Evolutionary trace-based peptides identify a novel asymmetric interaction that mediates oligomerization in nuclear receptors

J Biol Chem. 2005 Sep 9;280(36):31818-29. doi: 10.1074/jbc.M501924200. Epub 2005 Jul 1.


Germ cell nuclear factor (GCNF) is an orphan nuclear receptor that plays important roles in development and reproduction, by repressing the expression of essential genes such as Oct4, GDF9, and BMP15, through binding to DR0 elements. Surprisingly, whereas recombinant GCNF binds to DR0 sequences as a homodimer, endogenous GCNF does not exist as a homodimer but rather as part of a large complex termed the transiently retinoid-induced factor (TRIF). Here, we use evolutionary trace (ET) analysis to design mutations and peptides that probe the molecular basis for the formation of this unusual complex. We find that GCNF homodimerization and TRIF complex formation are DNA-dependent, and ET suggests that dimerization involves key functional sites on both helix 3 and helix 11, which are located on opposing surfaces of the ligand binding domain. Targeted mutations in either helix of GCNF disrupt the formation of both the homodimer and the endogenous TRIF complex. Moreover, peptide mimetics of both of these ET-determined sites inhibit dimerization and TRIF complex formation. This suggests that a novel helix 3-helix 11 heterotypic interaction mediates GCNF interaction and would facilitate oligomerization. Indeed, it was determined that the endogenous TRIF complex is composed of a GCNF oligomer. These findings shed light on an evolutionarily selected mechanism that reveals the unusual DNA-binding, dimerization, and oligomerization properties of GCNF.

Publication types

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

MeSH terms

  • Adaptor Proteins, Vesicular Transport / genetics
  • Adaptor Proteins, Vesicular Transport / metabolism
  • Adaptor Proteins, Vesicular Transport / physiology
  • Amino Acid Sequence
  • Cell Line
  • DNA-Binding Proteins / genetics
  • DNA-Binding Proteins / metabolism
  • DNA-Binding Proteins / physiology
  • Dimerization
  • Evolution, Molecular*
  • Genes, Reporter
  • Molecular Sequence Data
  • Nuclear Receptor Subfamily 6, Group A, Member 1
  • Peptides / genetics
  • Peptides / metabolism*
  • Peptides / physiology
  • Point Mutation
  • Protein Structure, Secondary
  • Protein Structure, Tertiary
  • Receptor Cross-Talk / physiology*
  • Receptors, Cytoplasmic and Nuclear / genetics
  • Receptors, Cytoplasmic and Nuclear / metabolism*
  • Receptors, Cytoplasmic and Nuclear / physiology
  • Receptors, Retinoic Acid / genetics
  • Receptors, Retinoic Acid / metabolism
  • Receptors, Retinoic Acid / physiology
  • Response Elements


  • Adaptor Proteins, Vesicular Transport
  • DNA-Binding Proteins
  • NR6A1 protein, human
  • Nuclear Receptor Subfamily 6, Group A, Member 1
  • Peptides
  • Receptors, Cytoplasmic and Nuclear
  • Receptors, Retinoic Acid
  • TICAM1 protein, human