The 5'-terminal sequence of U1 RNA complementary to the consensus 5' splice site of hnRNA is single-stranded in intact U1 snRNP particles

Abstract

The 5'-terminal region of U1 snRNA is highly complementary to the consensus exon-intron regions of hnRNA and it has been suggested that U1 snRNP might play a role in the splicing of the pre-mRNA by intermolecular base-pairing between these regions. Here the secondary structure of the 5' terminus of U1 RNA in the isolated native U1 snRNP particle has been investigated by site-directed enzymatic cleavage of the RNA. Individual oligodeoxynucleotides complementary to various sequences within the first 15 nucleotides of the 5' terminus of U1 RNA have been tested for their ability to form stable DNA X RNA hybrids, with subsequent cleavage of the U1 RNA by RNase H. Our results show unequivocally that the 9 nucleotides at the 5' terminus which are complementary to a consensus 5' splice site are indeed single-stranded in the intact U1 snRNP particle, and are not protected by snRNP proteins. However, they also indicate that the U1 sequence complementary to an intron's consensus 3' end is not readily available for intermolecular base-pairing, either in the intact U1 snRNP particle or in the deproteinized U1 RNA molecule. Therefore our data favour the possibility that U1 snRNP plays a role only in the recognition of a 5' splice site of hnRNA, rather than being involved in the alignment of both ends of an intron for splicing.