of these three genes were markedly decreased after hnf4a small interfering RNA (siRNA) transfection. The results indicated that Hnf4α has a regulatory effect on rabbitfish Δ6/Δ5 fad gene transcription, identifying Hnf4α as a TF of Δ6/Δ5 fad in vertebrates for the first time.