Characterization of nuclear polyadenylated RNA-binding proteins in Saccharomyces cerevisiae

To study the functions of heterogeneous nuclear ribonucleoproteins (hnRNPs), we have characterized nuclear polyadenylated RNA-binding (Nab) proteins from Saccharomyces cerevisiae. Nab1p, Nab2p, and Nab3p were isolated by a method which uses UV light to cross-link proteins directly bound to poly(A)+ RNA in vivo. We have previously characterized Nab2p, and demonstrated that it is structurally related to human hnRNPs. Here we report that Nab1p is identical to the Np13p/Nop3p protein recently implicated in both nucleocytoplasmic protein shuttling and pre-rRNA processing, and characterize a new nuclear polyadenylated RNA-binding protein, Nab3p. The intranuclear distributions of the Nab proteins were analyzed by three-dimensional immunofluorescence optical microscopy. All three Nab proteins are predominantly localized within the nucleoplasm in a pattern similar to the distribution of hnRNPs in human cells. The NAB3 gene is essential for cell viability and encodes an acidic ribonucleoprotein. Loss of Nab3p by growth of a GAL::nab3 mutant strain in glucose results in a decrease in the amount of mature ACT1, CYH2, and TPI1 mRNAs, a concomitant accumulation of unspliced ACT1 pre-mRNA, and an increase in the ratio of unspliced CYH2 pre-mRNA to mRNA. These results suggest that the Nab proteins may be required for packaging pre-mRNAs into ribonucleoprotein structures amenable to efficient nuclear RNA processing.

[1]  A. Krainer,et al.  General splicing factor SF2/ASF promotes alternative splicing by binding to an exonic splicing enhancer. , 1993, Genes & development.

[2]  M. Swanson,et al.  PUB1 is a major nuclear and cytoplasmic polyadenylated RNA-binding protein in Saccharomyces cerevisiae , 1993, Molecular and cellular biology.

[3]  I. Karsch-Mizrachi,et al.  The Rb97D gene encodes a potential RNA-binding protein required for spermatogenesis in Drosophila. , 1993, Nucleic acids research.

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

[5]  G. Dreyfuss,et al.  Association of individual hnRNP proteins and snRNPs with nascent transcripts , 1993, The Journal of cell biology.

[6]  R. Lin,et al.  Pre-mRNA splicing within an assembled yeast spliceosome requires an RNA-dependent ATPase and ATP hydrolysis. , 1993, Proceedings of the National Academy of Sciences of the United States of America.

[7]  M. Aebi,et al.  Nuclear PRP20 protein is required for mRNA export. , 1993, The EMBO journal.

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

[9]  J. Steitz,et al.  14 The Diverse World of Small Ribonucleoproteins , 1993 .

[10]  D. Tollervey,et al.  NOP3 is an essential yeast protein which is required for pre-rRNA processing , 1992, The Journal of cell biology.

[11]  R. Reed,et al.  Protein components specifically associated with prespliceosome and spliceosome complexes. , 1992, Genes & development.

[12]  M. Wickens,et al.  Defects in mRNA 3'-end formation, transcription initiation, and mRNA transport associated with the yeast mutation prp20: possible coupling of mRNA processing and chromatin structure. , 1992, Genes & development.

[13]  P. Silver,et al.  A mutant nuclear protein with similarity to RNA binding proteins interferes with nuclear import in yeast. , 1992, Molecular biology of the cell.

[14]  G. Dreyfuss,et al.  Differential binding of heterogeneous nuclear ribonucleoproteins to mRNA precursors prior to spliceosome assembly in vitro. , 1992, Molecular and cellular biology.

[15]  G. Dreyfuss,et al.  Shuttling of pre-mRNA binding proteins between nucleus and cytoplasm , 1992, Nature.

[16]  Adrian R. Krainer,et al.  Regulation of alternative pre-mRNA splicing by hnRNP A1 and splicing factor SF2 , 1992, Cell.

[17]  C. Guthrie.  Messenger RNA splicing in yeast: clues to why the spliceosome is a ribonucleoprotein. , 1991, Science.

[18]  S. Ruby,et al.  Pre-mRNA splicing in yeast. , 1991, Trends in genetics : TIG.

[19]  C. Guthrie,et al.  PRP16 is an RNA-dependent ATPase that interacts transiently with the spliceosome , 1991, Nature.

[20]  J. Keene,et al.  Nuclear RNA-binding proteins. , 1991, Progress in nucleic acid research and molecular biology.

[21]  M. Green,et al.  Biochemical mechanisms of constitutive and regulated pre-mRNA splicing. , 1991, Annual review of cell biology.

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

[23]  T. Shenk,et al.  A uridylate tract mediates efficient heterogeneous nuclear ribonucleoprotein C protein-RNA cross-linking and functionally substitutes for the downstream element of the polyadenylation signal , 1990, Molecular and cellular biology.

[24]  E. Myers,et al.  Basic local alignment search tool. , 1990, Journal of molecular biology.

