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Differential dynamics of splicing factor SC35 during the cell cycle

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Abstract

Pre-mRNA splicing factors are enriched in nuclear domains termed interchromatin granule clusters or nuclear speckles. During mitosis, nuclear speckles are disassembled by metaphase and reassembled in telophase in structures termed mitotic interchromatin granules (MIGs). We analysed the dynamics of the splicing factor SC35 in interphase and mitotic cells. In HeLa cells expressing green fluorescent protein (GFP)-SC35, this was localized in speckles during interphase and dispersed in metaphase. In telophase, GFP-SC35 was highly enriched within telophase nuclei and also detected in MIGs. Fluorescence recovery after photobleaching (FRAP) experiments revealed that the mobility of GFP-SC35 was distinct in different mitotic compartments. Interestingly, the mobility of GFP-SC35 was 3-fold higher in the cytoplasm of metaphase cells compared with interphase speckles, the nucleoplasm or MIGs. Treatment of cells with inhibitors of cyclin-dependent kinases (cdks) caused changes in the organization of nuclear compartments such as nuclear speckles and nucleoli, with corresponding changes in the mobility of GFP-SC35 and GFP-fibrillarin. Our results suggest that the dynamics of SC35 are significantly influenced by the organization of the compartment in which it is localized during the cell cycle.

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Abbreviations

ASF:

alternate splicing factor

cdk:

cyclin-dependent kinase

Clk:

Cdc2-like kinase

DAPI:

4′,6-diamidino-2-phenylindole

FBS:

foetal bovine serum

FITC:

fluorescein isothiocyanate

FRAP:

fluorescence recovery after photobleaching

GFP:

green fluorescent protein

GST:

glutathione S-transferase

IGC:

interchromatin granule cluster

IPTG:

isopropyl-β-D-thiogalactopyranoside

mAb:

monoclonal antibody

MIG:

mitotic interchromatin granule

NAP:

NOR-associated patch

NOR:

nucleolar organizing region

PBS:

phosphate-buffered saline

PVDF:

polyvinylidene fluoride

RFI:

relative fluorescence intensity

RS:

serine-rich

SDS-PAGE:

sodium dodecyl sulphate-polyacrylamide gel electrophoresis

snRNP:

small nuclear ribonucleoprotein

snoRNA:

small nucleolar RNA

SRPK 1:

serine/arginine-rich protein kinase 1

References

  • Axelrod D, Koppel D E, Schlessinger J, Elson E and Webb W W 1976 Mobility measurement by analysis of fluorescence photobleaching recovery kinetics; Biophys. J. 16 1055–1069

    Article  CAS  Google Scholar 

  • Ben-David Y, Lerwin K, Tannock L, Bernstein A and Pawson T 1991 A mammalian protein kinase with potential for serine/ threonine and tyrosine phosphorylation is related to cell cycle regulators; EMBO J. 10 317–325

    Article  CAS  Google Scholar 

  • Bubulya P A, Prasanth K V, Deerinck T J, Gerlich D, Beaudouin J, Ellisman M H, Ellenberg J and Spector D L 2004 SR splicing factors transiently localize around active nucleolar organizing regions in telophase daughter nuclei; J. Cell Biol. 167 1–15

    Article  Google Scholar 

  • Carmo-Fonseca M, Pepperkok R, Carvalho M T and Lamond A I 1992 Transcription-dependent colocalization of the U1, U2, U4/ U6 and U5 sn-RNPs in coiled bodies; J. Cell Biol. 117 1–14

    Article  CAS  Google Scholar 

  • Colwill K, Pawson T, Andrews B, Prasad J, Manley J L, Bell J C and Duncan P I 1996 The Clk/Sty protein kinase phosphorylates splicing factors and regulates their intranuclear distribution; EMBO J. 15 265–275

    Article  CAS  Google Scholar 

  • Fakan S 1994 Perichromatin fibrils are in situ forms of nascent transcripts; Trends Cell Biol. 4 86–90

