A composite double-/single-stranded RNA-binding region in protein Prp3 supports tri-snRNP stability and splicing

Elife. 2015 Jul 10:4:e07320. doi: 10.7554/eLife.07320.

Abstract

Prp3 is an essential U4/U6 di-snRNP-associated protein whose functions and molecular mechanisms in pre-mRNA splicing are presently poorly understood. We show by structural and biochemical analyses that Prp3 contains a bipartite U4/U6 di-snRNA-binding region comprising an expanded ferredoxin-like fold, which recognizes a 3'-overhang of U6 snRNA, and a preceding peptide, which binds U4/U6 stem II. Phylogenetic analyses revealed that the single-stranded RNA-binding domain is exclusively found in Prp3 orthologs, thus qualifying as a spliceosome-specific RNA interaction module. The composite double-stranded/single-stranded RNA-binding region assembles cooperatively with Snu13 and Prp31 on U4/U6 di-snRNAs and inhibits Brr2-mediated U4/U6 di-snRNA unwinding in vitro. RNP-disrupting mutations in Prp3 lead to U4/U6•U5 tri-snRNP assembly and splicing defects in vivo. Our results reveal how Prp3 acts as an important bridge between U4/U6 and U5 in the tri-snRNP and comparison with a Prp24-U6 snRNA recycling complex suggests how Prp3 may be involved in U4/U6 reassembly after splicing.

Keywords: S. cerevisiae; X-ray crystallography; biochemistry; biophysics; human; pre-mRNA splicing; protein–RNA interaction; small nuclear ribonucleoprotein particle; structural biology.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Crystallography, X-Ray
  • DNA Mutational Analysis
  • Humans
  • Models, Biological
  • Models, Molecular
  • Mutation
  • Nuclear Proteins / chemistry
  • Nuclear Proteins / genetics
  • Nuclear Proteins / metabolism*
  • Protein Binding
  • RNA / metabolism*
  • RNA Splicing*
  • Ribonucleoprotein, U4-U6 Small Nuclear / chemistry
  • Ribonucleoprotein, U4-U6 Small Nuclear / genetics
  • Ribonucleoprotein, U4-U6 Small Nuclear / metabolism*
  • Ribonucleoproteins, Small Nuclear / chemistry
  • Ribonucleoproteins, Small Nuclear / metabolism*

Substances

  • Nuclear Proteins
  • PRPF3 protein, human
  • Ribonucleoprotein, U4-U6 Small Nuclear
  • Ribonucleoproteins, Small Nuclear
  • RNA

Associated data

  • PDB/4YHU
  • PDB/4YHV
  • PDB/4YHW

Grants and funding

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.