Antisense probing of the human U4/U6 snRNP with biotinylated 2'-OMe RNA oligonucleotides.
We have used antisense 2'-OMe RNA oligonucleotides carrying four 5'-terminal biotin residues to probe the structure and function of the human U4/U6 snRNP. Nine oligonucleotides, complementary to multiple regions of U4 and U6 snRNAs, bound stably and specifically to U4/U6 snRNP. This allowed for efficient and selective removal of U4/U6 from HeLa cell nuclear extracts. Binding of oligonucleotides to certain snRNA domains inhibited splicing and affected the U4-U6 interaction. Pre-mRNA and splicing products could also be affinity-selected through binding of the oligonucleotides to U4/U6 snRNPs in splicing complexes. The results suggest that U4 snRNP is not released during spliceosome assembly.
Blencowe
B J
BJ
European Molecular Biology Laboratory, Heidelberg Federal Republic of Germany.
Sproat
B S
BS
Ryder
U
U
Barabino
S
S
Lamond
A I
AI
D001483:Base Sequence; D001710:Biotin; D015152:Blotting, Northern; D002846:Chromatography, Affinity; D006801:Humans; D008745:Methylation; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D009693:Nucleic Acid Hybridization; D015345:Oligonucleotide Probes; D012313:RNA / Q000235:genetics*; D012326:RNA Splicing / Q000235:genetics; D016372:RNA, Antisense; D012333:RNA, Messenger / Q000037:antagonists & inhibitors; D012261:Ribonucleoproteins / Q000235:genetics* / Q000302:isolation & purification; D017411:Ribonucleoproteins, Small Nuclear
D015345:Oligonucleotide Probes; D016372:RNA, Antisense; D012333:RNA, Messenger; D012261:Ribonucleoproteins; D017411:Ribonucleoproteins, Small Nuclear; D012313:RNA; D001710:Biotin