The Heterogeneous Nuclear Ribonucleoprotein C Protein Tetramer Binds U1, U2, and U6 snRNAs through Its High Affinity RNA Binding Domain (the bZIP-like Motif)

Based on UV cross-linking experiments, it has been reported that the C protein tetramer of 40 S heterogeneous nuclear ribonucleoprotein complexes specifically interacts with stem-loop I of U2 small nuclear RNA (snRNA) (Temsamani, J., and Pederson, T. (1996)J. Biol. Chem. 271, 24922–24926), that C pr...

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Published in:The Journal of biological chemistry 1998-08, Vol.273 (33), p.21359-21367
Main Authors: Shahied-Milam, Lillian, Soltaninassab, Syrus R., Iyer, Gopakumar V., LeStourgeon, Wallace M.
Format: Article
Language:English
Subjects:
HeLa Cells
Heterogeneous-Nuclear Ribonucleoproteins
Humans
Phosphorylation
Protein Binding
Ribonucleoproteins - chemistry
Ribonucleoproteins - metabolism
RNA, Small Nuclear - chemistry
RNA, Small Nuclear - metabolism
RNA-Binding Proteins - metabolism
Uridine - metabolism
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Summary:Based on UV cross-linking experiments, it has been reported that the C protein tetramer of 40 S heterogeneous nuclear ribonucleoprotein complexes specifically interacts with stem-loop I of U2 small nuclear RNA (snRNA) (Temsamani, J., and Pederson, T. (1996)J. Biol. Chem. 271, 24922–24926), that C protein disrupts U4:U6 snRNA complexes (Forne, T., Rossi, F., Labourier, E., Antoine, E., Cathala, G., Brunel, C., and Tazi, J. (1995) J. Biol. Chem. 270, 16476–16481), that U6 snRNA may modulate C protein phosphorylation (Mayrand, S. H., Fung, P. A., and Pederson, T. (1996) Mol. Cell. Biol. 16, 1241–1246), and that hyperphosphorylated C protein lacks pre-mRNA binding activity. These findings suggest that snRNA-C protein interactions may function to recruit snRNA to, or displace C protein from, splice junctions. In this study, both equilibrium and non-equilibrium RNA binding assays reveal that purified native C protein binds U1, U2, and U6 snRNA with significant affinity (∼7.5–50 nm) although nonspecifically. Competition binding assays reveal that U2 snRNA (the highest affinity snRNA substrate) is ineffective in C protein displacement from branch-point/splice junctions or as a competitor of C protein's self-cooperative RNA binding mode. Additionally, C protein binds snRNA through its high affinity bZLM and mutations in the RNA recognition motif at suggested RNA binding sites primarily affect protein oligomerization.
ISSN:0021-9258
1083-351X
DOI:10.1074/jbc.273.33.21359