Disabled-2; an autophagic and apoptotic switch

Epithelial to mesenchymal transition (EMT) is postulated to be required for the initiation of tumor cell metastasis.¹ Disabled-2 (Dab2) has been shown to play an essential role during TGFβ-mediated EMT;² yet, its expression is often lost in highly metastatic cell lines.³ Our recent work (highlighted herein) provides mechanistic clues for this apparent paradox.⁴ We observe that following TGFβ-mediated EMT in normal murine mammary epithelial (NMuMG) cells, prolonged and chronic TGFβ treatment leads to the proteolytic degradation and loss of Dab2 by cathepsin-B (CTSB), loss of the mesenchymal phenotype and the induction of autophagy, which promotes tumor metastasis and chemoresistance to the cytotoxic effects of doxorubicin. CTSB inhibition or expression of a CTSB-resistant mutant Dab2 will shift long-term TGFβ-treated cells from autophagy to apoptosis while maintaining chemosensitivity to doxorubicin. Mechanistically, Dab2 interacts with Beclin-1 and promotes its CK2-dependent phosphorylation at Ser337 and/or Ser341, thereby preventing Beclin-1/Vps34 interaction and initiation of autophagosome formation. This mechanism further stabilizes Bim by attenuating ERK-mediated Bim phosphorylation to promote apoptosis (Fig. 1). Thus, CTSB-modulated Dab2 expression plays an important role in metastasis and doxorubicin-mediated chemosensitivity.

Figure 1.

Role of Disabled-2 (Dab2) in TGFβ-mediated autophagy. Chronic TGFβ treatment induces the expression of the protease cathepsin B (CTSB) leading the cleavage and degradation of wild-type Dab2 (WT Dab2). Loss of Dab2 allows Beclin-1/Vps34 interactions, initiation of the autophagosome formation and an increase in autophagy. CTSB attenuation or expression of a non-CTSB cleavage mutant form of Dab2 (Mut. Dab2) facilitates CK2-mediated Beclin-1 phosphorylation resulting in loss of Beclin-1/Vps34 association, autophagosome platform dissociation, and subsequent inhibition of autophagy. Dab2 prevents ERK-mediated Bim phosphorylation and degradation and stabilizes TGFβ-induced Bim expression to promote apoptosis.

Autophagy plays a dual role in cancer. It is thought to prevent tumor initiation by inhibiting inflammation, chronic tissue damage, and genome instability, and yet it has also been shown to promote the growth of established tumors and contribute chemoresistance.⁵ Our data⁴ demonstrates that once cells have undergone EMT and are further exposed to chronic TGFβ, the subsequent loss of Dab2 promotes autophagy. Modulating CTSB and Dab2 expression levels in cells governs whether the autophagic or apoptotic pathways are activated. In NMuMG cells, preventing Dab2 loss through silencing of CTSB or expression of a non-cleavable Dab2 promotes apoptosis upon chronic TGFβ treatment. Loss of Dab2 through CTSB overexpression promotes autophagy in response to chronic TGFβ signaling. We also observed that cell lines with high CTSB and low Dab2 expression are more metastatic when injected into the mammary fat pads of female mice, suggesting that Dab2 is inhibitory for autophagy induction and confirming the contribution of autophagy to metastasis. Our data also supports a role for autophagy in the acquisition of chemoresistance. Cells expressing the non-cleavable Dab2 mutant are more sensitive to the cytotoxic effects of doxorubicin, while cells overexpressing CTSB acquire enhanced chemoresistance to doxorubicin. Modulating Dab2 levels in these cells or expressing the CK2-phosphomutant form of Beclin-1 regulates chemosensitivity. High Dab2 levels promotes doxorubicin sensitivity whereas low Dab2 expression or expression of a mutant Beclin-1 promotes doxorubicin resistance. Based on these data we conclude that autophagy induction and its regulation by Dab2 contribute to metastasis and chemoresistance.

Dab2 expression levels have previously been shown to have significant impact on cellular differentiation and cancer development. Dab2 expression is upregulated and essential during retinoic acid (RA)-induced differentiation of F9 teratocarcinoma cells into visceral endoderm-like cells⁶ and during TGFβ-mediated EMT in mammary epithelium.² In contrast, aberrantly low levels of Dab2 have been found in highly metastatic cell lines,³ and its ectopic expression in these cancer lines has been shown to result in cell cycle arrest and a decreased rate of cell proliferation. As such, Dab2 has been assigned the label of a tumor suppressor gene.⁷ CTSB-mediated proteolysis and loss of Dab2 may therefore represent a key regulatory checkpoint for differentiated cells. Initially, TGFβ-induced Dab2 may be required for differentiation; however, upon chronic exposure of differentiated cells to TGFβ, Dab2 expression is targeted as it mediates the checkpoint switch between autophagic survival and apoptotic cell death. Upregulation of CTSB expression, upon chronic TGFβ exposure, may represent the molecular mechanism targeting Dab2, thereby promoting cellular survival and tumorigenesis.

Disclosure of potential conflicts of interest

No potential conflicts of interest were disclosed.