miR-17 is involved in the regulation of LC-PUFA biosynthesis in vertebrates: Effects on liver expression of a fatty acyl desaturase in the marine teleost Siganus canaliculatus

The rabbitfish Siganus canaliculatus is the first marine teleost shown to be able to biosynthesize long-chain polyunsaturated fatty acids (LC-PUFA) from C18 PUFA precursors catalyzed by two fatty acyl desaturases (fad) including Δ4 Fad and Δ6/Δ5 Fad as well as two elongases (Elovl4 and Elovl5). Previously, hepatocyte nuclear factor 4α (Hnf4α) was demonstrated to be predominant in the transcriptional regulation of two fads. To clarify the regulatory mechanisms involved in rabbitfish lipogenesis, the present study focused on the regulatory role of Hnf4α to elovl5 expression and LC-PUFA biosynthesis. Bioinformatics analysis predicted two potential Hnf4α elements in elovl5 promoter, one binding site was confirmed to interact with Hnf4α by gel shift assays. Moreover, overexpression of hnf4α caused a remarkable increase both in elovl5 promoter activity and mRNA contents, while knock-down of hnf4α in S. canaliculatus hepatocyte line (SCHL) resulted in a significant decrease of elovl5 gene expression. Meanwhile, hnf4α overexpression enhanced LC-PUFA biosynthesis in SCHL cell, and intraperitoneal injection to rabbitfish juveniles with Hnf4α agonists (Alverine and Benfluorex) increased the expression of hnf4α, elvol5 and Δ4 fad, coupled with an increased proportion of total LC-PUFA in liver. The results demonstrated that Hnf4α is involved in LC-PUFA biosynthesis by up-regulating the transcription of the elovl5 gene in rabbitfish, which is the first report of Hnf4α as a transcription factor of the elovl5 gene in vertebrates.

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

Gene ◽

10.1016/j.gene.2018.01.031 ◽

2018 ◽

Vol 647 ◽

pp. 174-180 ◽

Cited By ~ 24

Author(s):

Yewei Dong ◽

Jianhong Zhao ◽

Junliang Chen ◽

Shuqi Wang ◽

Yang Liu ◽

...

Keyword(s):

Gene Promoter ◽

Fatty Acyl ◽

Marine Teleost ◽

Siganus Canaliculatus ◽

Fatty Acyl Desaturase

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The miR-33 gene is identified in a marine teleost: a potential role in regulation of LC-PUFA biosynthesis in Siganus canaliculatus

Scientific Reports ◽

10.1038/srep32909 ◽

2016 ◽

Vol 6 (1) ◽

Cited By ~ 9

Author(s):

Qinghao Zhang ◽

Cuihong You ◽

Shuqi Wang ◽

Yewei Dong ◽

Óscar Monroig ◽

...

Keyword(s):

Potential Role ◽

Marine Teleost ◽

Siganus Canaliculatus ◽

Pufa Biosynthesis

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Identification of miR-145 as a Key Regulator Involved in LC-PUFA Biosynthesis by Targeting hnf4α in the Marine Teleost Siganus canaliculatus

Journal of Agricultural and Food Chemistry ◽

10.1021/acs.jafc.0c04649 ◽

2020 ◽

Vol 68 (51) ◽

pp. 15123-15133

Author(s):

Cuiying Chen ◽

Mei Zhang ◽

Yuanyou Li ◽

Shuqi Wang ◽

Dizhi Xie ◽

...

Keyword(s):

Marine Teleost ◽

Siganus Canaliculatus ◽

Pufa Biosynthesis

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Transcriptional regulation mechanism of sterol regulatory element binding proteins on Δ6 fatty acyl desaturase in razor clam Sinonovacula constricta

British Journal Of Nutrition ◽

10.1017/s0007114520002068 ◽

2020 ◽

Vol 124 (9) ◽

pp. 881-889

Author(s):

Zhaoshou Ran ◽

Fei Kong ◽

Jilin Xu ◽

Kai Liao ◽

Xiaojun Yan

Keyword(s):

