基于核受体分子构象扰动的新型内分泌干扰物作用机制研究

Currently, large quantities of new type chemicals have been widely used, however, their endocrine disrupting effects and the underlying molecular mechanism still remain largely unknown. In this project, the endocrine disrupting effects of many emerging contaminants will be screened by the the recombinant yeast two-hybrid, dual-luciferase reporter gene assay, zebrafish embryo and mouse in vivo assay. From the perspective of molecular initiating events of nuclear receptors (NR), the combination of large-scale molecular dynamics simulations, NR biosensors, fluorescence spectroscopy, fluorescence lifetime and circular dichroism was planned to investigate the conformational disruption of NR ligand binding domain (LBD) monomer and dimer induced by contaminants. The binding mode and binding potency will be probed. The entry/exit pathways of contaminants from NR LBD and their effect on conformations of involved critical residues will be investigated by random accelerated molecular dynamics simulations and steered molecular dynamics simulations. We aim to reveal the structure-activity relationship on the endocrine disrupting effect of various contaminants and will elucidate the molecular mechanism at molecular level. The obtained results will provide essential scientific data for human health risk assessment of emerging pollutants.

目前大量新型化学品被逐渐广泛使用，然而对其内分泌干扰效应缺乏了解，并且其内分泌干扰效应分子机制尚不明晰。本项目利用重组基因酵母双杂交、双荧光素酶报告基因、斑马鱼胚胎及小鼠试验方法甄别多类新型污染物的潜在内分泌干扰效应。从核受体分子起始事件角度出发，采用大尺度分子动力学模拟并整合生物传感器、荧光光谱、荧光寿命及圆二色谱等研究手段系统考察污染物对多种核受体配体结合域以及核受体二聚体的构象扰乱程度，分析污染物与核受体的结合模式和结合能力。进一步利用随机加速动力学模拟和拉伸动力学模拟考察污染物进出核受体结合位点的关键通道及对重要氨基酸的构象影响。期望本项目在分子、基因、细胞和活体层次上的研究能揭示新型污染物内分泌干扰效应的构效关系，在分子水平诠释干扰效应分子机制，为新型污染物的人体健康风险评估提供重要科学数据支持。

内分泌干扰物是一类重要的新污染物，其如何干扰核受体以及核受体下游相关通路进而引起有害健康效应的分子机制尚不明晰。本项目主要研究了双酚A类似物、二苯甲酮紫外防晒剂、新型溴苯阻燃剂、叔丁基苯酚等新污染物对雌激素受体（Estrogen Receptor, ER）β、甲状腺激素受体 (Thyroid Hormone Receptor, TR) β、雄激素受体（Androgen Receptor, AR）、芳香烃受体(Aryl Hydrocarbon Receptor, AhR)、视黄酸受体(Retinoic Acid Receptor, RAR）β等多种核受体配体结合域（Ligand Binding Domain, LBD）及其二聚体的构象扰乱，表征了相关的分子起始事件及健康风险；BP-1、BP-8等典型二苯甲酮紫外防晒剂干扰ERβ通路后异常激活ERα与Wnt/β-catenin通路的交互作用，导致卵巢癌SKOV3细胞发生增殖、迁移和侵袭等异常生物学行为；BPA和BPS通过干扰ER-PI3K-eNOS通路促进小鼠体内外血管新生，而TBBPA、TBBPS通过干扰TR-PI3K-eNOS通路而抑制小鼠体内外血管新生；2,6-二叔丁基苯酚及其代谢产物抑制RARβ受体异常激活Erk1/2-COX2通路，促进肝癌细胞H4IIEs增殖、迁移、侵袭；2-巯基苯并噻唑通过激活AhR受体，促进膀胱癌T24细胞促T24细胞侵袭效应；基于机器学习方法构建了针对AhR受体、PXR受体干扰物质的高通量筛选模型。相关研究诠释各类污染物内分泌干扰效应的分子作用机制，揭示了相关健康风险，服务于我国“十四五”新污染物治理。本项目发表SCI收录论文17篇，其中Environ. Sci. Technol.杂志7篇，全国性学术会议大会报告1次、口头报告3次。