1. Academic Validation
  2. DRIP150 coactivation of estrogen receptor alpha in ZR-75 breast cancer cells is independent of LXXLL motifs

DRIP150 coactivation of estrogen receptor alpha in ZR-75 breast cancer cells is independent of LXXLL motifs

  • J Biol Chem. 2005 Mar 11;280(10):8819-30. doi: 10.1074/jbc.M413184200.
Jeongeun Eun Lee 1 Kyounghyun Kim James C Sacchettini Clare V Smith Stephen Safe
Affiliations

Affiliation

  • 1 Department of Veterinary Physiology and Pharmacology, Texas A&M University, College Station, Texas 77843-4466, USA.
Abstract

Vitamin D receptor-interacting protein 150 (DRIP150) has been identified as part of mediator-like complexes that enhance transcriptional activation of the Estrogen Receptor (ER) and other nuclear receptors (NRs). DRIP150 coactivates ligand-dependent ERalpha-mediated transactivation in ZR-75 and MDA-MB-231 breast Cancer cells transfected with a (luciferase) reporter construct (pERE3) regulated by three tandem estrogen-responsive elements. Coactivation of ERalpha by DRIP150 in ZR-75 cells was activation function 2-dependent and required an intact helix 12 that typically interacts with LXXLL motifs (NR box) in p160 steroid receptor coactivators. DRIP150 contains C- and N-terminal NR boxes (Amino acids 1182-1186 and 69-73, respectively), and deletion analysis of DRIP150 showed that regions containing these sequences were not necessary for coactivation of ERalpha. Analysis of multiple DRIP150 deletion mutants identified a 23-amino-acid sequence (789-811) required for coactivation activity. Analysis of the protein crystal structure data base identified two regions at Amino acids 789-794 and 795-804, which resembled alpha-helical motifs in Lanuginosa Lipase/histamine N-methyltransferase and hepatocyte nuclear factor 1, respectively. By using a squelching assay and specific amino acid point mutations within each alpha-helix, the NIFSEVRVYN (795-804) region was identified as the critical sequence required for the activity of DRIP150. These results demonstrate that coactivation of ERalpha by DRIP150 in ZR-75 cells is NR box-independent and requires a novel sequence with putative alpha-helical structure.

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