摘要
为了明确miR-21参与介导嗜水气单胞菌感染鲫皮肤免疫相关信号通路的分子机制,通过筛选鉴定miR-21及其靶基因富集免疫相关信号通路分析,利用实时荧光定量PCR(qRT-PCR)检测组织分布与时序表达水平。结果表明,miR-21与BCL2靶基因预测的目标结合位点序列在人类、小鼠、鱼类等不同物种之间高度保守。qRT-PCR检测显示miR-21主要在鲫鳃、肠道和肌肉组织中相对高表达,在嗜水气单胞菌感染鲫皮肤中,miR-21呈现短期波动时序表达模式;miR-21调控靶基因BCL2介导TLR/MAPK信号通路相关基因(如IRAK1、TAB1、IKKa)在鲫皮肤中以下调表达变化为主,而TLR4和RHO基因在感染12 h和72 h出现显著上调表达,并且BCL2与TAB1基因表达有显著相关性(r=0.87)。以上研究结果,有助于理解鱼类miR-21与靶基因功能及其介导皮肤免疫的分子调控机制。
In order to clarify the molecular mechanism of miR-21 involvement in mediating immune related signaling pathways in Carassius auratus skin infected with Aeromonas hydrophila,miR-21 and its target genes were identified through screening and enriched immune related signaling pathway analysis.Real time fluorescence quantitative PCR(qRT PCR)was used to detect tissue distribution and temporal expression levels.The results indicate that the predicted binding site sequences of miR-21 and BCL2 target genes are highly conserved among different species such as humans,mice,and fish.QRT PCR detection showed that miR-21 was mainly highly expressed in the gills,intestines,and muscle tissues of Carassius auratus,in Carassius auratus skin infected with Aeromonas hydrophila,miR-21 exhibited a short-term fluctuating temporal expression pattern;miR-21 regulates the target gene BCL2 to mediate the downregulatied of TLR/MAPK signaling pathway related genes(such as IRAK1,TAB1,IKKa)in the Carassius auratus skin,while TLR4 and RHO genes show significant upregulation at 12 and 72 hours of infection,and BCL2 is significantly correlated with TAB1 gene expression(r=0.87).The above research results contribute to the understanding of the molecular regulatory mechanisms of miR-21 and target gene functions in fish,as well as its mediation of skin immunity.
作者
金歌
霍佳昕
林轩
陈奕芯
周君敏
胡秀彩
吕爱军
JIN Ge;HUO Jiaxin;LIN Xuan;CHEN Yixin;ZHOU Junmin;HU Xiucai;LV Aijun(Aquatic College,Tianjin Agricultural University,Tianjin Key Laboratory of Aquatic ecology and Breeding,Tianjin 300392,China)
基金
天津市教委科研计划自然科学基金重点项目(2019ZD14)。
