Key Laboratory of Experimental Marine Biology, Institute of Oceanology, Chinese Academy of Sciences, Qingdao 266071, PR China.
Fish Shellfish Immunol. 2013 May;34(5):1209-15. doi: 10.1016/j.fsi.2013.01.009. Epub 2013 Feb 9.
The Rel/NF-κB transcription factors can function as key regulators to modulate the expression of immune-related genes in response to immune challenge or environmental stress. In the present study, a gene coding Rel/NF-κB homologue was identified from scallop Chlamys farreri (designated CfRel). Its deduced protein comprised 359 amino acids, and contained a conserved N-terminal Rel homology domain (RHD) and an IPT domain. There was an NF-κB/Rel/dorsal domain signature sequence in the RHD domain. The mRNA transcripts of CfRel could be detected in all the tested tissues including adductor muscle, mantle, gill, gonad, haemocytes, kidney and hepatopancreas, with the highest expression level in hepatopancreas. After LPS stimulation, there were two peaks of CfRel mRNA expression level in haemocytes at 6 h (25.25-fold, P < 0.05) and 24 h (59.66-fold, P < 0.05) respectively, while the mRNA expression of CfRel was only up-regulated at 3 h after PGN stimulation (2.35-fold, P < 0.05). By Western blotting technique, CfRel protein was observed in the cytoplasm and nucleus of scallop haemocytes, and its concentration in the haemocyte nucleus increased significantly at 3 h and 12 h after LPS stimulation. The noticeable NF-κB transcription activity of CfRel protein was determined by NF-κB luciferase reporter assays (122.43%, P < 0.05), and it decreased significantly (17.61%, P < 0.05) after the coexpression of scallop IκB protein. These results collectively suggested that CfRel mRNA transcripts and protein were induced by immune stimulation, and CfRel protein could extricate itself from IκB protein and transfer into the haemocyte nucleus to modulate the immune response in scallop.
Rel/NF-κB 转录因子可以作为关键调节因子，调节免疫相关基因的表达，以应对免疫挑战或环境压力。在本研究中，从扇贝 Chlamys farreri 中鉴定出一个编码 Rel/NF-κB 同源物的基因（命名为 CfRel）。其推导的蛋白质包含 359 个氨基酸，含有保守的 N 端 Rel 同源结构域（RHD）和 IPT 结构域。在 RHD 结构域中存在一个 NF-κB/Rel/dorsal 结构域签名序列。CfRel 的 mRNA 转录本可在所有测试组织中检测到，包括闭壳肌、套膜、鳃、性腺、血细胞、肾脏和肝胰腺，其中肝胰腺中的表达水平最高。在 LPS 刺激后，血细胞中 CfRel mRNA 表达水平有两个高峰，分别在 6 h（25.25 倍，P < 0.05）和 24 h（59.66 倍，P < 0.05），而在 PGN 刺激后仅在 3 h 时 CfRel mRNA 表达上调（2.35 倍，P < 0.05）。通过 Western blot 技术观察到 CfRel 蛋白存在于扇贝血细胞的细胞质和细胞核中，在 LPS 刺激后 3 h 和 12 h 时，其在血细胞核中的浓度显著增加。通过 NF-κB 荧光素酶报告基因检测，确定 CfRel 蛋白具有明显的 NF-κB 转录活性（122.43%，P < 0.05），而在共表达扇贝 IκB 蛋白后，其活性显著降低（17.61%，P < 0.05）。这些结果共同表明，CfRel mRNA 转录本和蛋白受到免疫刺激的诱导，CfRel 蛋白可以从 IκB 蛋白中释放出来，并转移到血细胞核中，调节扇贝的免疫反应。
