Here, we report the systematic integration of cellular and biochemical analyses, transcriptomic, PRPF8 interactome and proteomic datasets to reveal the importance of the fine-tuning of hBrr2 by PRPF8 in the spliceosome for pre-mRNA splicing and the functional consequences of its disruption by RP-related mutations, unravelling the mechanism of PRPF-RP pathogenesis. Here, PRPF8 is linked to retinitis pigmentosa 1.