    Article  CAS  Google Scholar 

  • Fakan S and Puvion E 1980 The ultrastructural visualization of nucleolar and extranucleolar RNA synthesis and distribution; Int. Rev. Cytol. 65 255–299

    Article  CAS  Google Scholar 

  • Ferreira J A, Carmo-Fonseca M and Lamond A I 1994 Differential interaction of splicing snRNPs with coiled bodies and interchromatin granules during mitosis and assembly of daughter cell nuclei; J. Cell Biol. 126 11–23

    Article  CAS  Google Scholar 

  • Fu X-D and Maniatis T 1990 Factor required for mammalian spliceosome assembly is localized to discrete regions in the nucleus; Nature (London) 343 437–441

    Article  CAS  Google Scholar 

  • Gui J F, Lane W S and Fu X-D 1994 A serine kinase regulates intracellular localization of splicing factors in the cell cycle; Nature (London) 369 678–682

    Article  CAS  Google Scholar 

  • Herrmann C H and Mancini M A 2001 The Cdk9 and cyclin T subunits of TAK/P-TEFb localize to splicing factor-rich nuclear speckle regions; J. Cell Sci. 114 1491–1503

    CAS  PubMed  Google Scholar 

  • Howell B W, Afar D E, Lew J, Douville E M, Icely P L, Gray D A and Bell J C 1991 STY, a tyrosine-phosphorylating enzyme with sequence homology to serine/threonine kinases; Mol. Cell. Biol. 11 568–572

    Article  CAS  Google Scholar 

  • Johnson K W and Smith K A 1991 Molecular cloning of a novel human cdc2/CDC28-like protein kinase; J. Biol. Chem. 266 3402–3407

    CAS  PubMed  Google Scholar 

  • Ko T K, Kelly E and Pines J 2001 CrkRS: a novel conserved Cdc2-related protein kinase that colocalises with SC35 speckles; J. Cell Sci. 114 2591–2603

    CAS  PubMed  Google Scholar 

  • Kruhlak M J, Lever M A, Fischle W, Verdin E, Bazett-Jones D P and Hendzel M J 2000 Reduced mobility of the alternate splicing factor (ASF) through the nucleoplasm and steady state speckle compartments; J. Cell Biol. 150 41–51

    Article  CAS  Google Scholar 

  • Lamond A I and Earnshaw W C 1998 Structure and function in the nucleus; Science 280 547–553

    Article  CAS  Google Scholar 

  • Lamond A I and Spector D L 2003 Nuclear speckles: a model for nuclear organelles; Nat. Rev. Mol. Cell Biol. 4 605–612

    Article  CAS  Google Scholar 

  • Leser G P, Fakan S and Martin T E 1989 Ultrastructural distribution of ribonucleoprotein complexes during mitosis. snRNP antigens are contained in mitotic granule clusters; Eur. J. Cell Biol. 50 376–389

    CAS  PubMed  Google Scholar 

  • Ljungman M and Paulsen M T 2001 The cyclin-dependent kinase inhibitor roscovitine inhibits RNA synthesis and triggers nuclear accumulation of p53 that is unmodified at Ser15 and Lys382; Mol. Pharmacol. 60 785–789

    CAS  PubMed  Google Scholar 

  • Meijer L, Borgne A, Mulner O, Chong J P J, Blow J J, Inagaki N, Inagaki M, Delcros J-G and Moulinoux J-P 1997 Biochemical and cellular effects of roscovitine, a potent and selective inhibitor of the cyclin-dependent kinases cdc2, cdk2 and cdk5; Eur. J. Biochem. 243 527–536

    Article  CAS  Google Scholar 

  • Mintz P J, Patterson S D, Neuwald A F, Spahr C S and Spector D L 1999 Purification and biochemical characterization of interchromatin granule clusters; EMBO J. 18 4308–4320

