(A) Domain composition of the related RBM5, RBM6 and RBM10 family proteins. (B) Sequence alignment of OCRE domains of human RBM5, RBM10 and RBM6 proteins. (C,D) Two views in a cartoon presentation of the RBM5 OCRE domain, showing side chains of conserved and exposed tyrosine residues, some of which were probed by mutational analysis. (C) At one side of the β-sheet surface the N-terminal extension shields the tyrosines (orange) from solvent exposure, (D) on the opposite surface of the β-sheet, numerous tyrosine residues (yellow) are solvent accessible. (E) Mutational analysis of conserved residues of the RBM5 OCRE domain. Specific mutations of conserved and accessible residues of RBM5 OCRE domain impair the activity of the protein in FAS alternative splicing regulation ex vivo. HeLa cells were co-transfected with a Fas wild type alternative splicing reporter (harbouring sequences between the 5’ end of exon 5 and the first 47 nucleotides of exon 7) and T7-RBM5 expression plasmids. RNA and proteins were isolated 24 hr after transfection. Patterns of alternative splicing were studied by RT-PCR using specific primers (PT1, PT2) and the percentage of inclusion was calculated and is presented in the histogram for a minimum of 16 replicas of the experiment. T-test (two-tailed distribution, homoscedastic) results are mentioned (**<0,01; ***<0,001). Full quantification and T-test results are provided in Figure 5—figure supplement 2.