Inefficient pre-spliceosome formation on FOXP3 intron 11. (A) Diagram of a pre-spliceosome complex. In the presence of ATP and two DExD-box helicases, DDX46 and DDX39B (DDX39A), SF1 is displaced from the BPS, allowing for U2 snRNP to bind and form the pre-spliceosome (created with BioRender). (B) Abundance of U1 snRNP components on F3i11wt-tobra and F3i11pU-tobra. (C) Abundance of SF1 and U2AF heterodimers on F3i11wt-tobra and F3i11pU-tobra. (D) Abundance of U2 snRNP components on F3i11wt-tobra and F3i11pU-tobra. In B–E, all incubations are in the presence of added ATP, and each bar graph is the median ± interquartile range of four replicates from two independent experiments; (*) P < 0.05, Mann–Whitney test.