Proteins associated with poly(A) and other regions of mRNA and hnRNA molecules as investigated by crosslinking

Setyono, Buddy; Greenberg, Jay

doi:10.1016/0092-8674(81)90103-3

Cited by 101 publications

(68 citation statements)

References 31 publications

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“…The immunofluorescence microscopy data on the C proteins, the 120,000-mol-wt hnRNP protein (12), and the A and B proteins (19,27,31) indicate that the major hnRNP proteins so far examined are segregated to the nucleus. The RNA transcripts that emerge in the cytoplasm must, therefore, exchange the major proteins with which it is associated, as previously suggested (12,25,42) i n d i c a t e t h a t t h e C p r o t e i n s a n d p o s s i b l y a l s o t h e o t h e r m a j o r h n R N P p r o t e i n s a r e n o t c h r o m a t i n p r o t e i n s a l t h o u g h t h e y a r e l i k e l y t o b e a s s o c i a t e d w i t h a c t i v e l y t r a n s c r i b i n g r e g i o n s t h r o u g h a s s o c i a t i o n w i t h n a s c e n t h n R N P t r a n s c r i p t s .…”

Section: Discussionmentioning

confidence: 90%

Monoclonal antibody characterization of the C proteins of heterogeneous nuclear ribonucleoprotein complexes in vertebrate cells.

Choi¹,

Dreyfuss²

1984

The Journal of Cell Biology

198

145

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The C proteins (C1 and C2) are major constituents of the 40S subparticle of heterogeneous nuclear ribonucleoprotein complexes (hnRNPs) (Beyer, A. L., M. E. Christensen, B. W. Walker, and W. M. LeStourgeon, 1977, Cell, 11:127-138) and are two of the most prominent proteins that become cross-linked by ultraviolet light to heterogeneous nuclear RNA (hnRNA) in vivo. Studies are described here on the characterization of the C proteins in vertebrate cells using monoclonal and polyclonal antibodies. Monoclonal antibodies to genuine RNP proteins, including the C proteins, were obtained by immunizing mice with purified complexes of poly(A)+hnRNA and poly(A)+mRNA with their contacting proteins in vivo obtained by ultraviolet cross-linking the complexes in intact cells (Dreyfuss, G., Y. D. Choi, and S. A. , Mol. Cell. Biol., 4:1104-1114. One of the monoclonal antibodies identified the C proteins in widely divergent species ranging from human to lizard. In all species examined, there were two C proteins in the molecular weight range of from 39,000 to 42,000 for C1, and from 40,000 to 45,000 for C2. The two C proteins were found to be highly related to each other; they were recognized by the same monoclonal antibodies and antibodies raised against purified C~ reacted also with C2. In avian, rodent, and human cells the C proteins were phosphorylated and were in contact with hnRNA in vivo. Immunofluorescence microscopy demonstrated that the C proteins are segregated to the nucleus. Within the nucleus the C proteins were not found in nucleoli and were not associated with chromatin as seen in cells in prophase. These findings demonstrate that C proteins with similar characteristics to those in humans are ubiquitous components of hnRNPs in vertebrates.Heterogeneous nuclear RNAs (hnRNAs), ~ the nuclear transcripts of which some are precursors to mRNAs, are associated in the cell with specific proteins to form heterogeneous nuclear ribonucleoprotein complexes (hnRNPs). The hnRNPs have been studied by both cytological (5, 7, 14, 37, 38) and biochemical (6, 10, 12, 13, 16, 20, 22, 25, 28-30, 32, 33, 35, 36, 39-42, 44--46, 48-50) approaches. These studies have revealed a major structural component of hnRNPs--the 40S particles. The major proteins of the 40S particles so far described are classified into three doublets: the A, B, and C groups (6). The C group proteins are the two proteins (by one-dimensional SDS PAGE) that appear to be the proteins most tightly associated with the hnRNA in vitro of the 40S

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Section: Discussionmentioning

confidence: 90%

Monoclonal antibody characterization of the C proteins of heterogeneous nuclear ribonucleoprotein complexes in vertebrate cells.

