Succession of splicing regulatory elements determines cryptic 5΄ss functionality.

A critical step in exon definition is the recognition of a proper splice donor (5'ss) by the 5' end of U1 snRNA. In the selection of appropriate 5' ss, cis-acting splicing regulatory elements (SREs) are indispensable. As amodel for 5' ss recognition, we investigated cryptic 5' ss selection within the human fibrinogen B beta-chain gene (FGB) exon 7, where we identified several exonic SREs that simultaneously acted on up-and downstream cryptic 5'ss. In the FGB exon 7 model system, 5'ss selection iteratively proceeded along an alternating sequence of U1 snRNA binding sites and interleaved SREs which in principle supported different 3'exon ends. Like in a relay race, SREs either suppressed a potential 5'ss and passed the splicing baton on or splicing actually occurred. From RNA-Seq data, we systematically selected 19 genes containing exons with silent U1 snRNA binding sites competing with nearby highly used 5'ss. Extensive SRE analysis by different algorithms found authentic 5'ss significantlymore supported by SREs than silent U1 snRNA binding sites, indicating that our concept may permit generalization to a model for 5'ss selection and 3'exon end definition.