Signaling Events Downstream of AHR Activation That Contribute to Toxic Responses: The Functional Role of an AHR-Dependent Long Noncoding RNA () Using the Zebrafish Model.

BACKGROUND: A structurally diverse group of chemicals, including dioxins [e.g., 2,3,7,8-tetrachlorodibenzo--dioxin (TCDD)] and polycyclic aromatic hydrocarbons (PAHs), can xenobiotically activate the aryl hydrocarbon receptor (AHR) and contribute to adverse health effects in humans and wildlife. In the zebrafish model, repression of has a causal role in several AHR-mediated toxic responses, including craniofacial cartilage malformations; however, the mechanism of repression remains unknown. We previously identified a long noncoding RNA, long intergenic noncoding RNA (), which is increased (in an AHR-dependent manner) by multiple AHR ligands and is required for the AHR-activated repression of .

OBJECTIVE: Using the zebrafish model, we aimed to enhance our understanding of the signaling events downstream of AHR activation that contribute to toxic responses by identifying: ) whether is enriched on the locus, ) 's functional contributions to TCDD-induced toxicity, ) PAHs that increase expression, and ) mammalian orthologs of .

METHODS: We used capture hybridization analysis of RNA targets (CHART), qRT-PCR, RNA sequencing, morphometric analysis of cartilage structures, and hemorrhaging screens.

RESULTS: The transcript was enriched at the 5' untranslated region (UTR) of the locus. Transcriptome profiling and human ortholog analyses identified processes related to skeletal and cartilage development unique to TCDD-exposed controls, and angiogenesis and vasculature development unique to TCDD-exposed zebrafish that were injected with a splice-blocking morpholino targeting . In comparison to TCDD exposed control morphants, morphants exposed to TCDD resulted in abnormal cartilage structures and a smaller percentage of animals displaying the hemorrhaging phenotype. In addition, expression was significantly increased in six out of the sixteen PAHs we screened.

CONCLUSION: Our study establishes that in zebrafish, is recruited to the 5' UTR to repress transcription, can regulate cartilage development, has a causal role in the TCDD-induced hemorrhaging phenotype, and is up-regulated by multiple environmentally relevant PAHs. These findings have important implications for understanding the ligand-specific mechanisms of AHR-mediated toxicity. https://doi.org/10.1289/EHP3281.