Disassembly of interchromatin granule clusters alters the coordination of transcription and pre-mRNA splicing.
To examine the involvement of interchromatin granule clusters (IGCs) in transcription and pre-mRNA splicing in mammalian cell nuclei, the serine-arginine (SR) protein kinase cdc2-like kinase (Clk)/STY was used as a tool to manipulate IGC integrity in vivo. Both immunofluorescence and transmission electron microscopy analyses of cells overexpressing Clk/STY indicate that IGC components are completely redistributed to a diffuse nuclear localization, leaving no residual structure. Conversely, overexpression of a catalytically inactive mutant, Clk/STY(K190R), causes retention of hypophosphorylated SR proteins in nuclear speckles. Our data suggest that the protein-protein interactions responsible for the clustering of interchromatin granules are disrupted when SR proteins are hyperphosphorylated and stabilized when SR proteins are hypophosphorylated. Interestingly, cells without intact IGCs continue to synthesize nascent transcripts. However, both the accumulation of splicing factors at sites of pre-mRNA synthesis as well as pre-mRNA splicing are dramatically reduced, demonstrating that IGC disassembly perturbs coordination between transcription and pre-mRNA splicing in mammalian cell nuclei.
Sacco-Bubulya
Paula
P
Cold Spring Harbor Laboratory, Cold Spring Harbor, NY 11724, USA.
Spector
David L
DL
D002467:Cell Nucleus / Q000378:metabolism* / Q000648:ultrastructure; D002478:Cells, Cultured; D002843:Chromatin / Q000378:metabolism* / Q000648:ultrastructure; D003594:Cytoplasmic Granules / Q000378:metabolism* / Q000648:ultrastructure; D003599:Cytoskeleton / Q000235:genetics / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D015971:Gene Expression Regulation, Enzymologic / Q000502:physiology; D049452:Green Fluorescent Proteins; D006801:Humans; D008164:Luminescent Proteins / Q000235:genetics; D008854:Microscopy, Electron; D009154:Mutation / Q000235:genetics; D017346:Protein Serine-Threonine Kinases / Q000235:genetics* / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000235:genetics* / Q000378:metabolism; D012326:RNA Splicing / Q000502:physiology*; D012333:RNA, Messenger / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D014158:Transcription, Genetic / Q000502:physiology*
D002843:Chromatin; D008164:Luminescent Proteins; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; C066780:Clk dual-specificity kinases; D011505:Protein-Tyrosine Kinases; D017346:Protein Serine-Threonine Kinases