F1Fo-ATP合酶δ亚基在白念珠菌致死性感染中的作用及其机制研究

白念珠菌是重要的致死性真菌，其复杂的致病因子主要靠ATP依赖Ras1-cAMP-PKA通路（“通路”）调控。线粒体ATP合成的终端酶是F1Fo-ATP合酶，其δ亚基功能未知。我们发现：δ亚基缺失后，并未影响细胞内ATP水平，却无法造成小鼠致死；Ras1活性、Cyr1显著抑制；Cdc19与δ亚基互作，生信分析Cdc19-Cyr1-Ras1互作；Cdc19别构激活剂FBP明显降低。据此我们提出，δ亚基并不影响ATP合成、却通过FBP/和Cdc19调控“通路”影响致死性感染。本项目拟以体内外实验确认δ亚基是白念珠菌致死性感染的关键，再以代谢组学、生化手段证明δ亚基不影响各环节ATP合成，以蛋白互作、代谢物蛋白互作等证明δ亚基通过FBP/和Cdc19调控“通路”，以SBVS寻找靶向小分子化合物。项目将从F1Fo-ATP合酶δ亚基这一新视角，揭示白念珠菌致死性感染的关键，为新靶点确认提供扎实证据。

Candida albicans is an important clinical pathogenic fungus causing life-threatening systemic infections, with the multiple pathogenic traits that are mainly regulated by the ATP-dependent Ras1-cAMP-PKA pathway. F1Fo-ATP synthase is the terminal enzyme responsible for mitochondrial ATP synthesis, with the function of the δ subunit remaining enigmatic. Our previous studies have demonstrated that: (i) The absence of the δ subunit in Candida albicans fails to lead to a lethal infection in mice disseminated models, without affecting ATP synthesis; (ii) The activity of Ras1 is significantly suppressed after the deletion of δ subunit, as was the expression level of CYR1; (iii) The evidences for the interaction of the δ subunit with Cdc19 are presented, and bioinformatics analysis indicates a potentially direct interaction between Cdc19-Cyr1-Ras1; (iv) The fructose 1,6-bisphospate (FBP), an allosteric activator of Cdc19, is also significantly reduced. Based on these, we propose that the δ subunit regulates this pathway through FBP/and Cdc19 to lead to a lethal infection rather than affecting ATP synthesis. This project intends to confirm the critical role of δ subunit in leading lethal infection in mice model though in vivo and in vitro studies. Moreover, the metabolomics and biochemical methods are performed to prove that δ subunit does not affect the ATP synthesis in all aspects. Then, protein-protein interactions together with the protein-metabolite interactions will be investigated to better reveal that δ subunit is most likely to regulate this pathway via FBP/and Cdc19. Last, the structure based virtual screening (SBVS) study is to present to identify a novel small molecule compound targeting the δ subunit. The present project will elucidate the key actor of the F1Fo-ATP synthase δ subunit of Candida albicans from a new perspective, and provide solid evidence for new antifungal drug target.