Choi¹,

Dreyfuss²

1984

The Journal of Cell Biology

198

145

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“…In this respect, the nucleocytoplasmic transport of ribosomal precursors seems to be similar to that of nuclear ribonucleoprotein particles containing mRNA sequences; drastic changes in protein composition during migration from nucleus to cytoplasm have been reported for those particles (e.g ., Martin et aI. , 1980;Pederson and Munroe, 1981 ;Setyono and Greenberg, 1981).…”

Section: Discussionmentioning

confidence: 99%

Ribocharin: a nuclear Mr 40,000 protein specific to precursor particles of the large ribosomal subunit

Hügle

Scheer

Franke

1985

Cell

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SummaryUsing a monoclonal antibody (No-194) we have identified, in Xenopus laevis and other amphibia, an acidic protein of M, 40,000 (ribocharin) which is specifically associated with the granular component of the nucleolus and nucleoplasmic 65S particles. These particles contain the nuclear 28S rRNA and apparently represent the precursor to the large ribosomal subunit in nucleocytoplasmic transit. By immunoelectron microscopy ribocharin has been localized in the granular component of the nucleolus and in interchromatin granules. During mitosis ribocharin-containing particles are associated with surfaces of chromosomes and are recollected in the reconstituting nucleoli in late telophase. We suggest that ribocharin is a specific component of precursor particles of the large ribosomal subunit, which dissociates from the 65S particle before passage through the nuclear envelope, and is reutilized in ribosome biogenesis.

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“…The RNA-protein ultraviolet-crosslinking method recently developed by Greenberg and coworker [20][21][22][23] Fig. 2.…”

Section: Resultsmentioning

confidence: 99%

“…Samples of 25 /tl contained: 1/tg 32p-labeled RNA (2.10 6 cpm)/10-20 /~g initiation factors/20 mM Tris-HC1 (pH 7.6)/ 100 mM KCI/3 mM MgAc2/7 mM 2-mercaptoethanol/10% glycerol. Irradiation at 254 nm was carried out by placing the samples under two 25 W germicidal tubes at a radiation intensity of 2000/~W/cm 2, at 0°C, for 8 min, as described previously [20,21,23]. The samples were then treated with ribonuclease T 2 and ribonuclease A for 1 h at 37°C [20][21][22][23].…”

Section: Methodsmentioning

confidence: 99%

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Ultraviolet-crosslinking reveals specific affinity of eukaryotic initiation factors for Semliki Forest virus mRNA

Setyono¹,

Steeg²,

Voorma³

1984

Biochimica et Biophysica Acta (BBA) - Gene Structure and Expres

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Eukaryotic initiation factors (elF) associate readily with 32 P-labeled Semliki Forest virus (SFV) mRNA in vitro, forming complexes which can be crosslinked by 254 nm ultraviolet irradiation. After ribonuclease digestion, the initiation factors were released and analysed by gel electrophoresis. Autoradiography revealed proteins by virtue of crosslinked 32 P-labeled mRNA fragments, elF-4A, -4B and -4C as well as three subunits of elF-3 could be crosslinked with SFV mRNA. None of these proteins bound to ribosomal RNAs.

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Proteins associated with poly(A) and other regions of mRNA and hnRNA molecules as investigated by crosslinking

Cited by 101 publications

References 31 publications

Monoclonal antibody characterization of the C proteins of heterogeneous nuclear ribonucleoprotein complexes in vertebrate cells.

Monoclonal antibody characterization of the C proteins of heterogeneous nuclear ribonucleoprotein complexes in vertebrate cells.

Ribocharin: a nuclear Mr 40,000 protein specific to precursor particles of the large ribosomal subunit

Ultraviolet-crosslinking reveals specific affinity of eukaryotic initiation factors for Semliki Forest virus mRNA

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