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

  • 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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Metadaten
Author:Philipp KeilGND, Alexander WulfGND, Nitin KachariyaORCiD, Samira Reuscher, Kristin HühnGND, Ivan SilbernORCiDGND, Janine AltmüllerORCiDGND, Mario KellerORCiDGND, Ralf Stehle, Katharina ZarnackORCiDGND, Michael SattlerORCiD, Henning UrlaubORCiD, Katja SträßerORCiDGND
URN:urn:nbn:de:hebis:30:3-830781
DOI:https://doi.org/10.1093/nar/gkac1206
ISSN:0305-1048
Pubmed Id:https://pubmed.ncbi.nlm.nih.gov/36583366
Parent Title (English):Nucleic acids research
Publisher:Oxford Univ. Press
Place of publication:Oxford
Document Type:Article
Language:English
Date of Publication (online):2022/12/30
Date of first Publication:2022/12/30
Publishing Institution:Universitätsbibliothek Johann Christian Senckenberg
Release Date:2024/03/30
Volume:51.2023
Issue:2
Page Number:21
First Page:831
Last Page:851
HeBIS-PPN:521812917
Institutes:Biowissenschaften
Fachübergreifende Einrichtungen / Buchmann Institut für Molekulare Lebenswissenschaften (BMLS)
Dewey Decimal Classification:5 Naturwissenschaften und Mathematik / 57 Biowissenschaften; Biologie / 570 Biowissenschaften; Biologie
Sammlungen:Universitätspublikationen
Licence (German):License LogoCreative Commons - CC BY-NC - Namensnennung - Nicht kommerziell 4.0 International