The Mr 70,000 protein of the U1 small nuclear ribonucleoprotein particle binds to the 5' stem-loop of U1 RNA and interacts with Sm domain proteins.

AUTOR(ES)

Patton, J R

RESUMO

The U1 small nuclear ribonucleoprotein (snRNP) particle, a cofactor in mRNA splicing, contains nine proteins, six of which are also present in other U snRNPs and three of which are specific to the U1 snRNP. Here we have used a reconstituted human U1 snRNP together with snRNP monoclonal antibodies to define the RNA binding sites of one of the U1 snRNP-specific proteins. When Sm monoclonal antibody (specific for the B', B, and D proteins of U snRNPs) was bound to U1 snRNPs prior to micrococcal nuclease digestion, the same approximately equal to 24 nucleotide fragment of U1 RNA (corresponding to nucleotides 120-143 and termed the "Sm domain") was protected as when no antibody was bound prior to digestion. In contrast, when RNP monoclonal antibody, which reacts with the U1 snRNP-specific Mr 70,000 protein, was bound, additional U1 RNA regions were protected against nuclease digestion. This phenomenon, which we term "antibody-mediated nuclease protection," was exploited to map the position of the Mr 70,000 protein to stem-loop I of U1 RNA. However, there were also sites of Mr 70,000 protein interaction with more 3'-ward regions of U1 RNA, particularly the Sm domain. This indicates that in the three-dimensional structure of the U1 snRNP, the RNP and Sm antigens are in contact with each other. The proximity of the Mr 70,000 protein's RNA binding site (stem-loop I) to the functionally important 5' end of U1 RNA suggests that this protein may be involved in the recognition of, or stabilization of base pairing with, pre-mRNA 5' splice sites.

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