DNAJB6 governs a novel regulatory loop determining Wnt/β-catenin signalling activity

DKK1 (dickkopf 1 homologue) is a secreted inhibitor of the Wnt signalling pathway and a critical modulator of tumour promotion and the tumour microenvironment. However, mechanisms regulating DKK1 expression are understudied. DNAJB6 {DnaJ [HSP40 (heat-shock protein 40 kDa)] homologue, subfamily B, member 6} is an HSP40 family member whose expression is compromised during progression of breast cancer and melanoma. Inhibition of the Wnt/β-catenin signalling pathway by up-regulation of DKK1 is one of the key mechanisms by which DNAJB6 suppresses tumour metastasis and EMT (epithelial-mesenchymal transition). Analysis of the DKK1 promoter to define the cis-site responsible for its up-regulation by DNAJB6 revealed the presence of two binding sites for a transcriptional repressor, MSX1 (muscle segment homeobox 1). Our investigations showed that MSX1 binds the DKK1 promoter and inhibits DKK1 transcription. Interestingly, silencing DNAJB6 resulted in up-regulation of MSX1 concomitant with increased stabilization of β-catenin. ChIP (chromatin immunoprecipitation) studies revealed that β-catenin binds the MSX1 promoter and stabilization of β-catenin elevates MSX1 transcription, indicating that β-catenin works as a transcription co-activator for MSX1. Functionally, exogenous expression of MSX1 in DNAJB6-expressing cells promotes the mesenchymal phenotype by suppression of DKK1. Thus we have identified a novel regulatory mechanism of DNAJB6-mediated DKK1 transcriptional up-regulation that can influence EMT. DKK1 is a feedback regulator of β-catenin levels and thus our studies also define an additional negative control of this β-catenin/DKK1 feedback loop by MSX1, which may potentially contribute to excessive stabilization of β-catenin.