Identification of the sequence determinants mediating the nucleo-cytoplasmic shuttling of TIAR and TIA-1 RNA-binding proteins

TIAR and TIA-1 are two closely related RNA-binding proteins which possess three RNA recognition motifs (RRMs) followed by an auxiliary region. These proteins are involved in several mechanisms of RNA metabolism, including alternative hnRNA splicing and regulation of mRNA translation. Here we characterize the subcellular localization of these proteins in somatic cells. We demonstrate that TIAR and TIA-1 continuously shuttle between the cytoplasm and the nucleus and belong to the class of RNA-binding proteins whose nuclear import is transcription-dependent. We identified RRM2 and the first half of the auxiliary region as important determinants for TIAR and TIA-1 nuclear accumulation. In contrast, the nuclear export of TIAR and TIA-1 is mediated by RRM3. Both RRMs contribute to TIAR and TIA-1 nuclear accumulation or export by their RNA-binding capacity. Indeed, whereas mutations of the highly conserved RNP2 or RNP1 peptides in RRM2 redistribute TIAR to the cytoplasm, similar modifications in RRM3 abolish TIAR nuclear export. Moreover, TIAR and TIA-1 nuclear accumulation is a Ran-GTP-dependent pathway, in contrast to its nuclear export which is unaffected by Ran-GTP depletion and which is independent of the major CRM1-exporting pathway. This study demonstrates the importance of TIAR and TIA-1 RNA-binding domains for their subcellular localization and provides the first evidence for distinct functions of TIAR and TIA-1 RRMs.

[1]  K. Williams,et al.  Phenylalanines that are conserved among several RNA-binding proteins form part of a nucleic acid-binding pocket in the A1 heterogeneous nuclear ribonucleoprotein. , 1988, The Journal of biological chemistry.

[2]  J. Steitz,et al.  Transportins 1 and 2 are redundant nuclear import factors for hnRNP A1 and HuR. , 2004, RNA.

[3]  David Levens,et al.  Nuclear targeting determinants of the far upstream element binding protein, a c-myc transcription factor. , 2000, Nucleic acids research.

[4]  M. Streuli,et al.  RNA-binding protein TIAR is essential for primordial germ cell development. , 1998, Proceedings of the National Academy of Sciences of the United States of America.

[5]  F. Bischoff,et al.  Identification of two novel RanGTP-binding proteins belonging to the importin beta superfamily. , 2000, The Journal of biological chemistry.

[6]  M. Schäfer,et al.  Identification of a novel putative Ran-binding protein and its close homologue. , 2000, Biochemical and biophysical research communications.

[7]  P. Anderson,et al.  TIA‐1 is a translational silencer that selectively regulates the expression of TNF‐α , 2000 .

[8]  N. Standart,et al.  The Xenopus ELAV Protein ElrB Represses Vg1 mRNA Translation during Oogenesis , 2005, Molecular and Cellular Biology.

[9]  P. Evans,et al.  Crystal structure of the RNA-binding domain of the U1 small nuclear ribonucleoprotein A , 1990, Nature.

[10]  W. Richardson,et al.  The nucleoplasmin nuclear location sequence is larger and more complex than that of SV-40 large T antigen , 1988, The Journal of cell biology.

[11]  P. Anderson,et al.  Regulation of Cyclooxygenase-2 Expression by the Translational Silencer TIA-1 , 2003, The Journal of experimental medicine.

[12]  F. Bischoff,et al.  Identification of Two Novel RanGTP-binding Proteins Belonging to the Importin β Superfamily* , 2000, The Journal of Biological Chemistry.

[13]  J. Steitz,et al.  Protein Ligands to Hur Modulate Its Interaction with Target Mrnas in Vivo , 2000, The Journal of cell biology.

[14]  A. Philpott,et al.  The role of nucleoplasmin in chromatin assembly and disassembly. , 1993, Philosophical transactions of the Royal Society of London. Series B, Biological sciences.

[15]  Xiao‐hui Huang,et al.  Increase of beta1-adrenergic receptor gene expression induced by nicotine in hippocampal slice of rat. , 2003, Acta pharmacologica Sinica.

[16]  G. Dreyfuss,et al.  Transportin-mediated Nuclear Import of Heterogeneous Nuclear RNP Proteins , 1997, The Journal of cell biology.

[17]  T. Ho,et al.  The mechanism of inhibition of Ran-dependent nuclear transport by cellular ATP depletion , 2002, The Journal of cell biology.

[18]  G. Dreyfuss,et al.  Characterization and primary structure of the poly(C)-binding heterogeneous nuclear ribonucleoprotein complex K protein , 1992, Molecular and cellular biology.

[19]  L. Dember,et al.  Individual RNA Recognition Motifs of TIA-1 and TIAR Have Different RNA Binding Specificities (*) , 1996, The Journal of Biological Chemistry.

