Fig. 3: U1-C is required for branaplam modulation of -1A 5′SS RNA binding.
From: A sequential binding mechanism for 5′ splice site recognition and modulation for the human U1 snRNP
A Fluorescence in arbitrary units (au) across time in seconds (s) of 1 nM 9bp-1A binding in the absence of U1-C with (top) DSMO and (bottom) 10 µM branaplam overlaid with idealizations (black lines). B Violin plots of bound dwell time distributions at 1 nM 9bp-1A RNA with and without U1-C and/or branaplam. DMSO was included in the absence of branaplam. C Change in fluorescence due to branaplam binding to a duplex of 11bp-1A and U1 snRNP in the absence (grey triangles) and presence (white circles) of U1-C by microscale thermophoresis (MST). The change in fluorescence in the presence of U1-C is overlaid with a fitted equation (solid purple) and 95% confidence interval of the fitted equation (shaded purple region) to estimate a KD value (2.69 ± 0.36 µM). D Violin plots showing the bound dwell time distributions across different permutations of U1-C and branaplam concentrations in solution across indicated RNA oligo sequences (SMN2, FOXM1, SF3B3, HTT*). Each violin plot is overlaid with box plot that show the median (horizontal line), interquartile range (IQR, box), and whiskers representing data within 1.5\(\times\)IQR. Highlighted nucleotides in the 5′SS sequences above each plot indicate predicted base pairs to the U1 snRNA. The lower case letters in HTT* indicate a +7 G:A and +8G:U substitutions included to enable RNA synthesis. For B and D, numbers above the violins indicate the number of bound lifetimes included in each distribution. Source data are provided as a Source data file.
