biostudies-literatureUnknown32(5)Soni KIntron removal during pre-mRNA splicing is of extraordinary complexity and its disruption causes a vast number of genetic diseases in humans. While key steps of the canonical spliceosome cycle have been revealed by combined structure-function analyses, structural information on an aberrant spliceosome committed to premature disassembly is not available. Here, we report two cryo-electron microscopy structures of post-B^act spliceosome intermediates from Schizosaccharomyces pombe primed for disassembly. We identify the DEAH-box helicase-G-patch protein pair (Gih35-Gpl1, homologous to human DHX35-GPATCH1) and show how it maintains catalytic dormancy. In both structures, Gpl1 recognizes a remodeled active site introduced by an overstabilization of the U5 loop I interaction with the 5Nature structural & molecular biology914-925https://www.ebi.ac.uk/biostudies/studies/S-EPMC12086092biostudies-literatureStructures of aberrant spliceosome intermediates on their way to disassembly.PMC12086092Dybkov OTrakansuebkul SFischer TSoni KSchwan MUrlaub HHorvath AFlemming DWild KSinning IfalseStructures of aberrant spliceosome intermediates on their way to disassembly.Intron removal during pre-mRNA splicing is of extraordinary complexity and its disruption causes a vast number of genetic diseases in humans. While key steps of the canonical spliceosome cycle have been revealed by combined structure-function analyses, structural information on an aberrant spliceosome committed to premature disassembly is not available. Here, we report two cryo-electron microscopy structures of post-B^act spliceosome intermediates from Schizosaccharomyces pombe primed for disassembly. We identify the DEAH-box helicase-G-patch protein pair (Gih35-Gpl1, homologous to human DHX35-GPATCH1) and show how it maintains catalytic dormancy. In both structures, Gpl1 recognizes a remodeled active site introduced by an overstabilization of the U5 loop I interaction with the 52025-01-01T00:00:00Z2025 May2026-06-01T06:16:01.727Z2026-04-08T09:54:03.051ZS-EPMC120860923983347010.1038/s41594-024-01480-7