Specific poly-A-binding protein of 76,000 molecular weight in polyribosomes is not present on poly A of free cytoplasmic mRNP

Venrooij, Walther J. van; Eekelen, Chris A.G. van; Jansen, Rob; Princen, H.M.G.

doi:10.1038/270189a0

Cited by 65 publications

(46 citation statements)

References 31 publications

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“…The set of proteins present in CmRNP but absent in PmRNP, whrch we have observed m a varrety of cell types, may regulate in vrvo the entry of mRNA into the polysomes. With regard to the possible role of the P78 component, it has been suggested that rt is involved m the transport of mRNA [ 151, and rt remains associated with mRNA only during translation [6]. More recently we have demonstrated that P78 is associated with the 3'-poly(A) tracts of the RNA moieties of thermally eluted CmRNP and PmRNP of chick embyronic muscles [27] .…”

Section: Resultsmentioning

confidence: 99%

“…The A& A 2s0 ratros of these fractions were 1.40-l .45 (for CmRNP) and 1.55-1.65 (for PmRNP). The remaming 5-10% of the bound material was eluted at 25'C with 50% formamide m low salt buffer, an eluant used for the isolation of eukaryotrc hnRNP [ 121 and PmRNP [6,12] . When subcellular fractrons prepared from embryos pulse-labeled with [jH]adenosine were chromatographed, the relative distnbutron of counts in the 4 fractions gave the same pattern as shown m fig 1 Moreover, rechromatography on ohgo(dT)-cellulose of the deprotemrzed labeled 45'C fractions showed that about 95% of the mitral counts could be rebound to and then eluted from the column at 4°C with the low salt buffer, as would be expected of typical poly(A)'RNA.…”

Section: Resultsmentioning

confidence: 99%

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Thermal chromatography of eukaryotic messenger ribonucleoprotein particles on oligo(dT)‐cellulose

1979

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Thermal chromatography of eukaryotic messenger ribonucleoprotein particles on oligo(dT)‐cellulose

1979

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“…Partial purification of the fusion protein, as described in Materials and Methods, yielded a preparation which was approximately 90% pure, as determined by Coomassie blue staining of an SDS gel. The about 60,000, which is restricted to the poly(A) tails of heterogeneous nuclear RNA (41,45), and a 72,000-molecular-weight mRNA poly(A)-binding protein in the cytoplasm (7,13,14,17,18,24,27,44,45,52,53). Indirect immunofluorescence on yeast cells with antibodies to the poly(A)-binding protein, produced by immunizing a mouse with the fusion protein of XYPA72.1 (Fig.…”

Section: Resultsmentioning

confidence: 99%

mRNA polyadenylate-binding protein: gene isolation and sequencing and identification of a ribonucleoprotein consensus sequence.

Adam¹,

Nakagawa²,

Swanson³

et al. 1986

Mol. Cell. Biol.

450

258

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We identified and produced antibodies to the major proteins that interact with poly(A)+ RNAs in the yeast Saccharomyces cerevisiae. The major proteins which were cross-linked by UV light to poly(A)+ RNA in intact yeast cells had apparent molecular weights of 72,000, 60,000, and 50,000. The poly(A) segment of the RNA was selectively cross-linked to the 72,000-molecular-weight protein (72K protein). Mice immunized with purified UV-cross-linked RNA-protein (RNP) complexes produced antibodies to the three major RNP proteins. A yeast genomic DNA library constructed in the Agtll expression vector was screened with the anti-RNP serum, and recombinant bacteriophage clones were isolated. One recombinant phage, XYPA72.1, bearing a 2.5-kilobase insert, produced a large ,I-galactosidase-RNP fusion protein. mRNA and heterogeneous nuclear RNA are found in eucaryotic cells in association with specific proteins to form RNA-protein complexes known as messenger ribonucleoprotein (RNP) particles (mRNPs) and heterogeneous nuclear ribonucleoprotein particles (hnRNPs), respectively. The complexes are the structural units of these polynucleotides and are likely to be involved in mRNA biogenesis, metabolism, and function (11,13,32). Therefore, the proteins which form hnRNP and mRNP complexes are of considerable interest and have been extensively studied (1, 11-13, 24, 48). Despite a great deal of effort, little is known about the biochemical functions of RNP proteins. Until recently, it has been difficult to identify unambiguously the genuine RNP proteins since nonspecific RNA-protein interactions are likely to occur during cell fractionation. UV cross-linking of proteins to RNA in intact cells overcomes these difficulties and allows the identification of proteins which bind heterogeneous nuclear RNA and mRNA in vivo (11,16,51,54). To learn more about these proteins and about the RNP complexes, we have immunized mice with purified UV-crosslinked RNP complexes and have produced antibodies to several of the major RNP proteins in vertebrate cells (1,12,13).Much of the knowledge of RNP proteins has come from work with higher eucaryotes (12,24,48 (7) and is one of the most conserved and extensively investigated eucaryotic RNP proteins. Although many important functions in mRNA metabolism have been attributed to this protein, little is known about its function, amino acid sequence, and metabolism. This is because specific probes, such as antibodies or gene sequences, have not been available from any species.We report here the identification of the proteins that interact with poly(A)+ RNAs in yeast by UV cross-linking and the production of antibodies to the proteins. Using these antibodies, we identified the poly(A)-binding protein in yeast

