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.
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4 February 2002
Article|
February 04 2002
Disassembly of interchromatin granule clusters alters the coordination of transcription and pre-mRNA splicing
Paula Sacco-Bubulya,
Paula Sacco-Bubulya
Cold Spring Harbor Laboratory, Cold Spring Harbor, NY 11724
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David L. Spector
David L. Spector
Cold Spring Harbor Laboratory, Cold Spring Harbor, NY 11724
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Paula Sacco-Bubulya
Cold Spring Harbor Laboratory, Cold Spring Harbor, NY 11724
David L. Spector
Cold Spring Harbor Laboratory, Cold Spring Harbor, NY 11724
Address correspondence to David L. Spector, Cold Spring Harbor Laboratory, 1 Bungtown Rd., Cold Spring Harbor, NY 11724. Tel.: (516) 367-8456. Fax: (516) 367-8876. E-mail: [email protected]
The online version of this article contains supplemental material.
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Abbreviations used in this paper: Clk, cdc2-like kinase; DIG, digoxygenin; GFP, green fluorescent protein; IGC, interchromatin granule cluster; PF, perichromatin fibril; RS, arginine-serine rich; SR, serine-arginine; TEM, transmission electron microscopy.
Received:
July 05 2001
Revision Received:
December 10 2001
Accepted:
December 14 2001
Online ISSN: 1540-8140
Print ISSN: 0021-9525
The Rockefeller University Press
2002
J Cell Biol (2002) 156 (3): 425–436.
Article history
Received:
July 05 2001
Revision Received:
December 10 2001
Accepted:
December 14 2001
Citation
Paula Sacco-Bubulya, David L. Spector; Disassembly of interchromatin granule clusters alters the coordination of transcription and pre-mRNA splicing . J Cell Biol 4 February 2002; 156 (3): 425–436. doi: https://doi.org/10.1083/jcb.200107017
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