Editors' ChoiceNuclear Receptors

Stable, But Transcriptionally Inactive

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Science Signaling  03 Jul 2012:
Vol. 5, Issue 231, pp. ec180
DOI: 10.1126/scisignal.2003357

Regulation of ligand-activated nuclear receptor activity is complex, involving ligand binding, chaperone interactions, multiple posttranslational modifications, and interactions with corepressors and coactivators. Picard et al. describe a complex interplay between phosphorylation, SUMOylation, and ubiquitylation of the estrogen receptor β (ERβ). SUMO is a small protein that is conjugated to proteins at lysine residues, sometimes at the same residues targeted for modification by ubiquitin. Polyubiquitylation typically destabilizes proteins by targeting them for proteasomal degradation. In vitro SUMOylation reactions and studies in transfected cells showed that ERβ was modified by SUMO-1 at an atypical IKNS motif and that this required phosphorylation at the serine in the motif, which may be a substrate for mitogen-activated protein kinase, or substitution of the serine with a charged glutamic acid or aspartic acid residue. Reporter gene assays showed that SUMOylation-deficient ERβ mutants resulted in enhanced transcriptional activity in response to estrogen. Conversely, overexpression of increasing amounts of SUMO-1 inhibited estrogen-responsive ERβ reporter gene activity. The abundance of ERβ was increased by mutation of the lysine residue in the SUMOylation motif, as well as by introduction of the S6D or S6E mutations that enhanced SUMOylation. This apparent conflict was explained by ubiquitylation of ERβ at this lysine and competition for this residue by SUMO or ubiquitin modification: Overexpression of SUMO-1 reduced the ubiquitylation of ERβ. Subnuclear fractionation experiments comparing the distribution of SUMO-competent and SUMO-deficient ERβ indicated that SUMO modification reduced the association of ERβ with chromatin, a result confirmed by chromatin immunoprecipitation experiments showing that overexpression of SUMO reduced the amount of ERβ bound to estrogen-responsive promoters. Treatment of cells with various kinase inhibitors indicated that SUMOylation of ERβ was reduced when glycogen synthase kinase 3β (GSK-3β) was inhibited. However, the serine in the IKNS motif does not match the consensus sequence for a GSK-3β phosphorylation site. Instead, a consensus GSK-3β motif was present adjacent to the IKNS motif, and analysis of SUMOylation of ERβ with mutation of the two serine residues in the GSK-3β motif suggested that these were targeted by GSK-3β. Thus, Picard et al. have identified a phosphorylation-dependent SUMOylation motif that is also regulated by phosphorylation outside the motif. SUMOylation of ERβ at this site stabilizes the receptor by reducing its ubiquitylation but reduces its transcriptional activity by preventing its interaction with target gene promoters.

N. Picard, V. Caron, S. Bilodeau, M. Sanchez, X. Mascle, M. Aubry, A. Tremblay, Identification of estrogen receptor β as a SUMO-1 target reveals a novel phosphorylated sumoylation motif and regulation by glycogen synthase kinase 3β. Mol. Cell. Biol. 32, 2709–2721 (2012). [Abstract] [Full Text]

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