hnRNP H and hnRNP F complex with Fox2 to silence fibroblast growth factor receptor 2 exon IIIc

Mol Cell Biol. 2008 Sep;28(17):5403-19. doi: 10.1128/MCB.00739-08. Epub 2008 Jun 23.

Abstract

The heterogeneous nuclear ribonucleoprotein H (hnRNP) family of proteins has been shown to activate exon inclusion by binding intronic G triplets. Much less is known, however, about how hnRNP H and hnRNP F silence exons. In this study, we identify hnRNP H and hnRNP F proteins as being novel silencers of fibroblast growth factor receptor 2 exon IIIc. In cells that normally include this exon, we show that the overexpression of either hnRNP H1 or hnRNP F resulted in the dramatic silencing of exon IIIc. In cells that normally skip exon IIIc, skipping was disrupted when RNA interference was used to knock down both hnRNP H and hnRNP F. We show that an exonic GGG motif overlapped a critical exonic splicing enhancer, which was predicted to bind the SR protein ASF/SF2. Furthermore, the expression of ASF/SF2 reversed the silencing of exon IIIc caused by the expression of hnRNP H1. We show that hnRNP H and hnRNP F proteins are present in a complex with Fox2 and that the presence of Fox allows hnRNP H1 to better compete with ASF/SF2 for binding to exon IIIc. These results establish hnRNP H and hnRNP F as being repressors of exon inclusion and suggest that Fox proteins enhance their ability to antagonize ASF/SF2.

Publication types

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

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Base Sequence
  • Binding, Competitive
  • Cell Line
  • Conserved Sequence
  • DNA Mutational Analysis
  • Exons / genetics*
  • Gene Silencing*
  • Heterogeneous-Nuclear Ribonucleoprotein Group F-H / chemistry
  • Heterogeneous-Nuclear Ribonucleoprotein Group F-H / metabolism*
  • Humans
  • Immunoprecipitation
  • Molecular Sequence Data
  • Nuclear Proteins / metabolism
  • Protein Binding
  • RNA Splicing Factors
  • RNA-Binding Proteins / metabolism*
  • Rats
  • Receptor, Fibroblast Growth Factor, Type 2 / genetics*
  • Regulatory Sequences, Nucleic Acid / genetics
  • Repressor Proteins / metabolism*
  • Sequence Alignment
  • Serine-Arginine Splicing Factors
  • Structure-Activity Relationship

Substances

  • Heterogeneous-Nuclear Ribonucleoprotein Group F-H
  • Nuclear Proteins
  • RBFOX2 protein, human
  • RNA Splicing Factors
  • RNA-Binding Proteins
  • Repressor Proteins
  • Serine-Arginine Splicing Factors
  • Receptor, Fibroblast Growth Factor, Type 2