Dysregulation of Neuronal Gαo Signaling by Graphene Oxide in Nematode Caenorhabditis elegans.

Exposure to graphene oxide (GO) induced some dysregulated microRNAs (miRNAs), such as the increase in mir-247, in nematode Caenorhabditis elegans. We here further identified goa-1 encoding a Gαo and pkc-1 encoding a serine/threonine protein kinase as the targets of neuronal mir-247 in the regulation of GO toxicity. GO exposure increased the expressions of both GOA-1 and PKC-1. Mutation of goa-1 or pkc-1 induced a susceptibility to GO toxicity, and suppressed the resistance of mir-247 mutant to GO toxicity. GOA-1 and PKC-1 could also act in the neurons to regulate the GO toxicity, and neuronal overexpression of mir-247 could not affect the resistance of nematodes overexpressing neuronal goa-1 or pkc-1 lacking 3'-UTR to GO toxicity. In the neurons, GOA-1 acted upstream of diacylglycerol kinase/DGK-1 and PKC-1 to regulate the GO toxicity. Moreover, DGK-1 and GOA-1 functioned synergistically in the regulation of GO toxicity. Our results highlight the crucial role of neuronal Gαo signaling in response to GO in nematodes.

Functional disruption in epidermal barrier enhances toxicity and accumulation of graphene oxide.

In Caenorhabditis elegans, mutation of mlt-7 causes the deficits in epidermal barrier. Using the nematodes with epidermal-specific RNA interference (RNAi) knockdown of mlt-7 as a genetic tool, we found that epidermal-specific RNAi knockdown of mlt-7 resulted in a susceptibility to graphene oxide (GO) toxicity, and enhanced GO accumulation in the body. Epidermal-development related proteins of BLI-1 and IFB-1 acted as downstream targets of MLT-7, and mediated the function of MLT-7 in maintaining the epidermal barrier. Antimicrobial proteins of NLP-30 and CNC-2 also acted as downstream targets of MLT-7 in the regulation of GO toxicity. Epidermal-specific RNAi knockdown of nlp-30 or cnc-2 enhanced GO toxicity and accumulation in bli-1(RNAi) or ifb-1(RNAi) nematodes. Our data highlights the importance of maintaining normal epidermal barrier for nematodes against the GO toxicity.

Developmental basis for intestinal barrier against the toxicity of graphene oxide.

BACKGROUND: Intestinal barrier is crucial for animals against translocation of engineered nanomaterials (ENMs) into secondary targeted organs. However, the molecular mechanisms for the role of intestinal barrier against ENMs toxicity are still largely unclear. The intestine of Caenorhabditis elegans is a powerful in vivo experimental system for the study on intestinal function. In this study, we investigated the molecular basis for intestinal barrier against toxicity and translocation of graphene oxide (GO) using C. elegans as a model animal. RESULTS: Based on the genetic screen of genes required for the control of intestinal development at different aspects using intestine-specific RNA interference (RNAi) technique, we identified four genes (erm-1, pkc-3, hmp-2 and act-5) required for the function of intestinal barrier against GO toxicity. Under normal conditions, mutation of any of these genes altered the intestinal permeability. With the focus on PKC-3, an atypical protein kinase C, we identified an intestinal signaling cascade of PKC-3-SEC-8-WTS-1, which implies that PKC-3 might regulate intestinal permeability and GO toxicity by affecting the function of SEC-8-mediated exocyst complex and the role of WTS-1 in maintaining integrity of apical intestinal membrane. ISP-1 and SOD-3, two proteins required for the control of oxidative stress, were also identified as downstream targets for PKC-3, and functioned in parallel with WTS-1 in the regulation of GO toxicity. CONCLUSIONS: Using C. elegans as an in vivo assay system, we found that several developmental genes required for the control of intestinal development regulated both the intestinal permeability and the GO toxicity. With the focus on PKC-3, we raised two intestinal signaling cascades, PKC-3-SEC-8-WTS-1 and PKC-3-ISP-1/SOD-3. Our results will strengthen our understanding the molecular basis for developmental machinery of intestinal barrier against GO toxicity and translocation in animals.

Long-term exposure to thiolated graphene oxide in the range of µg/L induces toxicity in nematode Caenorhabditis elegans.

The in vivo toxicity and translocation of thiolated graphene oxide (GO-SH) are still largely unclear. We hypothesized that long-term exposure to GO-SH may cause the adverse effects on environmental organisms. We here employed in vivo assay system of Caenorhabditis elegans to investigate the possible toxicity and translocation of GO-SH after long-term exposure. In wild-type nematodes, we observed that prolonged exposure to GO-SH at concentrations>100µg/L resulted in the toxicity on functions of both primary targeted organs such as the intestine and secondary targeted organs such as the neurons and the reproductive organs. The severe accumulation of GO-SH was further detected in the body of wild-type nematodes. The translocation of GO-SH into secondary targeted organs such as reproductive organs through intestinal barrier might be associated with the enhancement in intestinal permeability in GO-SH exposed wild-type nematodes. Prolonged exposure to GO-SH (100µg/L) decreased the expression of gas-1 encoding a subunit of mitochondrial complex I, and mutation of gas-1 caused the formation of GO-SH toxicity at concentration>10µg/L and more severe accumulation of GO-SH in the body of animals. Therefore, our results confirm the possibility for prolonged exposure to GO-SH in inducing adverse effects on nematodes. Our data highlight the potential adverse effects of GO-SH in the range of µg/L on environmental organisms after long-term exposure.