Through physical interactions between various transcription factors and specific Med subunits, the Mediator complex acts as an integrative hub for channeling different signaling pathways to direct diverse gene programs and biological functions (Malik and Roeder, 2010). Transforming growth factor beta (TGFβ) signaling plays a critical role in the progression of human cancer (Bierie and Moses, 2006), and the Smad2/3 co-regulatory factors can also contribute to TGFβ functions in development and tumorigenesis (Kato et al., 2002; Kim et al., 2003; Iyer et al., 2004). Here, we show that Mediator Med15 deficiency attenuates the TGFβ-targeted gene expression, relieves the TGFβ-mediated growth inhibition, and hinders the TGFβ-induced epithelial–mesenchymal transition (EMT), through the reduction of the phosphorylation and nuclear accumulation of Smad2/3. Furthermore, the Smad3-binding domain was mapped to a short region of MED15, which is important for Smad3-dependent transcriptional activation. Interestingly, MED15 is also highly expressed in clinical cancer tissues correlated with hyperactive TGFβ signaling, as indicated by Smad3 phosphorylation. Moreover, Med15 RNAi decreased the metastatic potential of a highly aggressive breast cancer cell line by reducing TGFβ/Smad signaling. Collectively, these data suggest that the Mediator MED15 regulates TGFβ/Smad signaling and breast cancer cell metastasis; thus it may serve as a potential therapeutic target.
To determine the function of Med15 in TGFβ signaling, we first used retrovirus-based shRNA to knock down Med15 in HaCaT cells, an immortalized human keratinocyte cell line that is often used in investigations of TGFβ signaling. Western blot confirmed the effects of RNAi (Figure 1A). Med15 RNAi attenuated the TGFβ-induced gene expression, as reflected by the TGFβ target genes PAI-1 and p21 (Figure 1B).