U1 small nuclear ribonucleoprotein studied by in vitro assembly.
The small nuclear RNAs are known to be complexed with proteins in the cell (snRNP). To learn more about these proteins, we developed an in vitro system for studying their interactions with individual small nuclear RNA species. Translation of HeLa cell poly(A)+ mRNA in an exogenous message-dependent reticulocyte lysate results in the synthesis of snRNP proteins. Addition of human small nuclear RNA U1 to the translation products leads to the formation of a U1 RNA-protein complex that is recognized by a human autoimmune antibody specific for U1 snRNP. This antibody does not react with free U1 RNA. Moreover, addition of a 10- to 20-fold molar excess of transfer RNA instead of U1 RNA does not lead to the formation of an antibody-recognized RNP. The proteins forming the specific complex with U1 RNA correspond to the A, B1, and B2 species (32,000, 27,000, and 26,000 mol wt, respectively) observed in previous studies with U1 snRNP obtained by antibody-precipitation of nuclear extracts. The availability of this in vitro system now permits, for the first time, direct analysis of snRNA-protein binding interactions and, in addition, provides useful information on the mRNAs for snRNP proteins.
D000918:Antibody Specificity; D006367:HeLa Cells / Q000032:analysis; D006801:Humans; D008970:Molecular Weight; D009698:Nucleoproteins / Q000378:metabolism*; D011061:Poly A / Q000378:metabolism; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000378:metabolism; D012261:Ribonucleoproteins / Q000378:metabolism*; D017411:Ribonucleoproteins, Small Nuclear
D009698:Nucleoproteins; D012333:RNA, Messenger; D012261:Ribonucleoproteins; D017411:Ribonucleoproteins, Small Nuclear; D011061:Poly A