核因子 kappa B/p65 在卵形鲳鲹 Epinephelus coioides 过氧化物酶体增殖物激活受体 δ 表达中发挥正向作用。
Nuclear factor kappa B/p65 plays a positive role in peroxisome proliferator-activated receptor δ expression in orange-spotted grouper Epinephelus coioides.
机构信息
Joint Laboratory of Guangdong Province and Hong Kong Region on Marine Bioresource Conservation and Exploitation, College of Marine Sciences, South China Agricultural University, Guangzhou, 510642, China; Guangdong Laboratory for Lingnan Modern Agriculture, Guangzhou, 510642, China.
Joint Laboratory of Guangdong Province and Hong Kong Region on Marine Bioresource Conservation and Exploitation, College of Marine Sciences, South China Agricultural University, Guangzhou, 510642, China; Guangdong Laboratory for Lingnan Modern Agriculture, Guangzhou, 510642, China; Laboratory for Marine Biology and Biotechnology, Qingdao National Laboratory for Marine Science and Technology, Qingdao, 266000, China.
出版信息
Fish Shellfish Immunol. 2020 Jul;102:101-107. doi: 10.1016/j.fsi.2020.03.060. Epub 2020 Apr 4.
The transcription factor nuclear factor kappa B (NF-κB) is a critical regulator of immune and inflammatory responses with crucial roles in various pathophysiologic conditions involving cell survival and death. Recent studies in mammals showed that NF-κB was also involved in peroxisome proliferator-activated receptors (PPARs)-mediated immune responses However, the mechanism by which NF-κB regulates PPARδ in teleosts remains unclear. In the present study, we analyzed the potential role of NF-κB/p65 (Ecp65) in the immune response stimulated by various pathogens in the grouper Epinephelus coioides. Ecp65 expression was significantly induced soon after infection with lipopolysaccharide, nervous necrosis virus, poly(I:C), and zymosan A. We also analyzed the promoter to determine the regulatory effect of Ecp65 on PPARδ expression, using progressive EcPPARδ promoter deletion mutations. Among the five truncated mutants, the luciferase reporter activity of the PPARδ-5 promoter region was highest in response to Ecp65, indicating that the core p65-binding region was located in the PPARδ-5 promoter region (+122 bp to +383 bp). Mutation analyses indicated that the luciferase reporter activity of the EcPPARδ promoter was dramatically decreased by mutation of the M3 (+305 bp to +324 bp) and M4 (+346 bp to +365 bp) binding sites, respectively. We further confirmed that Ecp65 bound to the M3 and M4 binding sites in the 5'-untranslated region of EcPPARδ by electrophoretic mobility shift assay. Finally, overexpression of Ecp65 in vitro notably promoted the transcription of EcPPARδ, interferon-related genes, and several inflammatory cytokines. This study demonstrated that Ecp65 plays an important role in modulating the innate immune responses in groupers. These results also further our understanding of the mechanisms involved in the transcriptional regulation of PPARs by p65 in bony fish.
转录因子核因子 kappa B（NF-κB）是免疫和炎症反应的关键调节剂，在涉及细胞存活和死亡的各种病理生理条件中具有重要作用。最近在哺乳动物中的研究表明，NF-κB 也参与过氧化物酶体增殖物激活受体（PPARs）介导的免疫反应。然而，NF-κB 调节硬骨鱼中 PPARδ 的机制尚不清楚。在本研究中，我们分析了 NF-κB/p65（Ecp65）在石斑鱼 Epinephelus coioides 中受各种病原体刺激的免疫反应中的潜在作用。Ecp65 的表达在感染脂多糖、神经坏死病毒、聚肌苷酸和酵母聚糖 A 后很快被诱导。我们还分析了启动子，以确定 Ecp65 对 PPARδ 表达的调节作用，使用渐进式 EcPPARδ 启动子缺失突变。在五个截短突变体中，Ecp65 对 PPARδ-5 启动子区域的荧光素酶报告基因活性最高，表明核心 p65 结合区域位于 PPARδ-5 启动子区域（+122 bp 至+383 bp）。突变分析表明，EcPPARδ 启动子的荧光素酶报告基因活性因 M3（+305 bp 至+324 bp）和 M4（+346 bp 至+365 bp）结合位点的突变而显著降低。我们进一步通过电泳迁移率变动分析证实，Ecp65 结合到 EcPPARδ 的 5'-非翻译区的 M3 和 M4 结合位点。最后，体外过表达 Ecp65 显著促进了 EcPPARδ、干扰素相关基因和几种炎症细胞因子的转录。本研究表明，Ecp65 在调节石斑鱼固有免疫反应中发挥重要作用。这些结果还进一步了解了 p65 在硬骨鱼中调节 PPARs 转录的机制。
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