Characterization of Pre‐messenger‐RNA‐Containing Nuclear Ribonucleoprotein Particles from Avian Erythroblasts

Maundrell, Kinsey; Scherrer, Klaus

doi:10.1111/j.1432-1033.1979.tb13249.x

Cited by 59 publications

(40 citation statements)

References 54 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…This protein, abundant in polyribosomes of selected young erythroblasts, is thus possibly associated with only a subfraction of ribosomes. It is interesting to recall that a similar protein is detected in small nRNP particles [47] as well as in free cytoplasmic complexes 1141. An interesting speculation might attribute to this polypeptide a role in transport of mRNA from the nucleus to the cytoplasm.…”

Section: Discussionmentioning

confidence: 99%

Comparisons of Proteins Associated with Duck‐Globin mRNA and Its Polyadenylated Segment in Polyribosomal and Repressed Free Messenger Ribonucleoprotein Complexes

Víncent

Goldenberg

Scherrer

1981

European Journal of Biochemistry

Self Cite

View full text Add to dashboard Cite

EDTA dissociation of polyribosomes from duck erythroblasts allowed us to isolate the 15‐S globin messenger ribonucleoproteins (mRNP) by sucrose gradient centrifugation or affinity chromatography on poly(U)‐Sepharose or oligo(dT)‐ce1lu1ose columns. Their protein composition was compared by one and two‐dimensional electrophoresis in sodium dodecyl sulfate to the free 20‐S mRNP containing the repressed fraction of globin mRNA [Vincent, A., Civelli, O., Maundrell, K., and Scherrer, K. (1980) Eur. J. Biochem. 112, 617–633]. The protein composition of the 15‐S mRNP isolated by these methods in different ionic strength conditions, was characterized by a major 73000‐Mr polypeptide and seven minor polypeptides with Mr ranging from 45000 to 68000, all of which are slightly basic, and about five acidic ones in the 80000–130000‐Mr range. All these are retained in the 15‐S mRNP core particle isolated at 0.5 M KCl. At low ionic strength, in addition, a specific group of acidic polypeptides in the Mr range 35000–105000 was also found associated with globin mRNA. Oligo(dT)‐cellulose chromatography of mRNP digested with ribonucleases A and T1 indicated that the 73000‐Mr major protein is bound to the poly(A) segment; some other proteins resolved as minor components interact with both the poly(A) and non‐poly(A) regions of globin mRNA. Characterization of proteins interacting with the poly(A) segment of non‐polyribosomal globin mRNA in 20‐S free mRNP demonstrated the absence of the polyribosomal 73000‐Mr poly(A)‐binding protein. Furthermore, it confirmed that the protein compositions of translatable polyribosomal and repressed free globin mRNP are very different. Indeed, the respective core (0.5 M KCl) particles contain only two possibly common polypeptides. The specificity of proteins associated with globin mRNA in two different functional states shown here supports the hypothesis of a role of mRNP proteins in translational control of mRNA.

show abstract

Section: Discussionmentioning

confidence: 99%

Comparisons of Proteins Associated with Duck‐Globin mRNA and Its Polyadenylated Segment in Polyribosomal and Repressed Free Messenger Ribonucleoprotein Complexes

Víncent

Goldenberg

Scherrer

1981

European Journal of Biochemistry

Self Cite

View full text Add to dashboard Cite

show abstract

“…servations (Samarina et al 1968;Martin et al 1974Martin et al , 1978Pederson 1974;Beyer et al 1977;Karn et al 1977;Maundrell and Scherrer 1979;Walker et al 1980;Le Stourgeon et al 1981;Steitz and Kamen 1981;Lothstein et al 1985;Wilk et al 1985), hnRNA sediments in sucrose gradients as heterogeneous material of greater than 30S and, after mild digestion with nuclease, as more homogeneous material corresponding to monoparticles at about 30S. Heparin converts the hnRNP particles to slightly slower sedimenting material, but it is evident that particles sedimenting only slightly slower than the control remain.…”

Section: Heparin-resistan T Hnrnp Particlesmentioning

confidence: 99%

Immunopurification of heterogeneous nuclear ribonucleoprotein particles reveals an assortment of RNA-binding proteins.

Piñol-Roma¹,

Choi²,

Matunis³

et al. 1988

Genes Dev.

