Role of glucocorticoid signalling in stress adaptation

发展适应策略以应对广泛的应激源是生物体整体健康的基础。在脊椎动物中协调应激反应的关键分子是皮质类固醇，它们的作用可由两个受体介导，即糖皮质激素受体(GR)和盐皮质激素受体(MR)。这些皮质类固醇受体在进化上是保守的，在脊椎动物中普遍表达。我的研究计划旨在了解糖皮质激素的作用机制，以及与应激相关的糖皮质激素信号在动物适应应激中的后果。尽管GR的作用被很好地描述，但对MR在鱼类中的生理作用几乎一无所知。因此，我的主要假设是，MR信号对应激轴激活是必不可少的，并且在调节高度保守的糖皮质激素介导的应激适应方面可以与GR互补地发挥作用。在这个为期5年的计划中，我的目标是专门探讨MR在感知和激活应激反应以及调节应激相关行为中的进化保守作用。我的实验室最近使用CRISPR/Cas9突变技术在斑马鱼中产生了GR和MR基因敲除突变。利用这些突变体，我们将进行功能丧失研究，以充分表征这些受体在调节应激应对机制中的作用。我们还在斑马鱼中产生了双重GR，MR基因敲除(MGR)，这些MGR突变与我现有的GR和MR突变相结合，将使我的实验室能够全面确定皮质类固醇受体在介导糖皮质激素相关应激反应中的保守作用。该方法将包括对不同发育阶段的斑马鱼(野生型和突变型)进行整体动物研究，以及组织特异性研究，包括肝脏、鳃、心脏、头肾、骨骼肌和脑。这些研究将确定皮质类固醇受体信号在调节与应激相关的能量底物重新分配中的作用。这些研究的结果将为压力应对机制提供新的见解，特别是MR信号在调节糖皮质激素在恢复体内平衡中的作用。此外，我们开发的新工具，包括受体基因敲除，将为培训和合作提供理想的环境，并将促进参与压力适应的新途径的发现。这项工作的结果将对鱼类应激研究的所有领域产生影响，包括水产养殖和环境影响监测。它还将在生物医学领域具有翻译潜力，因为哺乳动物中GR或MR的敲除都是致命的。最后，这一建议和所产生的结果将极大地影响我们对糖皮质激素在调节应激适应中的进化保守作用的理解。

The development of adaptive strategies to cope with a wide spectrum of stressors is fundamental to the overall fitness of an organism. Key players that orchestrate the stress response in vertebrates are corticosteroids, and their action can be mediated by two receptors, the glucocorticoid receptor (GR) and the mineralocorticoid receptor (MR). These corticosteroid receptors are evolutionarily conserved and are ubiquitously expressed in vertebrates. My research program aims to understand the mechanisms of glucocorticoid action, and the consequences of stress-related glucocorticoid signalling in adapting animals to stress. Despite the well characterized action of GR, almost nothing is known about the physiological role of MR in fish. Therefore, my overarching hypothesis is that MR signalling is essential for stress axis activation and can operate in a complementary fashion with GR in regulating the highly conserved glucocorticoid-mediated stress adaptation. In this 5-year proposal, my goal is to specifically explore the evolutionarily conserved role of MR in the perception and activation of the stress response, as well as the regulation of stress-related behaviour. My lab recently generated the GR and MR knockout mutants in zebrafish using CRISPR/Cas9 mutagenesis. Using these mutants, we will carry out loss of function studies to fully characterize the role of these receptors in modulating the stress coping mechanisms. We have also generated double GR,MR knockouts (MGR) in zebrafish, and these MGR mutants, in combination with my existing GR and MR mutants, will allow my lab to comprehensively determine the conserved action of corticosteroid receptors in mediating the glucocorticoid-related stress responses. The methodology will involve whole animal studies with zebrafish (wildtype and mutants) at various developmental stages, as well as tissue-specific studies, including liver, gill, heart, head kidney, skeletal muscle, and brain. The studies will determine the contribution of the corticosteroid receptor signalling in regulating the energy substrate re-partitioning associated with stress. The results from these studies will provide novel insights on stress coping mechanisms, and specifically the role of MR signalling in mediating the glucocorticoid effect in restoring homeostasis. Furthermore, the new tools, including receptor knockouts, we have generated will provide an ideal environment for training and collaboration, and will facilitate discoveries of novel pathways involved in stress adaptation. The results from this work will have an impact on all areas of fish stress research, including aquaculture and environmental effects monitoring. It will also have translational potential in the biomedical field, as knockout of either GR or MR in mammals is lethal. Finally, this proposal and the results generated will greatly impact our understanding of the evolutionary conserved role of glucocorticoids in mediating stress adaptation.