Alphafold 3 guided insights into the Importin b / Importin 7 heterodimer interaction and its binding to Histone H1

The nuclear import of H1 linker histones is facilitated by a heterodimer of the transport receptors Importinb (Impb) and Importin7 (Imp7). While both importins can individually interact with H1, only their preassembled hetero-dimer enables its proper binding for translocation through the nuclear pore. The interaction between Imp7 and Impb is mediated by a short stretch of residues in the C-terminal region of Imp7, which plays a key role in Imp7 activation by Impb. This interaction is allosterically regulated by Impb, finely tuning the activity of the Impb/Imp7 hetero-dimer. A complete model of the Impb:Imp7:H1 complex was predicted by Alphafold3 (AF3) and subsequently validated using cross-linking (X-link) data, isothermal titration calorimetry (ITC), and pull-down experiments, providing robust support for the models accuracy. This model positions the globular domain of H1 within the central cavity of Imp7, in agreement with cross-linking data. Refinement of this atomic model against a previously published cryo-EM map demonstrated significantly improved correspondence compared to the earlier interpretation, which placed the H1 globular domain within Impb. This enhanced structural consistency further substantiates the accuracy of the AI-driven prediction. Detailed analysis identified the nucleoporin-like binding (NlB) region of Imp7 as a short stretch interacting with the outer surface of Impb. This interaction mode implies that regulation of FG binding site accessibility on Impb, mediated by the FXFG nucleoporin motives within the Imp7 NlB region, likely modulates transport pathways and/or enhances the efficiency of H1 nuclear import.