Defining the developmental and toxicological roles of the AHR-regulated Sox9 lncRNA in zebrafish

PROJECT SUMMARY/ABSTRACTThe aryl hydrocarbon receptor (AHR) is required for proper vertebrate development and is activated by adiverse group of chemicals including common pollutants such as chlorinated dioxins, polycyclic aromatichydrocarbons (PAHs), and coplanar polychlorinated biphenyls. In vertebrates, exposure to chemicals thatactivate the AHR during early life stages is associated with adverse birth outcomes and impairedneurodevelopment; however, the signaling events downstream of AHR and the genes contributing to toxicresponses remain largely unknown. Our long-term goal is to decipher conserved AHR-mediated biologicalresponses via discovery and validation of the causal genes and signaling mechanisms involved in AHRregulated developmental and toxicological processes. We previously identified a novel and phylogeneticallyconserved long non-coding RNA (slincR) that is induced by multiple environmentally relevant AHRligands. SlincR is required for the AHR-induced repression of sox9b, a master regulator of cartilagedevelopment, and plays a causal role in AHR-mediated hemorrhaging and cartilage defects. The objectiveof this proposal is to identify the developmental and toxicological roles of the AHR-regulated lncRNA,slincR, in zebrafish. The central hypothesis is that slincR is a critical AHR-downstream signaling factorpotentiating adverse developmental responses to diverse xenobiotic chemicals. Our rationale is thatunderstanding how slincR functions and what other factors it interacts with will deepen the understandingof AHR-signaling with the potential to uncover human therapeutic targets for AHR-related disease. Wewill test our hypothesis by performing loss- and gain-of-function studies to determine the in vivo role ofslincR in AHR regulated developmental and toxicological processes (Aim 1). To understand the slincRstructure-function relationship with AHR signaling, we will perform structure-mapping experiments andidentify the slincR interacting proteins and genome-wide targets (Aim 2). We will use computationalapproaches to integrate the findings of Aims 1 & 2 into the construction of an AHR gene regulatory network,establishing a systems-level understanding of slincR in AHR-regulated biological processes (Aim 3). Thiscontribution is significant as it will unravel the functional impact of AHR activation on slincR expressionand provide a new paradigm for understanding AHR-dependent cellular responses. The proposed researchis innovative as we employ cutting-edge technologies new to the field of environmental health sciences.Our proposal will also help to describe a novel adverse outcome pathway (AOP) for dioxin-like AHR ligandsthat incorporate multiple biological levels including molecular structure, molecular interactions, co-regulated genes, signaling networks, tissue development and whole animal phenotypes. These studies willserve as a template for the systematic functional characterization of lncRNA potentiating biology anddisease.