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基于转录组学技术对斑马鱼肝脏组织低氧胁迫影响的研究

宋汝浩 胡瑞芹 李根芳 张智聪 许强华

宋汝浩, 胡瑞芹, 李根芳, 张智聪, 许强华. 基于转录组学技术对斑马鱼肝脏组织低氧胁迫影响的研究[J]. 南方水产科学. doi: 10.12131/20220038
引用本文: 宋汝浩, 胡瑞芹, 李根芳, 张智聪, 许强华. 基于转录组学技术对斑马鱼肝脏组织低氧胁迫影响的研究[J]. 南方水产科学. doi: 10.12131/20220038
SONG Ruhao, HU Ruiqin, LI Genfang, ZHANG Zhicong, XU Qianghua. Research on effect of hypoxia stress on liver tissue of zebrafish (Danio rerio) based on transcriptomics technology[J]. South China Fisheries Science. doi: 10.12131/20220038
Citation: SONG Ruhao, HU Ruiqin, LI Genfang, ZHANG Zhicong, XU Qianghua. Research on effect of hypoxia stress on liver tissue of zebrafish (Danio rerio) based on transcriptomics technology[J]. South China Fisheries Science. doi: 10.12131/20220038

基于转录组学技术对斑马鱼肝脏组织低氧胁迫影响的研究

doi: 10.12131/20220038
基金项目: 国家重点研发计划项目 (2018YFD0900601, 2018YFC0310600);国家自然科学基金面上项目 (31772826);上海市教委重点科技创新项目 (2017-01-07-00-10-E00060)
详细信息
    作者简介:

    宋汝浩 (1991—),男,硕士研究生,研究方向为渔业资源。E-mail: songruhao2015@163.com

    通讯作者:

    胡瑞芹 (1989—),女,博士后,从事分子生物学研究。E-mail: rqhu@shou.edu.cn

    许强华 (1974—),女,教授,博士,从事分子生态与功能基因组学研究。E-mail: qhxu@shou.edu.cn

  • 中图分类号: S 917.4

Research on effect of hypoxia stress on liver tissue of zebrafish (Danio rerio) based on transcriptomics technology

  • 摘要: 为探究鱼类响应低氧胁迫的调控机制,将1月龄的野生型斑马鱼 (Danio rerio) 在1.5 mg·L−1的低氧浓度下胁迫2个月后,对其肝脏组织进行转录组测序比较分析。对常氧与低氧组的3 270个差异基因进行KEGG分析,主要富集于细胞增殖、脂质代谢、糖类代谢和氨基酸代谢等通路。其中,上调的1 864个基因主要与细胞增殖相关,下调的1 406个基因主要参与脂质代谢。对差异基因进行GO富集分析,发现铁离子束 (Iron ion banding) 功能差异显著。对铁代谢相关基因的表达量进行分析,发现铁离子储存相关基因fthl28和fthl31变化显著,提示在低氧胁迫下斑马鱼肝脏 (Zebrafish liver, ZFL) 组织中铁离子含量发生显著变化。利用ZFL细胞进行体外验证实验,将ZFL细胞进行0.1% (体积分数) O2低氧胁迫,发现随着胁迫时间的延长,ZFL细胞的成活率降低,且细胞中与铁代谢相关基因和铁蛋白 (Ferritin) 的表达均显著降低。综上所述,铁代谢调节是低氧胁迫下的重要响应过程,低氧会导致细胞内铁代谢紊乱,延长低氧时间会形成新的铁稳态。研究结果为探究鱼类的低氧适应机制提供了理论基础和参考。
  • 图  1  低氧胁迫下肝脏组织KEGG Pathway富集分布

    Figure  1.  Enrichment distribution of KEGG Pathway in liver tissue under hypoxia stress

    图  2  低氧胁迫肝脏组织差异基因KEGG Pathway富集分布

    注:a. 上调基因 KEGG Pathway 富集分布;b. 下调基因 KEGG Pathway 富集分布。

    Figure  2.  Enrichment and distribution of differential gene KEGG Pathway in liver tissue under hypoxia stress

    Note: a. Up-regulated gene KEGG Pathway enrichment distribution; b. Down-regulated gene KEGG Pathway enrichment distribution.

    图  3  低氧胁迫后肝脏GO富集分布

    Figure  3.  Distribution of GO enrichment in liver after hypoxia stress

    图  4  低氧胁迫后斑马鱼肝脏组织中与铁代谢相关基因的表达量分析

    注:a. 铁代谢相关基因的相对表达量热图;b. 5 个铁代谢相关基因的 RT-qPCR 和 RNA-seq 相对表达量的对比分析。

    Figure  4.  Analysis of expression levels of genes related to iron metabolism in zebrafish liver tissues after hypoxia stress

    Note: a. Heat map of relative expression of iron metabolism relative genes; b. Comparative analysis of relative expression of five iron metabolism related genes of RT-qPCR and RNA-seq.

    图  5  低氧胁迫ZFL细胞及细胞成活率、铁代谢相关基因与铁蛋白鉴定

    注:柱状图上方*表明与低氧组 (0.1% O2) 相比在 P<0.05 水平存在差异;**表示在 P<0.01 水平差异显著;***表示在P<0.001 水平差异极显著。a. 0.1% O2 胁迫 3 d 与常氧条件下 ZFL 细胞存活量对比;b. 0.1% O2 胁迫下 ZFL 细胞活性检测;c. 0.1% O2胁迫 3 d 与常氧条件下 ZFL 细胞中铁代谢相关基因表达量分析;d. 0.1% O2 胁迫后铁蛋白表达量与常氧下对比。

    Figure  5.  Identification of ZFL cells and cell survival rate, iron metabolism-related genes and Ferritin protein under hypoxia stress

    Note: *. Difference (P<0.05) compared with the hypoxia group (0.1% O2); **. Siginificant difference at P<0.01 level; ***. Very significant difference at P<0.001 level. a. Comparison of the viable cell mass of ZFL cells under 0.1% O2 stress for 3 d and normoxia condition; b. Detection of ZFL cell viability under 0.1% O2 stress; c. Analysis of the expression of genes related to iron metabolism in ZFL cells under 0.1% O2 stress for 3 d and normoxia condition; d. Comparison of Ferritin protein expression after 0.1% O2 stress and normoxia condition.

    表  1  引物信息

    Table  1.   Primer information

    引物名
    Primer name
    引物序列
    Primer sequences (5'–3')
    基因ID
    Gene ID
    tfa-F AGCAGCAGACATTGAGTGTC 30255
    tfa-R TTTGCTCCATCTACTGTAAC
    tfr2-F AGCAGTTTACCTCACACTGAC 494476
    tfr2-R AGGAATGTTGTCCGGCTCG
    fth27-F TGCGAGGCTTTGATCAACAAG 436651
    fth27-R TGGCAAATCCAGGAAGAGCC
    fth28-F AAGATGATCAATCTGGAGC 100006523
    fth28-R TTGAAGAACTTGGCAAATCC
    fth31-F AGGCTGCGATCAACAAGATG 553552
    fth31-R AGGAAGAGCCACATCGTC
    fpn-F ACATGCCCTCTCGACATGG 58153
    fpn-R AGGAGTAAACTATTGCCATACAG
    actin-F TGTCCCTGTATGCCTCTGGT 57934
    actin-R AAGTCCAGACGGAGGATG
    下载: 导出CSV
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出版历程
  • 收稿日期:  2022-02-21
  • 修回日期:  2022-04-30
  • 录用日期:  2022-06-15
  • 网络出版日期:  2022-07-29

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