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“…This particular point, already discussed in a preliminary report [17], has also been reported in the case of mRNP from rabbit reticulocytes and ascites tumor cells isolated by chromatography on oligo (dT)-cellulose columns [13]. This protein, with a M , ranging from 73 000 to 80000 depending upon the authors, has been identified in polyribosomal mRNP isolated from different cellular systems (see review in [21]) and was shown to interact with the poly (A) segment of mRNA [18,201.…”

Section: Discussionmentioning

confidence: 79%

“…It is interesting to note that particles with identical protein composition are also found in polyribosomal mRNP preparations (Vincent, Goldenberg, and Scherrer, unpublished results). Moreover, a similar group of proteins has been observed in free globin mRNP isolated from rabbit reticulocytes [13,47].…”

Section: Discussionmentioning

confidence: 84%

Identification and Characterization of the Translationally Repressed Cytoplasmic Globin Messenger‐Ribonucleoprotein Particles from Duck Erythroblasts

Víncent

Civelli

Maundrell

et al. 1980

European Journal of Biochemistry

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Globin messenger ribonucleoprotein (mRNP) particles which have been isolated from duck erythroblast post-polyribosomal supernatant are translationally inactive in vivo and in uitro but contain translatable mRNA active after deproteinisation. They were characterized following pruification by successive sucrose gradient sedimentation in a buffer containing 0.05 M KCl. The complex, which sediments homogeneously at about 20 S, has a density of 1.39 g/cm3 and thus consists of four parts protein to one part RNA; 40% of this RNA is globin mRNA and no other mRNA could be detected.Sedimentation of the purified globin mRNP on sucrose gradients in 0.5 M KCl produced four components while polyacrylamide gel electrophoresis in non-denaturing conditions and in the presence of EDTA resulted in the separation of three components. Hybridization to globin cDNA and translation in vitro of the RNA extracted from these subparticles revealed the existence of two core particles containing globin mRNA with nominal sedimentation coefficients of 13 S and 16 s.Analysis of the protein components of the isolated sub-complexes by dodecyl sulfate and bidimensional gel electrophoresis indicated a very characteristic protein composition for each of these complexes. The 16-S and 13-S globin mRNPs differed essentially by the presence in the 13-S mRNP only of a group of major polypeptides. Of the other two sub-complexes, one consisted of 90% small RNA in the 4-S range; the second sedimented ahead of the globin mRNP core particles at about 19s and consisted of a very characteristic set of about 14 polypeptides. The polyrisobomal73OOO-M, poly(A)-binding protein was not detected in the purified free globin mRNP although the mRNA in the untranslatable particle is polyadenylated.The presence in the cytoplasm of duck erythroblasts of two forms of untranslated globin messenger ribonucleoprotein particles, distinct in their protein composition from polyribosomal globin mRNP. suggests that they may have a specific role in the regulation of translation of globin mRNA.

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Specific poly-A-binding protein of 76,000 molecular weight in polyribosomes is not present on poly A of free cytoplasmic mRNP

Cited by 65 publications

References 31 publications

Thermal chromatography of eukaryotic messenger ribonucleoprotein particles on oligo(dT)‐cellulose

Thermal chromatography of eukaryotic messenger ribonucleoprotein particles on oligo(dT)‐cellulose

mRNA polyadenylate-binding protein: gene isolation and sequencing and identification of a ribonucleoprotein consensus sequence.

Identification and Characterization of the Translationally Repressed Cytoplasmic Globin Messenger‐Ribonucleoprotein Particles from Duck Erythroblasts

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