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

项目摘要/摘要 芳香烃受体(AHR)是脊椎动物正常发育所必需的，由一种 多种化学品，包括常见污染物，如氯化二恶英、多环芳香族 碳氢化合物(PAHs)和共面多氯联苯。在脊椎动物中，接触到 在生命早期激活促肾上腺皮质激素释放激素与不良分娩结局和受损有关 神经发育；然而，AHR下游的信号事件和导致中毒的基因 人们对此的反应在很大程度上仍不得而知。我们的长期目标是破译保守的AHR介导的生物 通过发现和验证AHR中涉及的致病基因和信号机制来做出反应 受调控的发育和毒理过程。我们之前发现了一种新的和系统发育的 由多种环境相关的AHR诱导的保守的长非编码RNA(SlincR) 配基。SlincR在AHR诱导的软骨主要调节因子sox9b的抑制中是必需的 并在AHR介导的出血和软骨缺陷中起因果作用。目标是 该建议的目的是确定AHR调节的lncRNA的发育和毒理学作用， 斑马鱼中的SlincR。中心假设是slincR是一个关键的AHR下游信号因子 增强对各种外来化学物质的不良发育反应。我们的理由是 了解slincR如何发挥作用以及它与哪些其他因素相互作用将加深对 具有发现人类治疗AHR相关疾病靶点的潜力的AHR信号。我们 将通过进行功能丧失和功能获得的研究来验证我们的假设，以确定在体内的作用 SlincR在AHR调节的发育和毒理学过程中(目标1)。要了解SlincR 与AHR信号的结构-功能关系，我们将进行结构映射实验和 确定slincR相互作用蛋白和全基因组靶标(目标2)。我们将使用计算 将目标1和目标2的结果整合到构建AHR基因调控网络的方法， 在AHR调节的生物过程中建立对slincR的系统水平的理解(目标3)。这 这一贡献意义重大，因为它将揭示AHR激活对slincR表达的功能影响 为理解AHR依赖的细胞反应提供了一种新的范式。拟议的研究 是创新的，因为我们采用了环境健康科学领域的新尖端技术。 我们的建议还将有助于描述二恶英类AHR配体的一种新的不良结果途径(AOP) 它融合了包括分子结构、分子相互作用、共同作用在内的多个生物学水平 调控基因、信号网络、组织发育和整个动物的表型。这些研究将 作为lncRNA增强生物学和系统功能表征的模板 疾病。

PROJECT SUMMARY/ABSTRACT The aryl hydrocarbon receptor (AHR) is required for proper vertebrate development and is activated by a diverse group of chemicals including common pollutants such as chlorinated dioxins, polycyclic aromatic hydrocarbons (PAHs), and coplanar polychlorinated biphenyls. In vertebrates, exposure to chemicals that activate the AHR during early life stages is associated with adverse birth outcomes and impaired neurodevelopment; however, the signaling events downstream of AHR and the genes contributing to toxic responses remain largely unknown. Our long-term goal is to decipher conserved AHR-mediated biological responses via discovery and validation of the causal genes and signaling mechanisms involved in AHR regulated developmental and toxicological processes. We previously identified a novel and phylogenetically conserved long non-coding RNA (slincR) that is induced by multiple environmentally relevant AHR ligands. SlincR is required for the AHR-induced repression of sox9b, a master regulator of cartilage development, and plays a causal role in AHR-mediated hemorrhaging and cartilage defects. The objective of 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 factor potentiating adverse developmental responses to diverse xenobiotic chemicals. Our rationale is that understanding how slincR functions and what other factors it interacts with will deepen the understanding of AHR-signaling with the potential to uncover human therapeutic targets for AHR-related disease. We will test our hypothesis by performing loss- and gain-of-function studies to determine the in vivo role of slincR in AHR regulated developmental and toxicological processes (Aim 1). To understand the slincR structure-function relationship with AHR signaling, we will perform structure-mapping experiments and identify the slincR interacting proteins and genome-wide targets (Aim 2). We will use computational approaches 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). This contribution is significant as it will unravel the functional impact of AHR activation on slincR expression and provide a new paradigm for understanding AHR-dependent cellular responses. The proposed research is 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 ligands that incorporate multiple biological levels including molecular structure, molecular interactions, co- regulated genes, signaling networks, tissue development and whole animal phenotypes. These studies will serve as a template for the systematic functional characterization of lncRNA potentiating biology and disease.