[25]  David A. Agard,et al.  Interphase nuclear envelope lamins form a discontinuous network that interacts with only a fraction of the chromatin in the nuclear periphery , 1990, Cell.

[26]  R. Lin,et al.  Preparation and fractionation of yeast splicing extract. , 1990, Methods in enzymology.

[27]  G. Dreyfuss,et al.  A novel heterogeneous nuclear RNP protein with a unique distribution on nascent transcripts , 1989, The Journal of cell biology.

[28]  M. Company,et al.  Isolation and characterization of pre-mRNA splicing mutants of Saccharomyces cerevisiae. , 1989, Genes & development.

[29]  R. Sikorski,et al.  A system of shuttle vectors and yeast host strains designed for efficient manipulation of DNA in Saccharomyces cerevisiae. , 1989, Genetics.

[30]  G. Dreyfuss,et al.  RNA-binding proteins as developmental regulators. , 1989, Genes & development.

[31]  R. Sperling,et al.  Isolation and visualization of large compact ribonucleoprotein particles of specific nuclear RNAs. , 1989, Proceedings of the National Academy of Sciences of the United States of America.

[32]  B. Wold,et al.  Isolation and characterization of a Xenopus laevis C protein cDNA: structure and expression of a heterogeneous nuclear ribonucleoprotein core protein. , 1988, Proceedings of the National Academy of Sciences of the United States of America.

[33]  G. Dreyfuss,et al.  RNA binding specificity of hnRNP proteins: a subset bind to the 3′ end of introns. , 1988, The EMBO journal.

[34]  G. Blobel,et al.  Identification and characterization of a yeast nucleolar protein that is similar to a rat liver nucleolar protein , 1988, The Journal of cell biology.

[35]  G. Dreyfuss,et al.  Classification and purification of proteins of heterogeneous nuclear ribonucleoprotein particles by RNA-binding specificities , 1988, Molecular and cellular biology.

[36]  G. Dreyfuss,et al.  Immunopurification of heterogeneous nuclear ribonucleoprotein particles reveals an assortment of RNA-binding proteins. , 1988, Genes & development.

[37]  G. Dreyfuss,et al.  Primary structure of human nuclear ribonucleoprotein particle C proteins: conservation of sequence and domain structures in heterogeneous nuclear RNA, mRNA, and pre-rRNA-binding proteins , 1987, Molecular and cellular biology.

[38]  H. Bourbon,et al.  Nucleolin, the major nucleolar protein of growing eukaryotic cells: an unusual protein structure revealed by the nucleotide sequence. , 1987, Proceedings of the National Academy of Sciences of the United States of America.

[39]  Gerald R. Fink,et al.  Methods in Yeast Genetics: A Laboratory Course Manual , 1987 .

[40]  Halina Sierakowska,et al.  Antibodies to hnRNP core proteins inhibit in vitro splicing of human beta-globin pre-mRNA , 1986, Nucleic Acids Res..

[41]  G. Dreyfuss,et al.  Heterogeneous nuclear ribonucleoproteins: role in RNA splicing. , 1986, Science.

[42]  J. Wooley,et al.  Set of novel, conserved proteins fold pre‐messenger RNA into ribonucleosomes , 1986, Proteins.

[43]  G. Dreyfuss.  Structure and function of nuclear and cytoplasmic ribonucleoprotein particles. , 1986, Annual review of cell biology.

[44]  R. Lin,et al.  Yeast mRNA splicing in vitro. , 1985, The Journal of biological chemistry.

[45]  F. Sherman,et al.  A mutation allowing an mRNA secondary structure diminishes translation of Saccharomyces cerevisiae iso-1-cytochrome c , 1985, Molecular and cellular biology.

[46]  R. Kornberg,et al.  Abundant nuclear ribonucleoprotein form of CAD RNA , 1985, Molecular and cellular biology.

[47]  G. Dreyfuss,et al.  Isolation of the heterogeneous nuclear RNA-ribonucleoprotein complex (hnRNP): a unique supramolecular assembly. , 1984, Proceedings of the National Academy of Sciences of the United States of America.

[48]  J. Warner,et al.  Cycloheximide resistance in yeast: the gene and its protein. , 1983, Nucleic acids research.

[49]  T. Alber,et al.  Nucleotide sequence of the triose phosphate isomerase gene of Saccharomyces cerevisiae. , 1982, Journal of molecular and applied genetics.

[50]  J. Abelson,et al.  Isolation and sequence of the gene for actin in Saccharomyces cerevisiae. , 1980, Proceedings of the National Academy of Sciences of the United States of America.

[51]  D. Gallwitz,et al.  Structure of a split yeast gene: complete nucleotide sequence of the actin gene in Saccharomyces cerevisiae. , 1980, Proceedings of the National Academy of Sciences of the United States of America.

[52]  G. Fink,et al.  Methods in yeast genetics , 1979 .

[53]  J. Vlak,et al.  Nucleolar function of the dense crescent in the yeast nucleus. A biochemical and ultrastructural study. , 1973, Experimental cell research.