The RNA-splicing factor PSF/p54 controls DNA-topoisomerase I activity by a direct interaction

J Biol Chem. 1998 Oct 9;273(41):26261-4. doi: 10.1074/jbc.273.41.26261.

Abstract

DNA-topoisomerase I has been implied in RNA splicing because it catalyzes RNA strand transfer and activates serine/arginine-rich RNA-splicing factors by phosphorylation. Here, we demonstrate a direct interaction between topoisomerase I and pyrimidine tract binding protein-associated splicing factor (PSF), a cofactor of RNA splicing, which forms heterodimers with its smaller homolog, the nuclear RNA-binding protein of 54 kDa (p54). Topoisomerase I, PSF, and p54 copurified in a 1:1:1 ratio from human A431 cell nuclear extracts. Specific binding of topoisomerase I to PSF (but not p54) was demonstrated by coimmunoprecipitation and by far Western blotting, in which renatured blots were probed with biotinylated topoisomerase I. Chemical cross-linking of pure topoisomerase I revealed monomeric, dimeric, and trimeric enzyme forms, whereas in the presence of PSF/p54 the enzyme was cross-linked into complexes larger than homotrimers. When topoisomerase I was complexed with PSF/p54 it was 16-fold more active than the pure enzyme, which could be stimulated 5- and 16-fold by the addition of recombinant PSF or native PSF/p54, respectively. A physiological role of this stimulatory mechanism seems feasible, because topoisomerase I and PSF showed a patched colocalization in A431 cell nuclei, which varied with cell cycle.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Catalysis
  • Cell Line
  • DNA / metabolism
  • DNA Topoisomerases, Type I / metabolism*
  • DNA-Binding Proteins
  • Enzyme Activation
  • Humans
  • Nuclear Matrix-Associated Proteins*
  • Nuclear Proteins / metabolism*
  • Octamer Transcription Factors
  • Protein Binding
  • RNA-Binding Proteins / metabolism*
  • Recombinant Proteins / metabolism

Substances

  • DNA-Binding Proteins
  • NONO protein, human
  • Nuclear Matrix-Associated Proteins
  • Nuclear Proteins
  • Octamer Transcription Factors
  • RNA-Binding Proteins
  • Recombinant Proteins
  • DNA
  • DNA Topoisomerases, Type I