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

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Item Type:Article
Title:Npl3 functions in mRNP assembly by recruitment of mRNP components to the transcription site and their transfer onto the mRNA
Creators Name:Keil, P. and Wulf, A. and Kachariya, N. and Reuscher, S. and Hühn, K. and Silbern, I. and Altmüller, J. and Keller, M. and Stehle, R. and Zarnack, K. and Sattler, M. and Urlaub, H. and Sträßer, K.
Abstract: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.
Keywords:Nuclear Proteins, Messenger RNA, RNA-Binding Proteins, Ribonucleoproteins, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins, Nuclear Proteins
Source:Nucleic Acids Research
ISSN:0305-1048
Publisher:Oxford University Press
Page Range:gkac1206
Date:30 December 2022
Official Publication:https://doi.org/10.1093/nar/gkac1206
PubMed:View item in PubMed
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https://edoc.mdc-berlin.de/21842/Preprint version

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