Binding Proteins ◽

Regulatory Element ◽

Fatty Acyl ◽

Regulation Mechanism ◽

Sinonovacula Constricta ◽

Marine Molluscs ◽

Razor Clam ◽

Pufa Biosynthesis ◽

Sterol Regulatory Element ◽

Fatty Acyl Desaturase

AbstractThe razor clam, Sinonovacula constricta, contains high levels of long-chain PUFA (LC-PUFA), which are critical for human health. In addition, S. constricta is the first marine mollusc demonstrated to possess Δ6 fatty acyl desaturase (Fad) and complete LC-PUFA biosynthetic ability, providing a good representative to investigate the molecular mechanism of sterol regulatory element binding proteins (SREBP) in regulating Δ6 Fad for LC-PUFA biosynthesis in marine molluscs. Herein, S. constricta SREBP and Δ6 Fad promoter were cloned and characterised. Subsequently, dual luciferase and electrophoretic mobility shift assays were conducted to explore the SREBP binding elements in the core regulatory region of S. constricta Δ6 Fad promoter. Results showed that S. constricta SREBP had a very conservative basic helix-loop-helix-leucine zipper motif, while S. constricta Δ6 Fad promoter exhibited very poor identity with teleost Fads2 promoters, indicating their differentiation during evolution. A 454 bp region harbouring a core sequence in S. constricta Δ6 Fad promoter was predicted to be essential for the transcriptional activation by SREBP. This was the first report on the regulatory mechanism of LC-PUFA biosynthesis in marine molluscs, which would facilitate optimising the LC-PUFA biosynthetic pathway of bivalves in further studies.

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Sp1 is Involved in Vertebrate LC-PUFA Biosynthesis by Upregulating the Expression of Liver Desaturase and Elongase Genes

International Journal of Molecular Sciences ◽

10.3390/ijms20205066 ◽

2019 ◽

Vol 20 (20) ◽

pp. 5066 ◽

Cited By ~ 3

Author(s):

Yuanyou Li ◽

Jianhong Zhao ◽

Yewei Dong ◽

Ziyan Yin ◽

Yang Li ◽

...

Keyword(s):

Binding Site ◽

Direct Interaction ◽

Bioinformatic Analysis ◽

Fatty Acyl ◽

Luciferase Reporter ◽

Mrna Levels ◽

Mobility Shift ◽

Siganus Canaliculatus ◽

Pufa Biosynthesis ◽

Mobility Shift Assay

The rabbitfish Siganus canaliculatus was the first marine teleost demonstrated to have the ability for the biosynthesis of long-chain (≥C20) polyunsaturated fatty acids (LC-PUFA) from C18 PUFA precursors, and all the catalytic enzymes including two fatty acyl desaturase 2 (Δ4 Fads2 and Δ6/Δ5 Fads2) and two elongases (Elovl4 and Elovl5) have been identified, providing a good model for studying the regulatory mechanisms of LC-PUFA biosynthesis in fish. Stimulatory protein 1 (Sp1) has been speculated to be a vital transcription factor in determining the promoter activity of Fads-like genes in fish, however its regulatory effects on gene expression and LC-PUFA biosynthesis have not been demonstrated. Bioinformatic analysis predicted potential Sp1 binding sites in the promoters of the rabbitfish Δ6/Δ5 fads2 and elovl5, but not in Δ4 fads2 promoter. Here we cloned full-length cDNA of the rabbitfish sp1 gene, which encoded a putative protein of 701 amino acids, and was expressed in all tissues studied with highest levels in gill and eyes. The dual luciferase reporter assay in HepG2 line cells demonstrated the importance of the Sp1 binding site for the promoter activities of both Δ6/Δ5 fads2 and elovl5. Moreover, the electrophoretic mobility shift assay confirmed the direct interaction of Sp1 with the two promoters. Insertion of the Sp1 binding site of Δ6/Δ5 fads2 promoter into the corresponding region of the Δ4 fads2 promoter significantly increased activity of the latter. In the Siganus canaliculatus hepatocyte line (SCHL) cells, mRNA levels of Δ6/Δ5 fads2 and elovl5 were positively correlated with the expression of sp1 when sp1 was overexpressed or knocked-down by RNAi or antagonist (mithramycin) treatment. Moreover, overexpression of sp1 also led to a higher conversion of 18:2n−6 to 18:3n−6, 18:2n−6 to 20:2n−6, and 18:3n−3 to 20:3n−3, which related to the functions of Δ6/Δ5 Fads2 and Elovl5, respectively. These results indicated that Sp1 is involved in the transcriptional regulation of LC-PUFA biosynthesis by directly targeting Δ6/Δ5 fads2 and elovl5 in rabbitfish, which is the first report of Sp1 involvement in the regulation of LC-PUFA biosynthesis in vertebrates.

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