宿主内输蛋白β1促进登革病毒非结构蛋白NS1分泌机制的研究

登革病毒非结构蛋白1（NS1）能够分泌至胞外促使血管渗透性增加导致血管渗漏，引发重症登革热，是决定登革病毒致病性的关键性因素，但是NS1分泌的分子机制尚不明确。申请人前期实验发现，宿主核转运蛋白importin β1能够与NS1相互作用，沉默该蛋白能够特异性地降低NS1的分泌，但其中机理有待于进一步探究。本项目拟1）通过importin β1拯救实验和NS1-importin β1免疫共沉淀、生物膜干扰实验以及利用importin β1抑制剂进一步验证这一表型；2）通过研究importin β1分子结构对NS1分泌和importin β1对亚细胞定位的影响，以及importin β1调节外泌体和脂质代谢等角度，深入解析importin β1调控NS1分泌的分子机理。该研究将为解析登革热的发病机制以及研发治疗重症登革热的靶向药物提供重要的理论支撑。

Dengue virus non-structural protein 1 (NS1) can be secreted from cells causing vascular permeability and leakage, which leads to severe Dengue. The molecular mechanism of NS1 secretion is still unclear. Our previous experiments showed that host nuclear transport protein importin β1 interacts with NS1 and knockdown of importin β1 significantly reduced the secretion of NS1, but the mechanism needs to be further elucidated. Therefore, the purpose of this project is to 1) further validate this phenotype by performing the importin β1 rescue experiment, NS1-importin β1 co-immunoprecipitation, biolayer interferometry assay, and the importin β1 inhibition experiments, 2)study how the molecular structure of importin β1 affect NS1 secretion, how importin β1 affects NS1 subcellular localization, and how importin β1 facilitate NS1 secretion by regulating exosomes secretion and lipid metabolism. The knowledge gained in this project will facilitate our understanding of severe dengue fever-related pathogenesis and the development of novel antiviralsagainst Dengue infection.