    Article  CAS  Google Scholar 

  • Misteli T, Caceres J F, Clement J Q, Krainer A R, Wilkinson M F and Spector D L 1998 Serine phosphorylation of SR proteins is required for their recruitment to sites of transcription in vivo; J. Cell Biol. 143 297–307

    Article  CAS  Google Scholar 

  • Misteli T, Caceres J F and Spector D L 1997 The dynamics of a pre-mRNA splicing factor in living cells; Nature (London) 387 523–527

    Article  CAS  Google Scholar 

  • Misteli T and Spector D L 1999 RNA polymerase II targets premRNA splicing factors to transcription sites in vivo; Mol. Cell 3 697–705

    Article  CAS  Google Scholar 

  • Phair R D and Misteli T 2000 High mobility of proteins in the mammalian cell nucleus; Nature (London) 404 604–609

    Article  CAS  Google Scholar 

  • Politz J C R, Tuft R A, Prasanth K V, Baudendistel N, Fogarty K E, Lifshitz L M, Langowski J, Spector D L and Pederson T 2006 Rapid, diffusional shuttling of poly(A) RNA between nuclear speckles and the nucleoplasm; Mol. Biol. Cell 17 1239–1249

    Article  CAS  Google Scholar 

  • Prasanth K V, Sacco-Bubulya P A, Prasanth S G and Spector D L 2003 Sequential entry of components of gene expression machinery into daughter nuclei; Mol. Biol. Cell 14 1043–1057

    Article  CAS  Google Scholar 

  • Reuter R, Appel B, Rinke J and Luhrmann R 1985 Localization and structure of snRNPs during mitosis. Immunofluorescent and biochemical studies; Exp. Cell Res. 159 63–79

    Article  CAS  Google Scholar 

  • Sacco-Bubulya P and Spector D L 2002 Disassembly of interchromatin granule clusters alters the coordination of transcription and pre-mRNA splicing; J. Cell Biol. 156 425–436

    Article  CAS  Google Scholar 

  • Scheer U and Hock R 1999 Structure and function of the nucleolus; Curr. Opin. Cell Biol. 11 385–390

    Article  CAS  Google Scholar 

  • Sirri V, Hernandez-Verdun D and Roussel P 2002 Cyclin-dependent kinases govern formation and maintenance of the nucleolus; J. Cell Biol. 156 969–981

    Article  CAS  Google Scholar 

  • Spector D L 1993 Macromolecular domains within the cell nucleus; Annu. Rev. Cell Biol. 9 265–315

    Article  CAS  Google Scholar 

  • Spector D L 2003 The dynamics of chromosome organisation and gene regulation; Ann. Rev. Biochem. 72 573–608

    Article  CAS  Google Scholar 

  • Spector D L, Fu X-D and Maniatis T 1991 Associations between distinct pre-mRNA splicing components and the cell nucleus; EMBO J. 10 3467–3481

    Article  CAS  Google Scholar 

  • Verheijen R, Kuijpers H, Voojis P, van Venrooij W and Ramaekers F 1986 Protein composition of nuclear matrix preparations from HeLa cells: an immunochemical approach; J. Cell Sci. 80 103–122

    CAS  PubMed  Google Scholar 

  • Voit R, Hoffmann M and Grummt I 1999 Phosphorylation by G1-specific cdk-cyclin complexes activates the nucleolar transcription factor UBF; EMBO J. 18 1891–1899

    Article  CAS  Google Scholar 

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  1. Centre for Cellular and Molecular Biology, Hyderabad, 500 007, India

    Kaushlendra Tripathi & Veena K. Parnaik

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  1. Kaushlendra Tripathi
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  2. Veena K. Parnaik
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Correspondence to Veena K. Parnaik.

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Tripathi, K., Parnaik, V.K. Differential dynamics of splicing factor SC35 during the cell cycle. J Biosci 33, 345–354 (2008). https://doi.org/10.1007/s12038-008-0054-3

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