scholarly journals Recognition of subsets of the mammalian A/B-type core heterogeneous nuclear ribonucleoprotein polypeptides by novel autoantibodies

1996 ◽  
Vol 320 (3) ◽  
pp. 761-767 ◽  
Author(s):  
Amalia DANGLI ◽  
Angeliki PLOMARITOGLOU ◽  
Efrosini BOUTOU ◽  
Neratzoula VASSILIADOU ◽  
Haralampos M. MOUTSOPOULOS ◽  
...  
Keyword(s):  
Rna Binding ◽  
Rna Binding Proteins ◽  
Peptide Mapping ◽  
Protein A ◽  
Major Protein ◽  
Protein Species ◽  
Two Dimensional Gel Electrophoresis ◽  
Heterogeneous Nuclear Ribonucleoprotein ◽  
Core Proteins ◽  
Nuclear Ribonucleoprotein

The structurally related A/B-type core heterogeneous nuclear ribonucleoprotein (hnRNP) polypeptides of 34–39 kDa (A1, A2, B1 and B2) belong to a family of RNA-binding proteins that are major components of 40 S hnRNP complexes. By two-dimensional gel electrophoresis and peptide mapping analysis we compared each member of the A/B-type core proteins in the human and rat liver cells. This comparison revealed the unique presence in rat cells of major protein species, referred to as mBx polypeptides, that appeared as three charge isoforms at a position corresponding to the minor HeLa B1b protein spot. In addition, clear differences in the ratios of the A1 polypeptide to the A1b isoform were observed. The detection, in sera of patients with rheumatic autoimmune diseases, of two novel autoantibody specificities, one recognizing solely B2 protein and the second both the B2 and mBx polypeptides, helped to identify mBx proteins as new A/B-type hnRNP components, immunologically related to B2 protein. A common immunoreactive V8 protease peptide of approx. 17 kDa has been identified in B2 and mBx hnRNP polypeptides. mBx protein species are identified in cells of murine origin, and have a ubiquitous tissue distribution and developmental appearance.

scholarly journals The C-protein tetramer binds 230 to 240 nucleotides of pre-mRNA and nucleates the assembly of 40S heterogeneous nuclear ribonucleoprotein particles.

1994 ◽  
Vol 14 (1) ◽  
pp. 518-533 ◽  
Author(s):  
M Huang ◽  
J E Rech ◽  
S J Northington ◽  
P F Flicker ◽  
A Mayeda ◽  
...  
Keyword(s):  
Rna Binding ◽  
Binding Studies ◽  
C Protein ◽  
Heterogeneous Nuclear Ribonucleoprotein ◽  
Core Proteins ◽  
C Proteins ◽  
Assembly Intermediate ◽  
Nuclear Ribonucleoprotein

A series of in vitro protein-RNA binding studies using purified native (C1)3C2 and (A2)3B1 tetramers, total soluble heterogeneous nuclear ribonucleoprotein (hnRNP), and pre-mRNA molecules differing in length and sequence have revealed that a single C-protein tetramer has an RNA site size of 230 to 240 nucleotides (nt). Two tetramers bind twice this RNA length, and three tetramers fold monoparticle lengths of RNA (700 nt) into a unique 19S triangular complex. In the absence of this unique structure, the basic A- and B-group proteins bind RNA to form several different artifactual structures which are not present in preparations of native hnRNP and which do not function in hnRNP assembly. Three (A2)3B1 tetramers bind the 19S complex to form a 35S assembly intermediate. Following UV irradiation to immobilize the C proteins on the packaged RNA, the 19S triangular complex is recovered as a remnant structure from both native and reconstituted hnRNP particles. C protein-RNA complexes composed of three, six, or nine tetramers (one, two, or three triangular complexes) nucleate the stoichiometric assembly of monomer, dimer, and trimer hnRNP particles. The binding of C-protein tetramers to RNAs longer than 230 nt is through a self-cooperative combinatorial mode. RNA packaged in the 19S complex and in 40S hnRNP particles is efficiently spliced in vitro. These findings demonstrate that formation of the triangular C protein-RNA complex is an obligate first event in the in vitro and probably the in vivo assembly the 40S hnRNP core particle, and they provide insight into the mechanism through which the core proteins package 700-nt increments of RNA. These findings also demonstrate that unless excluded by other factors, the C proteins are likely to be located along the length of nascent transcripts.

