title: Evidence for cooperative tandem binding of hnRNP C RRMs in mRNA processing.
abstract: The human hnRNP C is a ubiquitous cellular protein involved in mRNA maturation. Recently, we have shown that this protein specifically recognizes uridine (U) pentamers through its single RNA recognition motif (RRM). However, a large fraction of natural RNA targets of hnRNP C consists of much longer contiguous uridine stretches. To understand how these extended sites are recognized, we studied the binding of the RRM to U-tracts of 8-11 bases. In vivo investigation of internal translation activation of unr (upstream of N-ras) mRNA indicates that the conservation of the entire hnRNP C binding site, UC(U)8, is required for hnRNP C-dependent IRES activation. The assays further suggest a synergistic interplay between hnRNP C monomers, dependent on the protein's ability to oligomerize. In vitro spectroscopic and thermodynamic analyses show that isolated RRMs bind to (U)11 oligomers as dimers. Structural modeling of a ternary double-RRM/RNA complex indicates additionally that two RRM copies can be accommodated on the canonical sequence UC(U)8. The proposed tandem RRM binding is in very good agreement with the transcriptome-wide recognition of extended U-tracts by full-length hnRNP C, which displays a cross-linking pattern consistent with a positively cooperative RRM dimer binding model.
authors.affiliation: Department of Biology, Institute of Molecular Biology and Biophysics, ETH Zürich, 8093 Zürich, Switzerland.
authors.forename: Zuzana
authors.initials: Z
authors.lastname: Cieniková
authors.affiliation: Department of Biology, Institute of Molecular Biology and Biophysics, ETH Zürich, 8093 Zürich, Switzerland.
authors.forename: Sandrine
authors.initials: S
authors.lastname: Jayne
authors.affiliation: Department of Biology, Institute of Molecular Biology and Biophysics, ETH Zürich, 8093 Zürich, Switzerland.
authors.forename: Fred Franz
authors.initials: FF
authors.lastname: Damberger
authors.affiliation: Department of Biology, Institute of Molecular Biology and Biophysics, ETH Zürich, 8093 Zürich, Switzerland.
authors.forename: Frédéric Hai-Trieu
authors.initials: FH
authors.lastname: Allain
authors.affiliation: Department of Biology, Institute of Molecular Biology and Biophysics, ETH Zürich, 8093 Zürich, Switzerland.
authors.forename: Christophe
authors.initials: C
authors.lastname: Maris
mesh_terms: D001665:Binding Sites / Q000235:genetics*; D034482:Heterogeneous-Nuclear Ribonucleoprotein Group C / Q000378:metabolism*; D034441:Heterogeneous-Nuclear Ribonucleoproteins / Q000235:genetics; D006801:Humans; D011485:Protein Binding / Q000235:genetics*; D000067776:RNA Recognition Motif Proteins / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics* / Q000378:metabolism*; D059467:Transcriptome / Q000235:genetics; D014529:Uridine / Q000235:genetics / Q000378:metabolism
chemical_list: D034482:Heterogeneous-Nuclear Ribonucleoprotein Group C; D034441:Heterogeneous-Nuclear Ribonucleoproteins; D000067776:RNA Recognition Motif Proteins; D012333:RNA, Messenger; D014529:Uridine
keywords: IRES; RRM; alternative splicing; hnRNP C; uridine