The 26S proteasome system degrades the ERM transcription factor and regulates its transcription-enhancing activity

Abstract

ERM is a member of the ETS transcription factor family. High levels of the corresponding mRNA are detected in a variety of human breast cancer cell lines, as well as in aggressive human breast tumors. As ERM protein is almost undetectable in these cells, high degradation of this transcription factor has been postulated. Here we have investigated whether ERM degradation might depend on the proteasome pathway. We show that endogenous and ectopically expressed ERM protein is short-lived protein and undergoes proteasome-dependent degradation. Deletion mutagenesis studies indicate that the 61 C-terminal amino acids of ERM are critical for its proteolysis and serve as a degradation signal. Although ERM conjugates with ubiquitin, this post-translational modification does not depend on the C-terminal domain. We have used an Ets-responsive ICAM-1 reporter plasmid to show that the ubiquitin-proteasome pathway can affect transcriptional function of ERM. Thus, ERM is subject to degradation via the 26S proteasome pathway, and this pathway probably plays an important role in regulating ERM transcriptional activity.

The E3 ubiquitin ligase complex component COP1 regulates PEA3 group member stability and transcriptional activity

“…A major ERM species was observed at approximately 85 kDa in the Ni 2 þ -bound protein fraction as a result of Ub overproduction ( Figure 4a). As described earlier, it corresponds to a monoubiquitylated form of ERM (Baert et al, 2007). Higher molecular mass species, characteristic of polyubiquitylation (Baert et al, 2007), were also observed (Figure 4a, compare lanes 1 and 2).…”

“…As described earlier, it corresponds to a monoubiquitylated form of ERM (Baert et al, 2007). Higher molecular mass species, characteristic of polyubiquitylation (Baert et al, 2007), were also observed (Figure 4a, compare lanes 1 and 2). In the presence of COP1, a slight increase in monoubiquitylation of ERM was observed, but polyubiquitylation was not enhanced (compare lanes 2 and 3).…”

“…To regulate their target genes, these transcription factors are subjected to multiple post-translational modifications such as phosphorylation and acetylation (de Launoit et al, 2006). Recently, it has been shown that they can also be posttranslationally covalently linked to polypeptides such as ubiquitin (Ub) (Takahashi et al, 2005;Baert et al, 2007) and SUMO (Degerny et al, 2005;Gocke et al, 2005). We thus have shown that SUMO modification of ERM negatively regulates its transcriptional activity without affecting DNA-binding or subcellular localization (Degerny et al, 2005).…”

“…ERM and the PEA3 group members interact with COP1 through two in-tandem conserved motifs ERM is subjected to Ub modification (Baert et al, 2007), and possesses in its sequence two consensusbinding motifs (D/ED/EXXXVPD/E) for the E3 Ub ligase COP1 (Yi and Deng, 2005). To test whether this transcription factor can interact with COP1, we first expressed ERM as a glutathione-S-transferase (GST)-fusion protein and tested the ability of GST-ERM to bind to COP1.…”

“…COP1 cooperates with DET1 to regulate ubiquitylation and protein level of the PEA3 group members It has been recently shown in vivo that ERM is subjected to ubiquitylation followed by degradation (Baert et al, 2007). We thus tested whether the expression of COP1 induced a variation of the ubiquitylation level of ERM.…”