Mechanism Of Circular RNAs Regulating Graphene Oxide In Vivo Toxicity In Caenorhabditis Elegans

Graphene oxide?GO?,as a kind of engineering nanomaterials,shows great promise in many fields due to its unique physical and chemical properties.At the same time,people pay more attention to the GO toxicity to human and environmental animals.It is very necessary to study the toxicity mechanism and safety assessment of GO exposure.The model animal Caenorhabditis elegans has been widely used in ecotoxicological research because of its unique advantages.It has gradually established a multi-index analysis system which continuously improved the research technology by improving the sensitivity and detection speed of ecotoxicity.The mechanism of the toxicity effect of GO in nematodes was studied in previous work from the levels of mRNA,miRNA and lncRNA.circRNAs are a class of RNA molecules which were circular structure by special splicing.circRNAs can regulate biological processes in many ways,but its role in regulating of response to engineering nanomaterials is still unclear.In this study,we employed the model animal of C.elegans as the in vivo assay system to examine the roles and the functions of circRNAs in the regulation of response of animals to GO,a carbon-based nanomaterial.Dysregulated circRNAs in GO exposed nematodes were screened and identified by high-throughput sequencing combined with qRT-PCR assay.After 100?g/L GO exposure at environmental relavant concentration,we further screened out 10 dysregulated circRNAs including 8 down-regulated circRNAs and 2 up-regulated circRNAs.Using RNA interference?RNAi?technique,we found that the decrease in circ₀000201 or circ₀000247 expression might mediate the induction of GO toxicity,whereas the reduction in circ₀000115 expression might mediate a protection mechanism against GO toxicity in GO exposed nematodes.In addition,the effects of GO toxicity on circ₀000201 and circ₀000247 interference nematodes could also be detected at the environmental relavant concentration of 10?g/L GO.This further indicates circRNAs have the importamt roles in the safety assessment of GO exposure.We further elucidate the molecular mechanism of circ₀000115 in regulating GO toxicity.The results showed that circ₀000115 played a role in regulating GO toxicity in the intestine of nematodes by binding to IFC-2 protein and then enhanced GO toxicity by suppressing downstream gene daf-16,a key gene in Insulin/IGF-1 signaling pathway.In addition,circ₀000115 also regulated GO toxicity in neurons by affecting the expression of jnk-1 and mek-1 in JNK signaling pathway.In this study,circRNAs-induced toxicity of GO in vivo was screened from the whole genome and its biological function was identified.The molecular mechanism of circ₀000115 in regulating the in vivo toxicity of GO in nematodes was further discussed.It provides a reference and basis for further study on the regulatory function and mechanism of circRNAs in vivo toxicity of engineering nanomaterials and other biological processes.