410

405

View full text Add to dashboard Cite

Heterogeneous nuclear RNA-ribonucleoprotein (hnRNP) particles can be efficiently purifed by a specific, rapid, and mild procedure using monoclonal antibodies to hnRNP proteins. We report here on the detailed analysis of the protein composition of immunopurified hnRNP particles from human HeLa cells. By two-dimensional gel electrophoresis, immunopurified hnRNP particles contain at least 24 polypeptides in the range of 34,000-120,000 daltons. The abundant 30,000-40,000 dalton proteins, A, B, and C, described previously, are a subset of these polypeptides. The protein compositions of hnRNP particles found in the nucleoplasm fraction and in the chromatin-nucleolar fraction are very similar. Upon addition of the polyanion heparin, most of the major proteins remain associated in heparin-resistant particles, and only several, mostly minor, proteins dissociate. This provides an aid in the classification of the proteins and an additional criterion for the definition of hnRNP particle components. Chromatography on single-stranded DNA (ssDNA)-agarose in a heparin-and moderate or high salt (higher than 300 mM NaCl)-resistant manner suggests that most, if not all, of these proteins are single-stranded nucleic acid-binding proteins. We describe a general method for the large-scale purification of hnRNP proteins by affinity chromatography on ssDNA columns and its use for the production of new monoclonal antibodies to hnRNP proteins.

show abstract

“…The relative amounts of the 41K and 43K proteins are constant in many different cellular and biochemical fractions, and they therefore appear to behave like two subunit polypeptides of one multisubunit protein or of a larger structure. The hnRNP 120K protein has not been previously described, although proteins of high molecular weight associated with hnRNA in vitro (7,22,25,31,36,47,49,50) and in vivo (13,14,39,52) have been identified. It is shown to be a genuine hnRNP protein because it is cross-linked to highmolecular-weight nonnucleolar nuclear RNA in vivo.…”

mentioning

confidence: 99%

Characterization of heterogeneous nuclear RNA-protein complexes in vivo with monoclonal antibodies.

Dreyfuss¹,

Choi²,

1984

View full text Add to dashboard Cite

Exposure of cells to UV light of sufficient intensity brings about cross-linking of RNA to proteins which are in direct contact with it in vivo. The major [35S]methionine-labeled proteins which become cross-linked to polyadenylated heterogeneous nuclear RNA in HeLa cells have molecular weights of 120,000 (120K), 68K, 53K, 43K, 41K, 38K, and 36K. Purified complexes of polyadenylated RNA with proteins obtained by UV cross-linking in intact cells were used to immunize mice and generate monoclonal antibodies to several of these proteins. Some properties of three of the proteins, 41K, 43K, and 120K, were characterized with these antibodies. The 41K and 43K polypeptides are highly related. They were recognized by the same antibody (2B12) and have identical isoelectric points (pl = 6.0 + 0.2) but different partial peptide maps. The 41K and 43K polypeptides were part of the 40S heterogeneous nuclear ribonucleoprotein particle and appear to correspond to the previously described C proteins (Beyer et al.. Cell 11:127-138, 1977). A different monoclonal antibody (3G6) defined a new major heterogeneous ribonucleoprotein of 120K. The 41K, 43K, and 120K polypeptides were associated in vivo with both polyadenylated and non-polyadenylated nuclear RNA, and all three proteins were phosphorylated. The monoclonal antibodies recognized similar proteins in human and monkey cells but not in several other vertebrates. Immunofluorescence microscopy demonstrated that these proteins are segregated to the nucleus, where they are part of a fine particulate nonnucleolar structure. In cells extracted in situ with nonionic detergent. all of the 41K and 43K polypeptides were associated with the nucleus at salt concentrations up to 0.5 M NaCl, whereas the 120K polypeptide was completely extracted at this NaCl concentration. A substantial fraction of the 41K and 43K polypeptides (up to 40%) was retained with a nuclear matrix-a structure which is resistant to digestion with DNase I and to extraction by 2 M NaCl, but the 41K and 43K polypeptides were quantitatively removed at 0.5 M NaCl after digestion with RNase.The proteins which are associated with heterogeneous nuclear RNA (hnRNA) to form ribonucleoprotein complexes (hnRNPs) are thought to be involved in the packaging and processing of hnRNA (for a review, see reference 35). Current models of hnRNP structure based primarily on morphological (5. 6. 8. 15. 28. 40, 41) and sedimentation (7,22, 33,34,44,46,55) studies of material released from nuclei by RNase digestion (22, 33, 34. 46. 55) or by mild sonication (7,22,44) (25, 36, 44, 47. 50

show abstract

Characterization of Pre‐messenger‐RNA‐Containing Nuclear Ribonucleoprotein Particles from Avian Erythroblasts

Cited by 59 publications

References 54 publications

Comparisons of Proteins Associated with Duck‐Globin mRNA and Its Polyadenylated Segment in Polyribosomal and Repressed Free Messenger Ribonucleoprotein Complexes

Comparisons of Proteins Associated with Duck‐Globin mRNA and Its Polyadenylated Segment in Polyribosomal and Repressed Free Messenger Ribonucleoprotein Complexes

Immunopurification of heterogeneous nuclear ribonucleoprotein particles reveals an assortment of RNA-binding proteins.

Characterization of heterogeneous nuclear RNA-protein complexes in vivo with monoclonal antibodies.

Contact Info

Product

Resources

About