Disruption of pre-mRNA splicing in vivo results in reorganization of splicing factors.
We have examined the functional significance of the organization of pre-mRNA splicing factors in a speckled distribution in the mammalian cell nucleus. Upon microinjection into living cells of oligonucleotides or antibodies that inhibit pre-mRNA splicing in vitro, we observed major changes in the organization of splicing factors in vivo. Interchromatin granule clusters became uniform in shape, decreased in number, and increased in both size and content of splicing factors, as measured by immunofluorescence. These changes were transient and the organization of splicing factors returned to their normal distribution by 24 h following microinjection. Microinjection of these oligonucleotides or antibodies also resulted in a reduction of transcription in vivo, but the oligonucleotides did not inhibit transcription in vitro. Control oligonucleotides did not disrupt splicing or transcription in vivo. We propose that the reorganization of splicing factors we observed is the result of the inhibition of splicing in vivo.
O'Keefe
R T
RT
Cold Spring Harbor Laboratory, New York 11724.
Mayeda
A
A
Sadowski
C L
CL
Krainer
A R
AR
Spector
D L
DL
D001345:Autoradiography; D001483:Base Sequence; D002467:Cell Nucleus / Q000737:chemistry / Q000378:metabolism* / Q000648:ultrastructure; D006367:HeLa Cells; D006801:Humans; D008845:Microinjections; D008854:Microscopy, Electron; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000032:analysis / Q000378:metabolism*; D009838:Oligodeoxyribonucleotides / Q000494:pharmacology; D012322:RNA Precursors / Q000235:genetics / Q000378:metabolism*; D012326:RNA Splicing; D012342:RNA, Small Nuclear / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D012261:Ribonucleoproteins; D017411:Ribonucleoproteins, Small Nuclear / Q000032:analysis / Q000378:metabolism*; D000068103:Serine-Arginine Splicing Factors; D017381:Spliceosomes / Q000378:metabolism / Q000648:ultrastructure; D014158:Transcription, Genetic
D009687:Nuclear Proteins; D009838:Oligodeoxyribonucleotides; D012322:RNA Precursors; D012342:RNA, Small Nuclear; D012261:Ribonucleoproteins; D017411:Ribonucleoproteins, Small Nuclear; C073288:SRSF2 protein, human; D000068103:Serine-Arginine Splicing Factors