Prp8 positioning of U5 snRNA is linked to 5' splice site recognition

RNA. 2018 Jun;24(6):769-777. doi: 10.1261/rna.065458.117. Epub 2018 Feb 27.


Prp8 is an essential protein that regulates spliceosome assembly and conformation during pre-mRNA splicing. Recent cryo-EM structures of the spliceosome model Prp8 as a scaffold for the spliceosome's catalytic U snRNA components. Using a new amino acid probing strategy, we identified a dynamic region in human Prp8 that is positioned to stabilize the pre-mRNA in the spliceosome active site through interactions with U5 snRNA. Mutagenesis of the identified Prp8 residues in yeast indicates a role in 5' splice site recognition. Genetic interactions with spliceosome proteins Isy1, which buttresses the intron branch point, and Snu114, a regulatory GTPase that directly contacts Prp8, further corroborate a role for the same Prp8 residues in substrate positioning and activation. Together the data suggest that adjustments in interactions between Prp8 and U5 snRNA help establish proper positioning of the pre-mRNA into the active site to enhance 5' splice site fidelity.

Keywords: 5′ exon; Prp8; U5 snRNA; chemical probing; pre-mRNA splicing; spliceosome.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Catalytic Domain
  • Humans
  • RNA Precursors / genetics*
  • RNA Splice Sites*
  • RNA, Small Nuclear / genetics*
  • RNA-Binding Proteins / genetics
  • RNA-Binding Proteins / metabolism*
  • Ribonucleoprotein, U4-U6 Small Nuclear / genetics
  • Ribonucleoprotein, U4-U6 Small Nuclear / metabolism*
  • Ribonucleoprotein, U5 Small Nuclear / genetics
  • Ribonucleoprotein, U5 Small Nuclear / metabolism*
  • Saccharomyces cerevisiae / genetics*
  • Saccharomyces cerevisiae / metabolism
  • Spliceosomes


  • PRPF8 protein, human
  • RNA Precursors
  • RNA Splice Sites
  • RNA, Small Nuclear
  • RNA-Binding Proteins
  • Ribonucleoprotein, U4-U6 Small Nuclear
  • Ribonucleoprotein, U5 Small Nuclear
  • U5 small nuclear RNA