Fig. 3: Biophysical model for the sequence specificity of risdiplam and branaplam.

A The “two-interaction-mode model” for how risdiplam and branaplam affect splicing. PSI is assumed to be 100 times the equilibrium occupancy of U1 binding to the 5’ss. Model assumes three sequence-dependent Gibbs free energies: \(\Delta {G}_{{{{{{\rm{U}}}}}}1}\), energy of U1 binding to the 5’ss; \(\Delta {G}_{{{{{{\rm{ris}}}}}}}\), energy of risdiplam binding to the U1/5’ss complex or of branaplam binding to the U1/5’ss complex in the “risdiplam mode”; \(\Delta {G}_{{{{{{\rm{hyp}}}}}}}\), Gibbs free energy of branaplam binding to the U1/5’ss complex in the “hyper-activation mode”. Model parameters were inferred from the PSI values measured by MPSA on cells treated with DMSO, risdiplam, or branaplam (Fig. 1D–F), as well as from drug effect values \(E\) for risdiplam or branaplam determined by the RNA-seq (Fig. 2D). See text, SI Sec. 3.2, and SI Sec. 4.2 for additional information. B–E Experimentally measured vs. model-predicted PSI values and drug-effect values. PSI values are from the SMN2 exon 7 MPSA performed on cells treated with risdiplam or branaplam (Fig. 1); drug-effect values are from the RNA-seq analysis in Fig. 2. F, G Inferred single-nucleotide effects for (F) the “risdiplam energy motif” and (G) the “hyper-activation energy motif”. Top panels show median parameter values illustrated as sequence logos⁷¹. Bottom panels show medians (colored dots, with colors corresponding to each of the four RNA bases as indicated) and 95% credible intervals (colored lines) for motif parameters. Colored squares, median values that lie outside the y-axis limits. 5’ss, 5’ splice site. MPSA, massively parallel splicing assay.