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Keil, P.* ; Wulf, A.* ; Kachariya, N. ; Reuscher, S.* ; Huhn, K.* ; Silbern, I.* ; Altmüller, J.* ; Keller, M.* ; Stehle, R. ; Zarnack, K.* ; Sattler, M. ; Urlaub, H.* ; Sträßer, K.*

Npl3 functions in mRNP assembly by recruitment of mRNP components to the transcription site and their transfer onto the mRNA.

Nucleic Acids Res. 51, 831-851 (2023)
Postprint Forschungsdaten DOI PMC
Open Access Gold
Creative Commons Lizenzvertrag
RNA-binding proteins (RBPs) control every RNA metabolic process by multiple protein-RNA and protein-protein interactions. Their roles have largely been analyzed by crude mutations, which abrogate multiple functions at once and likely impact the structural integrity of the large ribonucleoprotein particles (RNPs) these proteins function in. Using UV-induced RNA-protein crosslinking of entire cells, protein complex purification and mass spectrometric analysis, we identified >100 in vivo RNA crosslinks in 16 nuclear mRNP components in Saccharomyces cerevisiae. For functional analysis, we chose Npl3, which displayed crosslinks in its two RNA recognition motifs (RRMs) and in the connecting flexible linker region. Both RRM domains and the linker uniquely contribute to RNA recognition as revealed by NMR and structural analyses. Interestingly, mutations in these regions cause different phenotypes, indicating distinct functions of the different RNA-binding domains. Notably, an npl3-Linker mutation strongly impairs recruitment of several mRNP components to chromatin and incorporation of other mRNP components into nuclear mRNPs, establishing a so far unknown function of Npl3 in nuclear mRNP assembly. Taken together, our integrative analysis uncovers a specific function of the RNA-binding activity of the nuclear mRNP component Npl3. This approach can be readily applied to RBPs in any RNA metabolic process.
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Publikationstyp Artikel: Journalartikel
Dokumenttyp Wissenschaftlicher Artikel
Korrespondenzautor
Schlagwörter Heteronuclear Nmr-spectroscopy; Small-angle Scattering; Photo-cross-linking; Mass-spectrometry; Gene-expression; Splicing Regulation; Protein Structures; Backbone Dynamics; Binding Protein; Sr Proteins
ISSN (print) / ISBN 0305-1048
e-ISSN 1362-4962
Zeitschrift Nucleic Acids Research
Quellenangaben Band: 51, Heft: 2, Seiten: 831-851 Artikelnummer: , Supplement: ,
Verlag Oxford University Press
Verlagsort Great Clarendon St, Oxford Ox2 6dp, England
Nichtpatentliteratur Publikationen
Begutachtungsstatus Peer reviewed
Institut(e) Institute of Structural Biology (STB)
Förderungen EU
DFG