Cloning and characterization of ∆6/∆5 fatty acyl desaturase (Fad) gene promoter in the marine teleost Siganus canaliculatus

Citation

Dong Y, Zhao J, Chen J, Wang S, Liu Y, Zhang Q, You C, Monroig O, Tocher DR & Li Y (2018) Cloning and characterization of ∆6/∆5 fatty acyl desaturase (Fad) gene promoter in the marine teleost Siganus canaliculatus. Gene, 647, pp. 174-180. https://doi.org/10.1016/j.gene.2018.01.031

Abstract
The rabbitfish Siganus canaliculatus was the first marine teleost demonstrated to have the ability of biosynthesizing long-chain polyunsaturated fatty acids (LC-PUFA) from C18 PUFA precursors, and all genes encoding the key enzymes for LC-PUFA biosynthesis have been cloned and functionally characterized, which provides us a potential model to study the regulatory mechanisms of LC-PUFA biosynthesis in teleosts. As the primary step to clarify such mechanisms, present research focused on promoter analysis of gene encoding ∆6/∆5 fatty acyl desaturase (Fad), a rate-limiting enzyme catalyzing the first step in the conversion of C18 PUFA to LC-PUFA. First, 2044 bp promoter sequence was cloned by genome walking, and the sequence from -456 bp to + 51bp was determined as core promoter by progressive deletion mutation. Moreover, binding sites of transcription factors (TF) such as CCAAT enhancer binding protein (C/EBP), nuclear factor 1 (NF-1), stimulatory protein 1 (Sp1), nuclear factor Y (NF-Y), activated protein 1 (AP1), sterol regulatory element (SRE), hepatocyte nuclear factor 4α (HNF4α) and peroxisome proliferator activated receptor γ (PPARγ) were identified in the core promoter by site-directed mutation and functional assays. Moreover, NF-1 and HNF4α were confirmed to interact with the core promoter region by gel shift assay and mass spectrometry. This is the first report of the promoter structure of a ∆6/∆5 Fad in a marine teleost, and a novel discovery of NF-1 and HNF4α binding to the ∆6/∆5 Fad promoter.

Keywords
Fatty acyl desaturase; LC-PUFA biosynthesis; transcriptional regulation mechanism; rabbitfish Siganus canaliculatus; marine teleost; NF-1; HNF4α

Journal
Gene: Volume 647

• Dong D6D5 promoter (Gene 2018).pdf