School of Biological Sciences, Universiti Sains Malaysia, 11800, Minden, Penang, Malaysia.
Centre for Chemical Biology, Universiti Sains Malaysia, Sains@USM, Block B No. 10, Persiaran Bukit Jambul, 11900, Bayan Lepas, Penang, Malaysia.
Sci Rep. 2018 Mar 1;8(1):3874. doi: 10.1038/s41598-018-22157-4.
The front-end desaturases (Fads) are rate-limiting enzymes responsible for production of long-chain polyunsaturated fatty acids (LC-PUFA). The full spectrum of the transcriptional regulation of fads is still incomplete, as cloning of fads promoter is limited to a few species. Here, we described the cloning and characterisation of the zebrafish fads2 promoter. Using 5'-deletion and mutation analysis on this promoter, we identified a specific region containing the sterol regulatory element (SRE) which is responsible for the activation of the fads2 promoter. In tandem, two conserved CCAAT boxes were also present adjacent to the SRE and mutation of either of these binding sites attenuates the transcriptional activation of the fads2 promoter. An in vivo analysis employing GFP reporter gene in transiently transfected zebrafish embryos showed that this 1754 bp upstream region of the fads2 gene specifically directs GFP expression in the yolk syncytial layer (YSL) region. This indicates a role for LC-PUFA in the transport of yolk lipids through this tissue layer. In conclusion, besides identifying novel core elements for transcriptional activation in zebrafish fads2 promoter, we also reveal a potential role for fads2 or LC-PUFA in YSL during development.
前端去饱和酶（Fads）是负责长链多不饱和脂肪酸（LC-PUFA）生成的限速酶。由于 fad 启动子的克隆仅限于少数几种物种，因此 fad 的转录调控的全貌仍不完整。在这里，我们描述了斑马鱼 fad2 启动子的克隆和特性。通过对该启动子进行 5'-缺失和突变分析，我们确定了一个包含固醇调节元件（SRE）的特定区域，该区域负责 fad2 启动子的激活。同时，在 SRE 附近还存在两个保守的 CCAAT 盒，突变其中任何一个结合位点都会减弱 fad2 启动子的转录激活。体内分析利用 GFP 报告基因在瞬时转染的斑马鱼胚胎中的实验表明，fads2 基因的这个 1754bp 上游区域特异性地在卵黄合胞层（YSL）区域指导 GFP 的表达。这表明 LC-PUFA 在通过该组织层运输卵黄脂质中起作用。总之，除了鉴定出斑马鱼 fad2 启动子中用于转录激活的新核心元件外，我们还揭示了 fad2 或 LC-PUFA 在发育过程中在 YSL 中的潜在作用。
