Exon‐independent recruitment of SRSF1 is mediated by U1 snRNP stem‐loop 3.

SRSF1 protein and U1 snRNPs are closely connected splicing factors. They both stimulate exon inclusion, SRSF1 by binding to exonic splicing enhancer sequences (ESEs) and U1 snRNPs by binding to the downstream 5′ splice site (SS), and both factors affect 5′ SS selection. The binding of U1 snRNPs initiates spliceosome assembly, but SR proteins such as SRSF1 can in some cases substitute for it. The mechanistic basis of this relationship is poorly understood. We show here by single‐molecule methods that a single molecule of SRSF1 can be recruited by a U1 snRNP. This reaction is independent of exon sequences and separate from the U1‐independent process of binding to an ESE. Structural analysis and cross‐linking data show that SRSF1 contacts U1 snRNA stem‐loop 3, which is required for splicing. We suggest that the recruitment of SRSF1 to a U1 snRNP at a 5′SS is the basis for exon definition by U1 snRNP and might be one of the principal functions of U1 snRNPs in the core reactions of splicing in mammals. Synopsis: Exons are activated for splicing by the binding either of proteins, such as SRSF1, to the exon, or U1 snRNPs to 5′ splice sites, but it is unclear if they act via a common mechanism. Here, SRSF1 is found to be recruited to splice‐sites through an interaction with stem‐loop 3 of U1 snRNA, possibly constituting one of the major functions for the U1 snRNP in splicing. Single‐molecule colocalisation microscopy shows that U1 snRNPs form a stoichiometric complex with SRSF1 and can recruit it to a 5′‐splice site.NMR spectroscopy shows that RNA‐binding domains of SRSF1 interact with stem‐loop 3 of U1 snRNA.The association of SRSF1 with U1snRNP does not require pre‐mRNA.U1 snRNP and exonic splicing enhancer sequences independently recruit SRSF1. [ABSTRACT FROM AUTHOR]