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Mechanism of Mediator recruitment by tandem Gcn4 activation domains and three Gal11 activator-binding domains.

Targets of the tandem Gcn4 acidic activation domains in transcription preinitiation complexes were identified by site-specific cross-linking. The individual Gcn4 activation domains cross-link to three common targets, Gal11/Med15, Taf12, and Tra1, which are subunits of four conserved coactivator complexes, Mediator, SAGA, TFIID, and NuA4. The Gcn4 N-terminal activation domain also cross-links to the Mediator subunit Sin4/Med16. The contribution of the two Gcn4 activation domains to transcription was gene specific and varied from synergistic to less than additive. Gcn4-dependent genes had a requirement for Gal11 ranging from 10-fold dependence to complete Gal11 independence, while the Gcn4-Taf12 interaction did not significantly contribute to the expression of any gene studied. Complementary methods identified three conserved Gal11 activator-binding domains that bind each Gcn4 activation domain with micromolar affinity. These Gal11 activator-binding domains contribute additively to transcription activation and Mediator recruitment at Gcn4- and Gal11-dependent genes. Although we found that the conserved Gal11 KIX domain contributes to Gal11 function, we found no evidence of specific Gcn4-KIX interaction and conclude that the Gal11 KIX domain does not function by specific interaction with Gcn4. Our combined results show gene-specific coactivator requirements, a surprising redundancy in activator-target interactions, and an activator-coactivator interaction mediated by multiple low-affinity protein-protein interactions.

Pubmed ID: 20308326

Authors

  • Herbig E
  • Warfield L
  • Fish L
  • Fishburn J
  • Knutson BA
  • Moorefield B
  • Pacheco D
  • Hahn S

Journal

Molecular and cellular biology

Publication Data

May 27, 2010

Associated Grants

  • Agency: NIGMS NIH HHS, Id: 5R01 GM075114
  • Agency: NIGMS NIH HHS, Id: R01 GM075114
  • Agency: NCI NIH HHS, Id: T32 CA080416
  • Agency: NCI NIH HHS, Id: T32 CA09657
  • Agency: NCI NIH HHS, Id: T32 CA80416

Mesh Terms

  • Basic-Leucine Zipper Transcription Factors
  • Gene Expression Regulation, Fungal
  • Mediator Complex
  • Protein Binding
  • Protein Structure, Tertiary
  • Protein Subunits
  • Saccharomyces cerevisiae
  • Saccharomyces cerevisiae Proteins
  • TATA-Binding Protein Associated Factors
  • Transcription Factor TFIID
  • Transcriptional Activation