Structural basis of Nur77-CypA interaction and cyclosporine A-mediated competitive displacement.

Nuclear receptor Nur77, along with cyclophilin A (CypA) and cyclosporine A (CsA), has been shown to participate in crosstalk of signaling cascades associated with inflammation, apoptosis and metabolic regulation. However, the fundamental molecular mechanism governing their mutual interactions is unclear. Using NMR spectroscopy combined with AI-driven docking, we resolved the Nur77 ligand-binding domain (LBD)-CypA interface. NMR analysis mapped the binding mode, revealing a weak yet specific protein-protein interaction. Integration of experimental results with AI modeling produced a high-confidence complex structural model validated by mutational analysis and expanding investigations through the NR4A family paralogs. Strikingly, CsA does not directly bind Nur77 LBD. Rather, it occupies CypA and competitively displaces Nur77 from the same binding groove, as demonstrated by competition assays. Our findings elucidate the mechanism for the Nur77-CypA interaction and refine CsA's mode of action, with implications for therapeutic development.