The Mediator complex subunit MED25 is targeted by the N-terminal transactivation domain of the PEA3 group members

International audiencePEA3, ERM and ER81 belong to the PEA3 subfamily of Ets transcription factors and play important roles in a number of tissue-specific processes. Transcriptional activation by PEA3 subfamily factors requires their characteristic amino-terminal acidic transactivation domain (TAD). However, the cellular targets of this domain remain largely unknown. Using ERM as a prototype, we show that the minimal N-terminal TAD activates transcription by contacting the activator interacting domain (ACID)/Prostate tumor overexpressed protein 1 (PTOV) domain of the Mediator complex subunit MED25. We further show that depletion of MED25 disrupts the association of ERM with the Mediator in vitro . Small interfering RNA-mediated knockdown of MED25 as well as the overexpression of MED25-ACID and MED25-VWA domains efficiently inhibit the transcriptional activity of ERM. Moreover, mutations of amino acid residues that prevent binding of MED25 to ERM strongly reduce transactivation by ERM. Finally we show that siRNA depletion of MED25 diminishes PEA3-driven expression of MMP-1 and Mediator recruitment. In conclusion, this study identifies the PEA3 group members as the first human transcriptional factors that interact with the MED25 ACID/PTOV domain and establishes MED25 as a crucial transducer of their transactivation potential

Identification and characterisation of new partners of the transcription factor ERM

ERM est un facteur de transcription de la famille ETS appartenant au groupe PEA3 qui joue un rôle important dans divers processus biologiques dont la tumorigenèse mammaire. La régulation de son activité transcriptionnelle nécessite des modifications post-traductionnelles et des interactions avec des partenaires. Nous avons mis au point différentes techniques de chromatographie d’affinité dans le but d’identifier de nouveaux partenaires d’ERM. Parmi ceux-ci, trois ont été confirmés comme partenaires directs d’ERM : la protéine CoAA (CoActivator Activator) et les protéines MED23 et MED25. MED23 et MED25 sont des sous-unités du médiateur, complexe qui régule la transcription en intégrant les signaux entre des facteurs de transcription et la machinerie transcriptionnelle. Nous avons démontré que ces protéines interagissent in vitro et in vivo avec ERM et sont nécessaires à l’activation transcriptionnelle induite par ce facteur de transcription. Toutefois, ces sous-unités ont une capacité différente de recrutement du médiateur sur ERM in vitro.La ribonucléoprotéine CoAA régule l’expression de certains gènes et l’épissage alternatif des ARN messagers. CoAA interagit in vitro et in vivo avec ERM. L’activité d’ERM est augmentée par la surexpression de CoAA tandis qu’elle est diminuée par sa sous-expression. Cette activation médiée par CoAA est liée à une modulation du taux de sumoylation d’ERM. Ces travaux ont permis de définir de nouvelles voies de régulation de l’activité transcriptionnelle d’ERM et des autres membres du groupe PEA3. Il reste maintenant à préciser les mécanismes impliqués dans la modulation de l’activité des membres du groupe PEA3 par ces nouveaux interacteurs.ERM is an ETS transcription factor which belongs to the PEA3 group and is involved in several processes such as migration and dissemination during organogenesis and cancer development. Regulation of its transcriptional activity requires post-translational modifications and interactions with partner proteins. In order to identify new ERM partners, we have developed various affinity chromatography techniques to isolate new potential partners. Among these candidates, CoAA (CoActivator Activator), MED23 and MED25 directly interact with ERM.MED23 and MED25 are subunits of the mediator. The mediator is a 30 sub-units multi-protein complex which mediates signals from transcription factors bound at upstream promoter elements or enhancers to RNA polymerase II and the general initiation factors bound at the core promoter. We found that MED23 and MED25 interact with ERM in vitro and in vivo and are required for transcriptional activation induced by ERM. However, these sub-units display various ability to recruit the mediator on ERM in vitro. The heterogeneous nuclear ribonucleoprotein-like protein CoAA regulates gene expression and RNA splicing. We demonstrated that ERM interacts in vitro and in vivo with CoAA. ERM transcriptional activity is enhanced upon CoAA overexpression and is decreased by CoAA knock-down. We demonstrated that CoAA modulates ERM transcriptional activity by decreasing sumoylated ERM levels. This work demonstrated new ways to regulate the activity of ERM and the two other PEA3 group members. The molecular mechanisms involved in the modulation of PEA3 member activity by these partners remain to be clarified

Les facteurs de transcription du groupe PEA3: Régulateurs transcriptionnels dans le processus de cancérisation

Erm, Er81, and Pea3 are the three members of the PEA3 group which belong to the Ets transcription factors family. These proteins regulate transcription of multiple target genes, such as those encoding several matrix metalloproteinases (MMP), which are enzymes degrading the extracellular matrix during cancer metastasis. In fact, PEA3-group genes are often overexpressed in different types of human cancers that also over-express these MMP and display a disseminating phenotype. In experimental models, regulation of PEA3 group member expression has been shown to influence the metastatic process, thus suggesting that these factors play a key role in metastasis. © John Libbey Eurotext.SCOPUS: re.jinfo:eu-repo/semantics/publishe

The Ets transcription factors of the PEA3 group: Transcriptional regulators in metastasis

The PEA3 group is composed of three highly conserved Ets transcription factors: Erm, Er81, and Pea3. These proteins regulate transcription of multiple genes, and their transactivating potential is affected by post-translational modifications. Among their target genes are several matrix metalloproteases (MMPs), which are enzymes degrading the extracellular matrix during normal remodelling events and cancer metastasis. In fact, PEA3-group genes are often over-expressed in different types of cancers that also over-express these MMPs and display a disseminating phenotype. Experimental regulation of the synthesis of PEA3 group members influences the metastatic process. This suggests that these factors play a key role in metastasis. © 2006 Elsevier B.V. All rights reserved.SCOPUS: re.jinfo:eu-repo/semantics/publishe

TRP channel–associated factors are a novel protein family that regulates TRPM8 trafficking and activity

International audienceTRPM8 is a cold sensor that is highly expressed in the prostate as well as in other non-temperature-sensing organs, and is regulated by downstream receptor-activated signaling pathways. However, little is known about the intracellular proteins necessary for channel function. Here, we identify two previously unknown proteins, which we have named "TRP channel-associated factors" (TCAFs), as new TRPM8 partner proteins, and we demonstrate that they are necessary for channel function. TCAF1 and TCAF2 both bind to the TRPM8 channel and promote its trafficking to the cell surface. However, they exert opposing effects on TRPM8 gating properties. Functional interaction of TCAF1/TRPM8 also leads to a reduction in both the speed and directionality of migration of prostate cancer cells, which is consistent with an observed loss of expression of TCAF1 in metastatic human specimens, whereas TCAF2 promotes migration. The identification of TCAFs introduces a novel mechanism for modulation of TRPM8 channel activity