scholarly journals Fluorescence studies on the role of tryptophan in heterogeneous nuclear ribonucleoprotein particles of HeLa cells

1989 ◽  
Vol 263 (1) ◽  
pp. 279-283 ◽  
Author(s):  
J Schenkel ◽  
I Appel ◽  
R Schwarzwald ◽  
E K F Bautz ◽  
J Wolfrum ◽  
...  
Keyword(s):  
Protein Interactions ◽  
Hela Cells ◽  
Rna Binding ◽  
Tryptophan Fluorescence ◽  
Fluorescence Decay ◽  
Fluorescence Studies ◽  
Heterogeneous Nuclear Ribonucleoprotein ◽  
Core Proteins ◽  
Tryptophan Residues ◽  
Nuclear Ribonucleoprotein

The 40 S heterogeneous nuclear ribonucleoprotein (hnRNP) particles from HeLa cells reveal tryptophan fluorescence with a bi-exponential decay, indicating that only a few of the ‘core’ proteins contain tryptophan residues. The presence of tryptophan residues distinguishes hnRNP particles from nucleosomes, with which they otherwise share a number of properties. This difference, however, is not essential for protein-RNA binding, as the fluorescence decay remains unchanged when hnRNP particles are dissociated into protein and RNA. However, the Stern-Volmer quenching constant is doubled upon salt dissociation, i.e. tryptophan residues become more accessible to solvent. Thus tryptophan quenching is a useful parameter for monitoring protein-protein interactions in hnRNP particles.

scholarly journals The C-protein tetramer binds 230 to 240 nucleotides of pre-mRNA and nucleates the assembly of 40S heterogeneous nuclear ribonucleoprotein particles

1994 ◽  
Vol 14 (1) ◽  
pp. 518-533
Author(s):  
M Huang ◽  
J E Rech ◽  
S J Northington ◽  
P F Flicker ◽  
A Mayeda ◽  
...  
Keyword(s):  
Rna Binding ◽  
Binding Studies ◽  
C Protein ◽  
Heterogeneous Nuclear Ribonucleoprotein ◽  
Core Proteins ◽  
C Proteins ◽  
Assembly Intermediate ◽  
Nuclear Ribonucleoprotein

A series of in vitro protein-RNA binding studies using purified native (C1)3C2 and (A2)3B1 tetramers, total soluble heterogeneous nuclear ribonucleoprotein (hnRNP), and pre-mRNA molecules differing in length and sequence have revealed that a single C-protein tetramer has an RNA site size of 230 to 240 nucleotides (nt). Two tetramers bind twice this RNA length, and three tetramers fold monoparticle lengths of RNA (700 nt) into a unique 19S triangular complex. In the absence of this unique structure, the basic A- and B-group proteins bind RNA to form several different artifactual structures which are not present in preparations of native hnRNP and which do not function in hnRNP assembly. Three (A2)3B1 tetramers bind the 19S complex to form a 35S assembly intermediate. Following UV irradiation to immobilize the C proteins on the packaged RNA, the 19S triangular complex is recovered as a remnant structure from both native and reconstituted hnRNP particles. C protein-RNA complexes composed of three, six, or nine tetramers (one, two, or three triangular complexes) nucleate the stoichiometric assembly of monomer, dimer, and trimer hnRNP particles. The binding of C-protein tetramers to RNAs longer than 230 nt is through a self-cooperative combinatorial mode. RNA packaged in the 19S complex and in 40S hnRNP particles is efficiently spliced in vitro. These findings demonstrate that formation of the triangular C protein-RNA complex is an obligate first event in the in vitro and probably the in vivo assembly the 40S hnRNP core particle, and they provide insight into the mechanism through which the core proteins package 700-nt increments of RNA. These findings also demonstrate that unless excluded by other factors, the C proteins are likely to be located along the length of nascent transcripts.

