The U1-70K and SRSF1 interaction is modulated by phosphorylation during the early stages of spliceosome assembly.

Autor: Paul T; Department of Chemistry, University of Alabama at Birmingham, Birmingham, Alabama, USA., Zhang P; Department of Medicine, Division of Hematology/Oncology, University of Alabama at Birmingham, Birmingham, Alabama, USA., Zhang Z; Department of Chemistry, University of Alabama at Birmingham, Birmingham, Alabama, USA., Fargason T; Department of Chemistry, University of Alabama at Birmingham, Birmingham, Alabama, USA., De Silva NIU; Department of Chemistry, University of Alabama at Birmingham, Birmingham, Alabama, USA., Powell E; Department of Chemistry, University of Alabama at Birmingham, Birmingham, Alabama, USA., Ekpenyong E; Department of Chemistry, University of Alabama at Birmingham, Birmingham, Alabama, USA., Jamal S; Department of Chemistry, University of Alabama at Birmingham, Birmingham, Alabama, USA., Yu Y; Department of Chemistry and Biochemistry, University of Delaware, Newark, Delaware, USA., Prevelige P; Department of Microbiology, University of Alabama at Birmingham, Birmingham, Alabama, USA., Lu R; Department of Medicine, Division of Hematology/Oncology, University of Alabama at Birmingham, Birmingham, Alabama, USA., Zhang J; Department of Chemistry, University of Alabama at Birmingham, Birmingham, Alabama, USA.
Jazyk: angličtina
Zdroj: Protein science : a publication of the Protein Society [Protein Sci] 2024 Aug; Vol. 33 (8), pp. e5117.
DOI: 10.1002/pro.5117
Abstrakt: In eukaryotes, pre-mRNA splicing is vital for RNA processing and orchestrated by the spliceosome, whose assembly starts with the interaction between U1-70K and SR proteins. Despite the significance of the U1-70K/SR interaction, the dynamic nature of the complex and the challenges in obtaining soluble U1-70K have impeded a comprehensive understanding of the interaction at the structural level for decades. We overcome the U1-70K solubility issues, enabling us to characterize the interaction between U1-70K and SRSF1, a representative SR protein. We unveil specific interactions: phosphorylated SRSF1 RS with U1-70K BAD1, and SRSF1 RRM1 with U1-70K RRM. The RS/BAD1 interaction plays a dominant role, whereas the interaction between the RRM domains further enhances the stability of the U1-70K/SRSF1 complex. The RRM interaction involves the C-terminal extension of U1-70K RRM and the conserved acid patches on SRSF1 RRM1 that is involved in SRSF1 phase separation. Our circular dichroism spectra reveal that BAD1 adapts an α-helical conformation and RS is intrinsically disordered. Intriguingly, BAD1 undergoes a conformation switch from α-helix to β-strand and random coil upon RS binding. In addition to the regulatory mechanism via SRSF1 phosphorylation, the U1-70K/SRSF1 interaction is also regulated by U1-70K BAD1 phosphorylation. We find that U1-70K phosphorylation inhibits the U1-70K and SRSF1 interaction. Our structural findings are validated through in vitro splicing assays and in-cell saturated domain scanning using the CRISPR method, providing new insights into the intricate regulatory mechanisms of pre-mRNA splicing.
(© 2024 The Author(s). Protein Science published by Wiley Periodicals LLC on behalf of The Protein Society.)
Databáze: MEDLINE
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