Functional antagonism between RNA binding proteins HuR and CUGBP2 determines the fate of COX-2 mRNA translation

Gastroenterology. 2007 Mar;132(3):1055-65. doi: 10.1053/j.gastro.2006.12.031. Epub 2006 Dec 19.


Background and aims: Cyclooxygenase-2 (COX-2) expression is regulated at the levels of messenger RNA (mRNA) stability and translation by AU-rich elements (ARE) located in its 3' untranslated region (3'UTR). Although structurally homologous RNA binding proteins HuR and CUGBP2 stabilize COX-2 mRNA, HuR induces whereas CUGBP2 inhibits COX-2 mRNA translation. This study aimed to determine the antagonism between these proteins on COX-2 expression.

Methods: COX-2 ARE binding activity was determined by nitrocellulose filter binding and UV cross-linking assays using recombinant glutathione S-transferase (GST)/HuR and GST/CUGBP2. Protein:protein interactions were determined by GST pull-down, yeast 2-hybrid, and immunocytochemistry assays. Nucleocytoplasmic shutting was determined by heterokaryon analyses. The effect of CUGBP2 and HuR on COX-2 ARE-dependent translation was shown by a chimeric luciferase mRNA containing COX-2 3'UTR. HT-29 cells were subjected to 12 Gy gamma-irradiation in a cesium irradiator.

Results: CUGBP2 and HuR bind with similar affinities to COX-2 ARE, but CUGBP2 competes with HuR for binding. In vitro, HuR and CUGBP2 heterodimerize. Furthermore, FLAG-tagged HuR and myc-tagged CUGBP2 colocalize in the nucleus of HCT-116 cells. Moreover, both proteins shuttled between the nucleus and cytoplasm. In vitro, HuR enhanced whereas CUGBP2 inhibited translation of the chimeric luciferase COX-2 3'UTR mRNA. Furthermore, CUGBP2 competitively inhibited HuR-mediated translation of the transcript. In HT-29 cells transfected with HuR and CUGBP2, a switch in COX-2 mRNA binding from predominantly HuR to CUGBP2 occurred after radiation treatment, which was coupled with increased silencing of the COX-2 mRNA.

Conclusions: CUGBP2 overrides HuR and suppresses COX-2 mRNA translation.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • 3' Untranslated Regions / metabolism*
  • 3' Untranslated Regions / radiation effects
  • Animals
  • Antigens, Surface / genetics
  • Antigens, Surface / metabolism*
  • Binding, Competitive
  • CELF Proteins
  • Cell Nucleus / metabolism
  • Cyclooxygenase 2 / genetics
  • Cyclooxygenase 2 / metabolism*
  • Cytoplasm / metabolism
  • Dimerization
  • ELAV Proteins
  • ELAV-Like Protein 1
  • Gamma Rays
  • HCT116 Cells
  • HT29 Cells
  • Humans
  • Membrane Proteins / genetics
  • Membrane Proteins / metabolism*
  • Mice
  • NIH 3T3 Cells
  • Nerve Tissue Proteins / genetics
  • Nerve Tissue Proteins / metabolism*
  • Protein Binding
  • Protein Biosynthesis* / radiation effects
  • Protein Interaction Mapping / methods
  • Protein Transport
  • RNA Stability
  • RNA, Messenger / metabolism*
  • RNA, Messenger / radiation effects
  • RNA-Binding Proteins / genetics
  • RNA-Binding Proteins / metabolism*
  • Recombinant Fusion Proteins / metabolism
  • Repressor Proteins / metabolism
  • Trans-Activators / metabolism
  • Transfection


  • 3' Untranslated Regions
  • Antigens, Surface
  • CELF Proteins
  • CELF2 protein, human
  • ELAV Proteins
  • ELAV-Like Protein 1
  • ELAVL1 protein, human
  • Membrane Proteins
  • Nerve Tissue Proteins
  • RNA, Messenger
  • RNA-Binding Proteins
  • Recombinant Fusion Proteins
  • Repressor Proteins
  • Trans-Activators
  • Cyclooxygenase 2
  • PTGS2 protein, human