The 5' terminus of the RNA moiety of U1 small nuclear ribonucleoprotein particles is required for the splicing of messenger RNA precursors.
We have investigated the role of small nuclear ribonucleoprotein particles (snRNPs) in the in vitro splicing of messenger RNA precursors by a variety of procedures. Removal of the U-type snRNPs from the nuclear extracts of HeLa cells with protein A-Sepharose-coupled human autoimmune antibodies leads to complete loss of splicing activity. The inhibition of splicing can be prevented by saturating the coupled antibodies with purified nucleoplasmic U snRNPs prior to incubation with nuclear extract. We further demonstrate that an intact 5' terminus of U1 snRNA is required for the functioning of U1 snRNP in the splicing reaction. Antibodies directed against the trimethylated cap structure of the U snRNAs inhibit splicing. Upon removal of the first eight nucleotides of the U1 snRNA in the particles by site-directed hydrolysis with ribonuclease H in the presence of a synthetic complementary oligodeoxynucleotide splicing is completely abolished. These results are in strong support of current models suggesting that a base-pairing interaction between the 5' terminus of the U1 snRNA and the 5' splice site of a mRNA precursor is a prerequisite for proper splicing.
D000936:Antigen-Antibody Complex; D001483:Base Sequence; D002467:Cell Nucleus / Q000378:metabolism; D006367:HeLa Cells / Q000378:metabolism; D006801:Humans; D007106:Immune Sera; D009694:Nucleic Acid Precursors / Q000235:genetics*; D010957:Plasmids; D012322:RNA Precursors; D012326:RNA Splicing; D012333:RNA, Messenger / Q000235:genetics*; D012261:Ribonucleoproteins / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism*; D017411:Ribonucleoproteins, Small Nuclear
D000936:Antigen-Antibody Complex; D007106:Immune Sera; D009694:Nucleic Acid Precursors; D012322:RNA Precursors; D012333:RNA, Messenger; D012261:Ribonucleoproteins; D017411:Ribonucleoproteins, Small Nuclear