关键词: 斑马鱼, F4 细胞, 低温, miRNA 

Abstract: Objective To investigate the expression and mechanism of miR-183-5p in zebrafish (Danio rerio) ZF4 cells during low temperature acclimatization. Methods ZF4 cells were cultured under 18℃ for 30 days. The expression of miR-183-5p in ZF4 cells was detected by RT-qPCR. Cell viability was determined by CCK-8 cell viability assay after transfection of miR-183-5p mimics/ miRNC with Turbofect cell transfection kit. Targetscan Fish database was used to predict the potential target genes, and dual luciferase activity reporting assay was used to confirm the binding site of miR-183-5p and its target gene egr1. Results The expression level of miR-183-5p increased significantly. The conservation analysis showed that miR-183-5p was highly conserved among species. Overexpression of miR-183-5p could enhance the viability of ZF4 cells under low temperature. The potential target genes of miR-183-5p were predicted by bioinformatics methods and tentatively verified by RT-qPCR. The dual luciferase activity reporting assay further confirmed that miR-183-5p could bind to the 3′UTR of egr1. Conclusion This study demonstrates that miR-183-5p plays a protective role in ZF4 cells during low temperature acclimatization through egr1 in zebrafish. It lays a foundation for further exploration of the mechanisms of zebrafish during low temperature acclimation and provides a new idea for the researches of molecular mechanisms of hypothermia treatment.

Key words: zebrafish, ZF4 cells, low temperature, miRNA