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母尹楠, 李婉茹, 何亮华, 陈炯, 陈新华. 大黄鱼HIF-1α基因的克隆鉴定与表达分析[J]. 水生生物学报, 2020, 44(2): 296-302. DOI: 10.7541/2020.036
引用本文: 母尹楠, 李婉茹, 何亮华, 陈炯, 陈新华. 大黄鱼HIF-1α基因的克隆鉴定与表达分析[J]. 水生生物学报, 2020, 44(2): 296-302. DOI: 10.7541/2020.036
MU Yin-Nan, LI Wan-Ru, HE Liang-Hua, CHEN Jiong, CHEN Xin-Hua. MOLECULAR CHARACTERIZATION AND EXPRESSION ANALYSIS OF HIF- IN LARGE YELLOW CROAKER (LARIMICHTHYS CROCEA)[J]. ACTA HYDROBIOLOGICA SINICA, 2020, 44(2): 296-302. DOI: 10.7541/2020.036
Citation: MU Yin-Nan, LI Wan-Ru, HE Liang-Hua, CHEN Jiong, CHEN Xin-Hua. MOLECULAR CHARACTERIZATION AND EXPRESSION ANALYSIS OF HIF- IN LARGE YELLOW CROAKER (LARIMICHTHYS CROCEA)[J]. ACTA HYDROBIOLOGICA SINICA, 2020, 44(2): 296-302. DOI: 10.7541/2020.036

大黄鱼HIF-1α基因的克隆鉴定与表达分析

MOLECULAR CHARACTERIZATION AND EXPRESSION ANALYSIS OF HIF- IN LARGE YELLOW CROAKER (LARIMICHTHYS CROCEA)

  • 摘要: 为研究低氧诱导因子1α(Hypoxia-inducible factor-1α, HIF-1α)在鱼类免疫应答中的作用, 研究克隆得到了大黄鱼(Larimichthys crocea)HIF-1α基因(LcHIF-1α)的cDNA序列, 其开放阅读框全长2256个核苷酸, 编码一个751个氨基酸的蛋白。通过氨基酸序列比对发现, LcHIF-1α蛋白具有5个典型的功能结构域, 包括1个helix-loop-helix(HLH)结构域, 2个PAS结构域, 1个DNA结合结构域(HIF-1)和1个羧基端反式激活结构域(HIF-1a_CTAD)。系统进化分析显示, LcHIF-1α和其他硬骨鱼类HIF-1α聚成一支, 与两栖类、鸟类和哺乳类HIF-1α的亲缘关系相对较远。序列分析结果表明, LcHIF-1α具有保守的特征结构域, 预示着其功能可能与哺乳动物HIF-1α类似。荧光定量PCR结果显示, LcHIF-1α在所检测的健康大黄鱼各个组织中都有表达, 在溶藻弧菌(Vibrio alginolyticus)感染后, 大黄鱼脾脏和头肾组织中的LcHIF-1α表达水平显著上调, 达到峰值时分别上升了6.8倍和2.9倍。此外, LcHIF-1α在大黄鱼的3种免疫细胞(中性粒细胞、巨噬细胞和淋巴细胞)中也都有表达, 在中性粒细胞和巨噬细胞中的表达量相对较高; 在脂多糖(LPS)诱导后, 中性粒细胞和巨噬细胞中LcHIF-1α的表达量显著增加, 峰值分别是对照组的2.6倍和1.8倍, 这些结果表明LcHIF-1α可能参与调控大黄鱼抗细菌免疫应答。

     

    Abstract: Hypoxia-inducible factor-1α (HIF-1α) is considered as the master regulator of the response to hypoxia, through activating target gene expression involved in angiogenesis, erythropoiesis and glycolysis. In recent years, HIF-1α has been found to regulate the immune response. However, very little information was available about its functions in fish immune response. In this study, the cDNA sequence of HIF-1α gene (LcHIF-1α) was cloned from large yellow croaker Larimichthys crocea. Its open reading frame (ORF) was 2256 nucleotides long, encoding a peptide of 751 amino acids. The deduced LcHIF-1α protein contained five typical motifs: a helix-loop-helix (HLH) domain, two PAS domains, a DNA binding domain (HIF-1), and a HIF-1α C-terminal transactivation domain (HIF-1a_CTAD). Phylogenetic analysis showed LcHIF-1α formed a separate clade with sequences from other fish species, far from the clades formed by avian, amphibian and mammalian HIF-1α proteins. The LcHIF-1α transcripts were constitutively expressed in all examined tissues and significantly up-regulated in the spleen and head kidney tissues by Vibrio alginolyticus, and reached the peak at 24h, with 6.8- and 2.9-fold increases, respectively. Its transcripts were also detected in neutrophils, macrophages and lymphocytes from large yellow croaker, and significantly up-regulated by lipopolysaccharide (LPS) in neutrophils and macrophages with 2.6- and 1.8-fold increases at the peak. These results indicate that LcHIF-1α may play a role in mediating the immune response of large yellow croaker.

     

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