RESUMO
Triazophos is a highly toxic organophosphorus pesticide, causing acute toxicity to brain tissue, and neurotoxicity and embryotoxicity to animals. Therefore, triazophos is considered as a public health problem due to its acute hazard index. MicroRNAs (miRNAs), a class of endogenous noncoding RNAs, can regulate the expression of target gene(s) by mediating mRNA cleavage or translational repression in organisms exposed to environmental chemicals. We found that nup43 is targeted by miR-217, which was significantly regulated in adult zebrafish (Danio rerio) exposed to triazophos (phenyl-1,2,4-triazolyl-3-(o,o-diethyl thionophosphate)). The expression of nup43 in both mRNA and protein levels was downregulated in a dose-dependent manner upon stimulation with triazophos. A dual luciferase reporter assay demonstrated that miR-217 interacted with the 3'-untranslated regions (3'-UTR) of nup43. The expression of nup43 in both mRNA and protein level was reduced in ZF4 cells when transfected with an miR-217 mimic, but increased when transfected with an miR-217 inhibitor. As a result, nup43 is targeted by miR-217 upon triazophos exposure. We suggest that miR-217 could be a potential toxicological biomarker for triazophos.
RESUMO
MicroRNA (miRNA) can regulate the expression of its target gene by mediating mRNA cleavage or by translational repression at a post-transcriptional level. Usually, one miRNA may regulate many genes as its targets, while one gene may also be targeted by many miRNAs. We previously demonstrated that cyb561d2, whose protein product is involved in cell defense, and chemical stress, is targeted by miR-155 in adult zebrafish (Danio rerio) when exposed to fipronil (5-amino-1-[2,6-dichloro-4-(trifluoromethyl) phenyl]-4-[(trifluoromethyl) sulphinyl]-1H-pyrazole-3-carbonitrile). Microcosm Targets prediction showed that the cyb561d2 gene is also highly possibly targeted by miR-194a, miR-216b, miR-429, and miR-499. These interactions need to be further validated experimentally. In this study, we evaluated the effects of fipronil on miR-194a, miR-216b, miR-429, miR-499 and cyb561d2 in zebrafish and investigated whether these four miRNAs could regulate the expression of cyb561d2 in both mRNA and protein levels. The expression of cyb561d2 was upregulated in both mRNA and protein level in a dose-dependent manner upon stimulation of fipronil, and miR-216b and miR-499 were downregulated concurrently, whereas there was no significant changes were observed in the expression level of miR-194a and miR-429. The dual luciferase report assay demonstrated that miR-216b and miR-499 interacted with cyb561d2 3'-untranslated regions (3'-UTR), miR-194a and miR-429 did not stimulate degradation of cyb561d2 mRNA. The expression of cyb561d2 was reduced in both mRNA and protein level when ZF4 cells were transfected with miR-499 mimic, whereas expression level of both mRNA and protein was increased when endogenous miR-499 was inhibited by transfection with miR-499 inhibitor. Likewise, the mRNA and protein level of cyb561d2 was affected by treatment with the mimics and the inhibitor of miR-216b. In contrast, when ZF4 cells were transfected with a mimic of miR-194a or miR-429, the expression of cyb561d2 mRNA was not significantly changed. As a result, cyb561d2 is targeted by miR-155, miR-216b and miR-499 upon fipronil exposure, and miR-194a and miR-429 can not target cyb561d2. The expression pattern of these 3 miRNAs presents novel fipronil responses that could be used as a toxicological biomarker.
