Association with Ets-1 Causes Ligand- and AF2-Independent Activation of Nuclear Receptors

Abstract

The vitamin D receptor (VDR) normally functions as a ligand-dependent transcriptional activator. Here we show that, in the presence of Ets-1, VDR stimulates the prolactin promoter in a ligand-independent manner, behaving as a constitutive activator. Mutations in the AF2 domain abolish vitamin D-dependent transactivation but do not affect constitutive activation by Ets-1. Therefore, in contrast with the actions of vitamin D, activation by Ets-1 is independent of the AF2 domain. Ets-1 also conferred a ligand-independent activation to the estrogen receptor and to peroxisome proliferator-activated receptor α. In addition, Ets-1 cooperated with the unliganded receptors to stimulate the activity of reporter constructs containing consensus response elements fused to the thymidine kinase promoter. There is a direct interaction of the receptors with Ets-1 which requires the DNA binding domains of both proteins. Interaction with Ets-1 induces a conformational change in VDR which can be detected by an increased resistance to proteolytic digestion. Furthermore, a retinoid X receptor-VDR heterodimer in which both receptors lack the core C-terminal AF2 domain can recruit coactivators in the presence, but not in the absence, of Ets-1. This suggests that Ets-1 induces a conformational change in the receptor which creates an active interaction surface with coactivators even in the AF2-defective mutants. These results demonstrate the existence of a novel mechanism, alternative to ligand binding, which can convert an unliganded receptor from an inactive state into a competent transcriptional activator.

ACKNOWLEDGMENTS

We thank R. Evans and M. Parker for plasmids used in this study. The Ets-1 fragments were a kind gift from A. Gutierrez-Hartmann.

This work was supported by grant PM97-0135 from the D.G.E.S., by grants 08.1/0032 and 08.6/0010.1/1999 from the Comunidad de Madrid, and by the Fundación Salud 2000 (Serono).

R.M.T. and A.I.C. contributed equally to this work.