The U1 snRNP protein U1C recognizes the 5′ splice site in the absence of base pairing

Splicing of precursor messenger RNA takes place in the spliceosome, a large RNA/protein macromolecular machine1. Spliceosome assembly occurs in an ordered pathway in vitro and is conserved between yeast and mammalian systems. The earliest step is commitment complex formation in yeast or E complex formation in mammals, which engages the pre-mRNA in the splicing pathway and involves interactions between U1 small nuclear ribonucleoprotein (snRNP) and the pre-mRNA 5′ splice site2,3. Complex formation depends on highly conserved base pairing between the 5′ splice site and the 5′ end of U1 snRNA, both in vivo and in vitro4,5,6,7. U1 snRNP proteins also contribute to U1 snRNP activity8,9,10. Here we show that U1 snRNP lacking the 5′ end of its snRNA retains 5′-splice-site sequence specificity. We also show that recombinant yeast U1C protein, a U1 snRNP protein, selects a 5′-splice-site-like sequence in which the first four nucleotides, GUAU, are identical to the first four nucleotides of the yeast 5′-splice-site consensus sequence. We propose that a U1C 5′-splice-site interaction precedes pre-mRNA/U1 snRNA base pairing and is the earliest step in the splicing pathway.