Functional specificity of shuttling hnRNPs revealed by genome-wide analysis of their RNA binding profiles.

Nab2, Npl3, and Nab4/Hrp1 are essential RNA binding proteins of the shuttling hnRNP class that are required for the efficient export of mRNA. To characterize the in vivo transcript specificity of these proteins, we identified their mRNA binding partners using a microarray-based assay. Each of the three proteins was coimmunoprecipitated with many different mRNA transcripts. Interestingly, each protein exhibits preferential associations with a distinct set of mRNAs. Notably, some of these appear to denote specific functional classes. For example, the ribosomal protein mRNAs and other highly expressed transcripts significantly favor association with Npl3 over Nab2, and Nab4/Hrp1 is strongly enriched with transcripts required for amino acid metabolism. Significantly, nab4 mutants showed a striking, desensitized growth phenotype when exposed to amino acid stress conditions suggesting a biological consequence to the associations we observed. Supporting the hypothesis that these proteins display transcript specificity, we identified a unique 7-nucleotide sequence overrepresented in the transcripts highly associated with Nab2 and Nab4/Hrp1 using the REDUCE algorithm. Validating our approach, our bioinformatics analysis correctly identified the known binding site for Nab4/Hrp1. These specialized associations of the hnRNP proteins of Saccharomyces cerevisiae suggest the opportunity to regulate the processing of particular transcripts between transcription and translation.

[1]  H. Krebber,et al.  Yeast Shuttling SR Proteins Npl3p, Gbp2p, and Hrb1p Are Part of the Translating mRNPs, and Npl3p Can Function as a Translational Repressor , 2004, Molecular and Cellular Biology.

[2]  J. Steitz,et al.  Evidence for reassociation of RNA-binding proteins after cell lysis: implications for the interpretation of immunoprecipitation analyses. , 2004, RNA.

[3]  Steven E Brenner,et al.  Genome-wide analysis reveals an unexpected function for the Drosophila splicing factor U2AF50 in the nuclear export of intronless mRNAs. , 2004, Molecular cell.

[4]  G. Crooks,et al.  WebLogo: a sequence logo generator. , 2004, Genome research.

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

[6]  N. Gray,et al.  A novel role for shuttling SR proteins in mRNA translation. , 2004, Genes & development.

[7]  P. Brown,et al.  Extensive Association of Functionally and Cytotopically Related mRNAs with Puf Family RNA-Binding Proteins in Yeast , 2004, PLoS biology.

[8]  C. Guthrie,et al.  Identification of Lhp1p-associated RNAs by microarray analysis in Saccharomyces cerevisiae reveals association with coding and noncoding RNAs. , 2004, Proceedings of the National Academy of Sciences of the United States of America.

[9]  P. Brown,et al.  Widespread cytoplasmic mRNA transport in yeast: Identification of 22 bud-localized transcripts using DNA microarray analysis , 2003, Proceedings of the National Academy of Sciences of the United States of America.

[10]  M. Rosbash,et al.  Localization of nuclear retained mRNAs in Saccharomyces cerevisiae. , 2003, RNA.

[11]  M. Henry,et al.  The yeast hnRNP-like protein Hrp1/Nab4 sccumulates in the cytoplasm after hyperosmotic stress: a novel Fps1-dependent response. , 2003, Molecular biology of the cell.

[12]  E. Izaurralde,et al.  Genome‐wide analysis of nuclear mRNA export pathways in Drosophila , 2003, The EMBO journal.

[13]  A. Corbett,et al.  Domain Analysis of the Saccharomyces cerevisiaeHeterogeneous Nuclear Ribonucleoprotein, Nab2p , 2003, The Journal of Biological Chemistry.

[14]  A. Corbett,et al.  Domain analysis of the Saccharomyces cerevisiae heterogeneous nuclear ribonucleoprotein, Nab2p. Dissecting the requirements for Nab2p-facilitated poly(A) RNA export. , 2003, The Journal of biological chemistry.

[15]  H. Hieronymus,et al.  Genome-wide analysis of RNA–protein interactions illustrates specificity of the mRNA export machinery , 2003, Nature Genetics.

[16]  C. Guthrie,et al.  Expression of the essential mRNA export factor Yra1p is autoregulated by a splicing-dependent mechanism. , 2002, RNA.

[17]  S. Tenenbaum,et al.  Eukaryotic mRNPs may represent posttranscriptional operons. , 2002, Molecular cell.

[18]  M. Rosbash,et al.  T7 RNA polymerase-directed transcripts are processed in yeast and link 3' end formation to mRNA nuclear export. , 2002, RNA.

[19]  John D. Storey,et al.  Precision and functional specificity in mRNA decay , 2002, Proceedings of the National Academy of Sciences of the United States of America.

[20]  Temple F. Smith,et al.  Probabilistic prediction of Saccharomyces cerevisiae mRNA 3'-processing sites. , 2002, Nucleic acids research.

[21]  Lionel Minvielle-Sebastia,et al.  Dual requirement for yeast hnRNP Nab2p in mRNA poly(A) tail length control and nuclear export , 2002, The EMBO journal.

[22]  A. Kornblihtt,et al.  Alternative splicing: multiple control mechanisms and involvement in human disease. , 2002, Trends in genetics : TIG.

[23]  Xiaodong Cheng,et al.  Nab2p Is Required for Poly(A) RNA Export in Saccharomyces cerevisiae and Is Regulated by Arginine Methylation via Hmt1p* , 2002, The Journal of Biological Chemistry.

[24]  G. Dreyfuss,et al.  Messenger-RNA-binding proteins and the messages they carry , 2002, Nature Reviews Molecular Cell Biology.

