The human U5 snRNP 52K protein (CD2BP2) interacts with U5-102K (hPrp6), a U4/U6.U5 tri-snRNP bridging protein, but dissociates upon tri-snRNP formation

RNA. 2005 May;11(5):598-608. doi: 10.1261/rna.2300805.

Abstract

The U5 snRNP plays an essential role in both U2- and U12-dependent splicing. Here, we have characterized a 52-kDa protein associated with the human U5 snRNP, designated U5-52K. Protein sequencing revealed that U5-52K is identical to the CD2BP2, which interacts with the cytoplasmic portion of the human T-cell surface protein CD2. Consistent with it associating with an snRNP, immunofluorescence studies demonstrated that the 52K protein is predominantly located in the nucleoplasm of HeLa cells, where it overlaps, at least in part, with splicing-factor compartments (or "speckles"). We further demonstrate that the 52K protein is a constituent of the 20S U5 snRNP, but is not found in U4/U6.U5 tri-snRNPs. Thus, it is the only 20S U5-specific protein that is not integrated into the tri-snRNP and resembles, in this respect, the U4/U6 di-snRNP assembly factor Prp24p/p110. Yeast two-hybrid screening and pulldown assays revealed that the 52K protein interacts with the U5-specific 102K and 15K proteins, suggesting that these interactions are responsible for its integration into the U5 particle. The N-terminal two-thirds of 52K interact with the 102K protein, whereas its C-terminal GYF-domain binds the 15K protein. As the latter lacks a proline-rich tract, our data indicate that a GYF-domain can also engage in specific protein-protein interactions in a polyproline-independent manner. Interestingly, the U5-102K protein has been shown previously to play an essential role in tri-snRNP formation, binding the U4/U6-61K protein. The interaction of 52K with a tri-snRNP bridging protein, coupled with its absence from the tri-snRNP, suggests it might function in tri-snRNP assembly.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adaptor Proteins, Signal Transducing
  • Amino Acid Sequence
  • Carrier Proteins / chemistry
  • Carrier Proteins / metabolism*
  • Cell Nucleus / metabolism
  • Conserved Sequence
  • HeLa Cells
  • Humans
  • Molecular Sequence Data
  • Molecular Weight
  • Multiprotein Complexes / chemistry
  • Multiprotein Complexes / metabolism
  • Protein Binding
  • Protein Transport
  • Ribonucleoprotein, U4-U6 Small Nuclear / chemistry
  • Ribonucleoprotein, U4-U6 Small Nuclear / metabolism*
  • Ribonucleoprotein, U5 Small Nuclear / chemistry
  • Ribonucleoprotein, U5 Small Nuclear / metabolism*
  • Substrate Specificity
  • Two-Hybrid System Techniques

Substances

  • Adaptor Proteins, Signal Transducing
  • CD2BP2 protein, human
  • Carrier Proteins
  • Multiprotein Complexes
  • Ribonucleoprotein, U4-U6 Small Nuclear
  • Ribonucleoprotein, U5 Small Nuclear
  • TOM translocase