Recruitment of the human TREX complex to mRNA during splicing.

In yeast, the TREX complex contains the THO transcription elongation complex, which functions in direct cotranscriptional recruitment of the mRNA export proteins Sub2 and Yra1 to nascent transcripts. Here we report the identification of the human THO complex and show that it associates with spliced mRNA, but not with unspliced pre-mRNA in vitro. Transcription is not required for this recruitment. We also show that the human THO complex colocalizes with splicing factors in nuclear speckle domains in vivo. Considering that splicing occurs cotranscriptionally in humans, our data indicate that recruitment of the human TREX complex to spliced mRNA is not directly coupled to transcription, but is instead coupled to transcription indirectly through splicing.

[1]  E. Hurt,et al.  The protein Aly links pre-messenger-RNA splicing to nuclear export in metazoans , 2000, Nature.

[2]  Paul Tempst,et al.  Nab2p and the Thp1p-Sac3p Complex Functionally Interact at the Interface between Transcription and mRNA Metabolism* , 2003, Journal of Biological Chemistry.

[3]  Henning Urlaub,et al.  Small Nuclear Ribonucleoprotein Remodeling During Catalytic Activation of the Spliceosome , 2002, Science.

[4]  Steven P. Gygi,et al.  Comprehensive proteomic analysis of the human spliceosome , 2002, Nature.

[5]  S. Chávez,et al.  Hpr1 Is Preferentially Required for Transcription of Either Long or G+C-Rich DNA Sequences in Saccharomyces cerevisiae , 2001, Molecular and Cellular Biology.

[6]  R. Reed,et al.  Coupling transcription, splicing and mRNA export. , 2003, Current opinion in cell biology.

[7]  D. Bentley.  The mRNA assembly line: transcription and processing machines in the same factory. , 2002, Current opinion in cell biology.

[8]  B. Blencowe,et al.  In vivo recruitment of exon junction complex proteins to transcription sites in mammalian cell nuclei. , 2004, RNA.

[9]  P. Silver,et al.  Messenger RNAs are recruited for nuclear export during transcription. , 2001, Genes & development.

[10]  E. Harlow,et al.  Antibodies: A Laboratory Manual , 1988 .

[11]  Kevin Struhl,et al.  TREX is a conserved complex coupling transcription with messenger RNA export , 2002, Nature.

[12]  Nikolaus Grigorieff,et al.  Purification and characterization of native spliceosomes suitable for three-dimensional structural analysis. , 2002, RNA.

[13]  E. Izaurralde.  A novel family of nuclear transport receptors mediates the export of messenger RNA to the cytoplasm. , 2002, European journal of cell biology.

[14]  T. Köcher,et al.  The DExH/D box protein HEL/UAP56 is essential for mRNA nuclear export in Drosophila , 2001, Current Biology.

[15]  T. Maniatis,et al.  An extensive network of coupling among gene expression machines , 2002, Nature.

[16]  N. Krogan,et al.  Transitions in RNA polymerase II elongation complexes at the 3′ ends of genes , 2004, The EMBO journal.

[17]  Juri Rappsilber,et al.  Mass spectrometry and EST-database searching allows characterization of the multi-protein spliceosome complex , 1998, Nature Genetics.

[18]  R. Luna,et al.  The yeast THO complex and mRNA export factors link RNA metabolism with transcription and genome instability , 2002, The EMBO journal.

[19]  Trevor Lithgow,et al.  A protein complex containing Tho2, Hpr1, Mft1 and a novel protein, Thp2, connects transcription elongation with mitotic recombination in Saccharomyces cerevisiae , 2000, The EMBO journal.

[20]  Daniel Zenklusen,et al.  Coupling of Termination, 3′ Processing, and mRNA Export , 2002, Molecular and Cellular Biology.

[21]  P. Silver,et al.  Intron status and 3'-end formation control cotranscriptional export of mRNA. , 2002, Genes & development.

[22]  Ed Hurt,et al.  A Conserved mRNA Export Machinery Coupled to pre-mRNA Splicing , 2002, Cell.

[23]  J. Manley,et al.  RNA polymerase II and the integration of nuclear events. , 2000, Genes & development.

[24]  Andrés Aguilera,et al.  Cotranscriptionally formed DNA:RNA hybrids mediate transcription elongation impairment and transcription-associated recombination. , 2003, Molecular cell.

[25]  A. Aguilera,et al.  Molecular Evidence That the Eukaryotic THO/TREX Complex Is Required for Efficient Transcription Elongation* , 2003, Journal of Biological Chemistry.

[26]  T. Köcher,et al.  Genome-wide analysis of mRNAs regulated by the THO complex in Drosophila melanogaster , 2004, Nature Structural &Molecular Biology.

[27]  M. Rosbash,et al.  Interactions between mRNA Export Commitment, 3′-End Quality Control, and Nuclear Degradation , 2002, Molecular and Cellular Biology.

[28]  Daniel Zenklusen,et al.  Stable mRNP Formation and Export Require Cotranscriptional Recruitment of the mRNA Export Factors Yra1p and Sub2p by Hpr1p , 2002, Molecular and Cellular Biology.

[29]  D. Spector,et al.  Macromolecular domains within the cell nucleus. , 1993, Annual review of cell biology.

[30]  Andrés Aguilera,et al.  A novel yeast gene, THO2, is involved in RNA pol II transcription and provides new evidence for transcriptional elongation‐associated recombination , 1998, The EMBO journal.

[31]  Tamás Fischer,et al.  The mRNA export machinery requires the novel Sac3p–Thp1p complex to dock at the nucleoplasmic entrance of the nuclear pores , 2002, The EMBO journal.

[32]  A. Aguilera,et al.  Molecular evidence for a positive role of Spt4 in transcription elongation , 2003, The EMBO journal.

[33]  M. Mann,et al.  Large-scale Proteomic Analysis of the Human Spliceosome References , 2006 .

[34]  S. Lacadie,et al.  Biochemical analysis of TREX complex recruitment to intronless and intron‐containing yeast genes , 2004, The EMBO journal.

[35]  M. Mann,et al.  Pre-mRNA splicing and mRNA export linked by direct interactions between UAP56 and Aly , 2001, Nature.