[25]  H. Dohlman,et al.  Analysis of RGS proteins in Saccharomyces cerevisiae. , 2002, Methods in enzymology.

[26]  J. Darnell,et al.  Microarray Identification of FMRP-Associated Brain mRNAs and Altered mRNA Translational Profiles in Fragile X Syndrome , 2001, Cell.

[27]  M. Rosbash,et al.  Quality control of mRNA 3′-end processing is linked to the nuclear exosome , 2001, Nature.

[28]  S. Peltz,et al.  Nonsense-mediated mRNA decay in Saccharomyces cerevisiae. , 2001, Gene.

[29]  A. Krainer,et al.  Pre-mRNA splicing in the new millennium. , 2001, Current opinion in cell biology.

[30]  R. Tibshirani,et al.  Significance analysis of microarrays applied to the ionizing radiation response , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[31]  M. Rosbash,et al.  A block to mRNA nuclear export in S. cerevisiae leads to hyperadenylation of transcripts that accumulate at the site of transcription. , 2001, Molecular cell.

[32]  H. Bussemaker,et al.  Regulatory element detection using correlation with expression , 2001, Nature Genetics.

[33]  S. Tenenbaum,et al.  Identifying mRNA subsets in messenger ribonucleoprotein complexes by using cDNA arrays. , 2000, Proceedings of the National Academy of Sciences of the United States of America.

[34]  D. Botstein,et al.  Genomic expression programs in the response of yeast cells to environmental changes. , 2000, Molecular biology of the cell.

[35]  J. Derisi,et al.  Plasma membrane compartmentalization in yeast by messenger RNA transport and a septin diffusion barrier. , 2000, Science.

[36]  B. Graveley.  Sorting out the complexity of SR protein functions. , 2000, RNA.

[37]  M. Jia,et al.  Global expression profiling of yeast treated with an inhibitor of amino acid biosynthesis, sulfometuron methyl. , 2000, Physiological genomics.

[38]  C. Guthrie,et al.  A putative ubiquitin ligase required for efficient mRNA export differentially affects hnRNP transport , 2000, Current Biology.

[39]  M. Ashburner,et al.  Gene Ontology: tool for the unification of biology , 2000, Nature Genetics.

[40]  P. Silver,et al.  Uncoupling of the hnRNP Npl3p from mRNAs during the stress-induced block in mRNA export. , 1999, Genes & development.

[41]  M. Swanson,et al.  Control of cleavage site selection during mRNA 3′ end formation by a yeast hnRNP , 1998, The EMBO journal.

[42]  D. Botstein,et al.  Cluster analysis and display of genome-wide expression patterns. , 1998, Proceedings of the National Academy of Sciences of the United States of America.

[43]  Michael Ruogu Zhang,et al.  Comprehensive identification of cell cycle-regulated genes of the yeast Saccharomyces cerevisiae by microarray hybridization. , 1998, Molecular biology of the cell.

[44]  Michael R. Green,et al.  Dissecting the Regulatory Circuitry of a Eukaryotic Genome , 1998, Cell.

[45]  M. Rosbash,et al.  Nuclear RNA export. , 1998, Genes & development.

[46]  S. Chen,et al.  A specific RNA-protein interaction at yeast polyadenylation efficiency elements. , 1998, Nucleic acids research.

[47]  P. Philippsen,et al.  Additional modules for versatile and economical PCR‐based gene deletion and modification in Saccharomyces cerevisiae , 1998, Yeast.

[48]  M. Mann,et al.  A comprehensive biochemical and genetic analysis of the yeast U1 snRNP reveals five novel proteins. , 1998, RNA.

[49]  P. Brown,et al.  Exploring the metabolic and genetic control of gene expression on a genomic scale. , 1997, Science.

[50]  Marco M. Kessler,et al.  Hrp1, a sequence-specific RNA-binding protein that shuttles between the nucleus and the cytoplasm, is required for mRNA 3'-end formation in yeast. , 1997, Genes & development.

[51]  L. Minvielle-Sebastia,et al.  The major yeast poly(A)-binding protein is associated with cleavage factor IA and functions in premessenger RNA 3'-end formation. , 1997, Proceedings of the National Academy of Sciences of the United States of America.

[52]  C. Ball,et al.  Genetic and physical maps of Saccharomyces cerevisiae. , 1997, Nature.

[53]  P. Silver,et al.  A protein that shuttles between the nucleus and the cytoplasm is an important mediator of RNA export. , 1996, Genes & development.

[54]  P. Silver,et al.  Potential RNA binding proteins in Saccharomyces cerevisiae identified as suppressors of temperature-sensitive mutations in NPL3. , 1996, Genetics.

[55]  S. Chen,et al.  A yeast protein that bidirectionally affects nucleocytoplasmic transport. , 1995, Journal of cell science.

[56]  J. Swedlow,et al.  Characterization of nuclear polyadenylated RNA-binding proteins in Saccharomyces cerevisiae , 1994, The Journal of cell biology.

[57]  Charles Elkan,et al.  Fitting a Mixture Model By Expectation Maximization To Discover Motifs In Biopolymer , 1994, ISMB.

[58]  M. Swanson,et al.  NAB2: a yeast nuclear polyadenylated RNA-binding protein essential for cell viability , 1993, Molecular and cellular biology.

[59]  C. Burd,et al.  hnRNP proteins and the biogenesis of mRNA. , 1993, Annual review of biochemistry.

[60]  Janina Maier,et al.  Guide to yeast genetics and molecular biology. , 1991, Methods in enzymology.

[61]  T. D. Schneider,et al.  Sequence logos: a new way to display consensus sequences. , 1990, Nucleic acids research.