A conserved protein motif is required for full modulatory activity of negative elongation factor subunits NELF-A and NELF-B in modifying glucocorticoid receptor-regulated gene induction properties

J Biol Chem. 2013 Nov 22;288(47):34055-72. doi: 10.1074/jbc.M113.512426. Epub 2013 Oct 6.

Abstract

NELF-B is a BRCA1-interacting protein and subunit (with NELF-A, -C/D, and -E) of the human negative elongation factor (NELF) complex, which participates in RNA polymerase II pausing shortly after transcription initiation, especially for synchronized gene expression. We now report new activities of NELF-B and other NELF complex subunits, which are to attenuate glucocorticoid receptor (GR)-mediated gene induction, reduce the partial agonist activity of an antagonist, and increase the EC50 of an agonist during nonsynchronized expression of exogenous and endogenous reporters. Stable knockdown of endogenous NELF-B has the opposite effects on an exogenous gene. The GR ligand-binding domain suffices for these biological responses. ChIP assays reveal that NELF-B diminishes GR recruitment to promoter regions of two endogenous genes. Using a new competition assay, NELF-A and NELF-B are each shown to act independently as competitive decelerators at two steps after the site of GR action and before or at the site of reporter gene activity. A common motif in each NELF was identified that is required for full activity of both NELF-A and NELF-B. These studies allow us to position the actions of two new modulators of GR-regulated transactivation, NELF-A and NELF-B, relative to other factors in the overall gene induction sequence.

Keywords: Cofactor Action; Competitive Decelerator; EC50; Gene Induction; Gene Transcription; Glucocorticoid Receptor; Mathematical Modeling; NELF-A and NELF-B; Steroid Hormone; Transcription Factors.

Publication types

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

MeSH terms

  • Animals
  • COS Cells
  • Chlorocebus aethiops
  • Gene Expression Regulation / physiology*
  • Gene Knockdown Techniques
  • Humans
  • Protein Structure, Tertiary
  • Rats
  • Receptors, Glucocorticoid / agonists
  • Receptors, Glucocorticoid / genetics
  • Receptors, Glucocorticoid / metabolism*
  • Response Elements / physiology*
  • Transcription Factors / genetics
  • Transcription Factors / metabolism*
  • Transcription, Genetic / physiology*
  • Transcriptional Elongation Factors / genetics
  • Transcriptional Elongation Factors / metabolism*

Substances

  • NELFA protein, human
  • Receptors, Glucocorticoid
  • Transcription Factors
  • Transcriptional Elongation Factors
  • negative elongation factor