Fig. 5: Molecular Dynamics (MD) data for Lin28 and YB-1 heterotrimers indicates the mechanism behind the mixing between YB-1 and Lin28 in the presence of RNA.

a MD structures of YB-1 and Lin28 hetero-and homotypic trimer formed in the presence of 16 nt-long Poly(C) RNA (see Materials and Methods for details). Zooms in on the interaction between CSD loop 3 and CTD that controls the cooperative association to RNA. b Interaction energies at the protein-protein and protein-RNA levels for indicated homo- and heterotrimers (ABC) interacting with 16-nt long Poly(C) RNA. Upper panel: global RNA-protein interactions for each protein, A, B, and C. The interaction energy of protein A with RNA is low since protein A interacts with less nucleotides than proteins B and C in the presence of the 16-nt-long RNA. When RRM1 of TDP-43 is located in the middle of the trimer (as molecule B), it significantly reduces the interaction of flanking proteins, A and C, with RNA. Middle panel: Interaction energies of Q76 and S77 of protein C with RNA for indicated homo-or hetero-trimers. Q76 and S77 of protein C are located at the interface between protein B and C. We noticed that Q76 and S77 poorly interact with RNA when protein B is TDP-43 RRM1 compared to YB-1 or Lin28. Lower panel: energies of interaction between Q76, S77 being mutated into alanines, with RNA in indicated trimers. In comparison to wild type, the contact of mutated Lin28 residues to RNA decreases dramatically. Energies were averaged over 200 ns of MD simulation (for RRM1 of TDP-43–10 ns were sufficient to observe a reduced interaction) and values are reported in kJ/mol with variant of fluctuations being ± 0.4 kJ/mol. c Representative images of the mixing/demixing Lin28 mutants and YB-1 along microtubules in HeLa cells. The relative enrichment of Lin28 and YB-1 compartments was measured as described in Methods. G119A and S120A mutations lead to a marked demixing between YB-1 and Lin28. H80A is a negative control. Scale bar: 5 µm.