scholarly journals Myoparr-Associated and -Independent Multiple Roles of Heterogeneous Nuclear Ribonucleoprotein K during Skeletal Muscle Cell Differentiation

2021 ◽  
Vol 23 (1) ◽  
pp. 108
Author(s):  
Keisuke Hitachi ◽  
Yuri Kiyofuji ◽  
Masashi Nakatani ◽  
Kunihiro Tsuchida
Keyword(s):  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Myogenic Differentiation ◽  
Multiple Roles ◽  
Cell Physiology ◽  
Transcriptional Level ◽  
Multiple Functions ◽  
Heterogeneous Nuclear Ribonucleoprotein ◽  
Nuclear Ribonucleoprotein

RNA-binding proteins (RBPs) regulate cell physiology via the formation of ribonucleic-protein complexes with coding and non-coding RNAs. RBPs have multiple functions in the same cells; however, the precise mechanism through which their pleiotropic functions are determined remains unknown. In this study, we revealed the multiple inhibitory functions of heterogeneous nuclear ribonucleoprotein K (hnRNPK) for myogenic differentiation. We first identified hnRNPK as a lncRNA Myoparr binding protein. Gain- and loss-of-function experiments showed that hnRNPK repressed the expression of myogenin at the transcriptional level. The hnRNPK-binding region of Myoparr was required to repress myogenin expression. Moreover, hnRNPK repressed the expression of a set of genes coding for aminoacyl-tRNA synthetases in a Myoparr-independent manner. Mechanistically, hnRNPK regulated the eIF2α/Atf4 pathway, one branch of the intrinsic pathways of the endoplasmic reticulum sensors, in differentiating myoblasts. Thus, our findings demonstrate that hnRNPK plays lncRNA-associated and -independent multiple roles during myogenic differentiation, indicating that the analysis of lncRNA-binding proteins will be useful for elucidating both the physiological functions of lncRNAs and the multiple functions of RBPs.

scholarly journals Identification of two RNA-binding proteins in Balbiani ring premessenger ribonucleoprotein granules and presence of these proteins in specific subsets of heterogeneous nuclear ribonucleoprotein particles.

1996 ◽  
Vol 16 (4) ◽  
pp. 1425-1435 ◽  
Author(s):  
Wurtz-T ◽  
E Kiseleva ◽  
G Nacheva ◽  
A Alzhanova-Ericcson ◽  
A Rosén ◽  
...  
Keyword(s):  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Affinity Column ◽  
Balbiani Ring ◽  
Western Blots ◽  
Heterogeneous Nuclear Ribonucleoprotein ◽  
Ribonucleoprotein Particles ◽  
Specific Subset ◽  
Nuclear Ribonucleoprotein

Balbiani ring (BR) granules are premessenger ribonucleoprotein particles (RNPs) generated in giant chromosomal puffs, the BRs, in the larval salivary glands of the dipteran chironomus tentans. Monoclonal antibodies were raised against nuclear proteins collected on a single-stranded-DNA-agarose affinity column, and two of them were used to identify RNA-binding proteins in BR granules. First, in Western blots (immunoblots), one of the antibodies recognized a 36-kDa protein and the other recognized a 45-KDa protein. Second, both antibodies bound to the BRs in immunocytological experiments. It was shown in cross-linking experiments that the two proteins are associated with heterogeneous nuclear RNP (hnRNP) complexes extracted from C. tentans nuclei. By immunoelectron microscopy of isolated and partly unfolded BR RNPs, it was specifically demonstrated that the BR granules contain the two proteins and, in addition, that both proteins are distributed frequently along the RNP fiber of the particles. Thus, the 36- and 45-KDa proteins are likely to be abundant, RNA-binding proteins in the BR particles. To elucidate to what extent the two proteins are also present in other hnRNPs, we studied the binding of the antibodies to chromosomal puffs in general. It was observed that many puffs in addition to the BRs harbor the two proteins, but there are also puffs containing only one of the components, either the 36- or the 45-kDa protein. We conclude that the two proteins are not randomly bound to all hnRNPs but that each of them seems to be linked to a specific subset of the particles.

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