| 引用本文: | 于金玉,梁健,梁爽,王晓宇,陈丽梅,李永仁,刘惠茹,郭永军.硬壳蛤MmDR2基因克隆及对溶藻弧菌的响应模式研究[J].海洋科学,2025,49(3):51-63. |
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| 硬壳蛤MmDR2基因克隆及对溶藻弧菌的响应模式研究 |
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于金玉1, 梁健1,2, 梁爽1,2, 王晓宇1,2, 陈丽梅1,2, 李永仁1,2, 刘惠茹1,2, 郭永军1,2
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1.天津农学院水产学院 天津市水产生态养殖重点实验室, 天津 300384;2.天津农学院 农业农村部智慧养殖重点实验室(省部共建), 天津 300384
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| 摘要: |
| 本研究探究多巴胺神经内分泌系统在调控硬壳蛤应对溶藻弧菌胁迫的分子机制。实验采用RACE技术克隆硬壳蛤多巴胺受体2(MmDR2)基因的全长cDNA序列, 采用实时定量PCR技术检测MmDR2基因在不同组织的表达模式, 溶藻弧菌攻毒和多巴胺刺激后, 在不同时间检测免疫和抗氧化相关酶活力及MmDR2基因的表达变化。基因克隆结果显示MmDR2基因全长1 464 bp, 共编码487个氨基酸, 具有1个DR2核心结构域, 且存在典型的多巴胺结合位点; 多序列比对结果显示MmDR2与斑马贻贝(Dreissena polymorpha)和砂海螂(Myaarenaria)的相似性较高, 分别为79.69 %和75.10 %; 系统发育进化树结果显示MmDR2与斑马贻贝和砂海螂的DR2聚为一支; 组织分布结果显示, MmDR2基因在各组织中均有表达, 其中在鳃、外套膜和血细胞中的表达量较高, 在性腺中表达量最低; 溶藻弧菌胁迫下, MmDR2在血细胞中的表达明显上升, 且在6 h达到峰值; 硬壳蛤感染溶藻弧菌, 会显著影响其消化腺中免疫和抗氧化相关酶的活性, 其中免疫相关酶活(ACP、AKP和LZM)首先被激活, 之后逐步恢复至正常水平, 抗氧化相关酶活则呈现波动, 总体在感染初期被显著抑制; 而当多巴胺刺激硬壳蛤后, 再对其进行溶藻弧菌攻毒, 其体内免疫和抗氧化相关酶活性总体表现为抑制状态。研究结果表明, MmDR2在硬壳蛤的抗感染免疫中发挥重要生物学作用。 |
| 关键词: 硬壳蛤 多巴胺受体2 溶藻弧菌 |
| DOI：10.11759/hykx20241117001 |
| 分类号:S-3 |
| 基金项目:天津市教委科研计划项目(2022KJ020) |
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| Cloning of the MmDR2 gene in Mercenaria mercenaria and its response pattern to Vibrio alginolyticus |
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YU Jinyu1, LIANG Jian1,2, LIANG Shuang1,2, WANG Xiaoyu1,2, CHEN Limei1,2, LI Yongren1,2, LIU Huiru1,2, GUO Yongjun1,2
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1.Key Laboratory of Aquatic Ecological Farming of Tianjin, College of Fisheries, Tianjin Agricultural University, Tianjin 300384, China;2.Key Laboratory of Smart Farming, Ministry of Agriculture and Rural Affairs (Provincial and Ministerial Co-construction), Tianjin Agricultural University, Tianjin 300384, China
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| Abstract: |
| This study investigated the molecular mechanisms by which the dopamine neuroendocrine system in Mercenaria mercenaria regulates the response to stress induced by Vibrio alginolyticus. We employed RACE technology to clone the full-length cDNA sequence of the dopamine receptor 2 (MmDR2) gene in the hard clam. Real-time quantitative PCR was employed to detect the expression patterns of MmDR2 in different tissues. Following a challenge with V. alginolyticus and dopamine stimulation, changes in the immune and antioxidant-related enzyme activity, as well as the expression of MmDR2, were measured at different time points. Results of the gene cloning revealed that MmDR2 is 1 464 bp in length, encoding 487 amino acids, and contains one DR2 core domain with a typical dopamine binding site. The multiple sequence alignment results showed that MmDR2 has high similarity with Dreissena polymorpha and Mya arenaria, at 79.69% and 75.10%, respectively. The phylogenetic tree results showed that MmDR2 and DR2 of D. polymorpha and M. arenaria clustered into one branch. The tissue distribution results demonstrated that the MmDR2 gene was expressed in all tissues, with higher expression levels in the gills, mantle, and hemocytes, and the lowest expression was found in the gonads. Under V. alginolyticus stress, the expression of MmDR2 in hemocytes significantly increased, peaking at 6 h. V. alginolyticus infection of M. mercenaria significantly affected the immune and antioxidant-related enzyme activity in the digestive glands. Among these, the immune-related enzymes ACP, AKP, and LZM were initially activated and then gradually returned to normal levels, while the activities of antioxidant-related enzymes fluctuated; significant inhibition was revealed during the early stages of infection. However, when M. mercenaria was stimulated with dopamine and subsequently challenged with V. alginolyticus, the overall immune and antioxidant-related enzyme activities in their bodies were suppressed. These results indicate that MmDR2 plays an important biological role in the anti-infection immunity of M. mercenaria. |
| Key words: Mercenaria mercenaria dopamine receptor 2 Vibrio alginolyticus |
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