[20]  J. Raymond,et al.  Translational Control of β2-Adrenergic Receptor mRNA by T-cell-restricted Intracellular Antigen-related Protein* , 2005, Journal of Biological Chemistry.

[21]  P. Anderson,et al.  Stress granules: sites of mRNA triage that regulate mRNA stability and translatability. , 2002, Biochemical Society transactions.

[22]  R. Dohmen.  SUMO protein modification. , 2004, Biochimica et biophysica acta.

[23]  J. Steitz,et al.  Delineation of mRNA Export Pathways by the Use of Cell-Permeable Peptides , 2001, Science.

[24]  A. Shyu,et al.  RNA stabilization by the AU‐rich element binding protein, HuR, an ELAV protein , 1998, The EMBO journal.

[25]  K. Hilse,et al.  Binding of mRNA by an oligopeptide containing an evolutionarily conserved sequence from RNA binding proteins , 1989, FEBS Letters.

[26]  A. Plet,et al.  Multiple interdependent sequence elements control splicing of a fibroblast growth factor receptor 2 alternative exon , 1997, Molecular and cellular biology.

[27]  J. Valcárcel,et al.  The apoptosis-promoting factor TIA-1 is a regulator of alternative pre-mRNA splicing. , 2000, Molecular cell.

[28]  P. Anderson,et al.  Stress granule assembly is mediated by prion-like aggregation of TIA-1. , 2004, Molecular biology of the cell.

[29]  J. Keene,et al.  RNA recognition: towards identifying determinants of specificity. , 1991, Trends in biochemical sciences.

[30]  David E. Golan,et al.  Dynamic Shuttling of Tia-1 Accompanies the Recruitment of mRNA to Mammalian Stress Granules , 2000, The Journal of cell biology.

[31]  Wei Li,et al.  RNA-Binding Proteins Tia-1 and Tiar Link the Phosphorylation of Eif-2α to the Assembly of Mammalian Stress Granules , 1999, The Journal of cell biology.

[32]  H. Lou,et al.  U1 snRNP-Dependent Function of TIAR in the Regulation of Alternative RNA Processing of the Human Calcitonin/CGRP Pre-mRNA , 2003, Molecular and Cellular Biology.

[33]  G. Dreyfuss,et al.  Signal sequences that target nuclear import and nuclear export of pre-mRNA-binding proteins. , 1995, Cold Spring Harbor symposia on quantitative biology.

[34]  J. Toulmé,et al.  Mapping of a Minimal AU-rich Sequence Required for Lipopolysaccharide-induced Binding of a 55-kDa Protein on Tumor Necrosis Factor-α mRNA* , 1998, The Journal of Biological Chemistry.

[35]  P. Schwarzmaier,et al.  Importins fulfil a dual function as nuclear import receptors and cytoplasmic chaperones for exposed basic domains , 2002, The EMBO journal.

[36]  B. S. Baker,et al.  Isolation of RRM-type RNA-binding protein genes and the analysis of their relatedness by using a numerical approach , 1993, Molecular and cellular biology.

[37]  Nobutoshi Ito,et al.  Crystal structure at 1.92 Å resolution of the RNA-binding domain of the U1A spliceosomal protein complexed with an RNA hairpin , 1994, Nature.

[38]  R. Laskey,et al.  A polypeptide domain that specifies migration of nucleoplasmin into the nucleus , 1982, Cell.

[39]  B. Golden,et al.  RNA-binding domain of the A protein component of the U1 small nuclear ribonucleoprotein analyzed by NMR spectroscopy is structurally similar to ribosomal proteins. , 1991, Proceedings of the National Academy of Sciences of the United States of America.

[40]  M. Gaestel,et al.  Leptomycin B‐sensitive nuclear export of MAPKAP kinase 2 is regulated by phosphorylation , 1998, The EMBO journal.

[41]  D. Goldfarb,et al.  Yeast Nucleoporins Involved in Passive Nuclear Envelope Permeability , 2000, The Journal of cell biology.

[42]  K. Scharf,et al.  Cytoplasmic heat shock granules are formed from precursor particles and are associated with a specific set of mRNAs , 1989, Molecular and cellular biology.

[43]  V. Kruys,et al.  Identification of Ribosomal Proteins Specific to Higher Eukaryotic Organisms* , 2002, The Journal of Biological Chemistry.

[44]  U. Kutay,et al.  Nucleocytoplasmic transport: taking an inventory , 2003, Cellular and Molecular Life Sciences CMLS.

[45]  P. Anderson,et al.  Structure, tissue distribution and genomic organization of the murine RRM-type RNA binding proteins TIA-1 and TIAR. , 1996, Nucleic acids research.

[46]  G. Dreyfuss,et al.  Transcription-dependent and transcription-independent nuclear transport of hnRNP proteins. , 1991, Science.