AZGP1 is a tumor suppressor in pancreatic cancer inducing mesenchymal-to-epithelial transdifferentiation by inhibiting TGF-Β-mediated ERK signaling

Article (PDF Available)inOncogene 29(37):5146-58 · September 2010with38 Reads
DOI: 10.1038/onc.2010.258 · Source: PubMed
Abstract
Epithelial-to-mesenchymal transdifferentiation (EMT) mediated by transforming growth factor-β (TGF-β) signaling leads to aggressive cancer progression. In this study, we identified zinc-α2-glycoprotein (AZGP1, ZAG) as a tumor suppressor in pancreatic ductal adenocarcinoma whose expression is lost due to histone deacetylation. In vitro, ZAG silencing strikingly increased invasiveness of pancreatic cancer cells accompanied by the induction of a mesenchymal phenotype. Expression analysis of a set of EMT markers showed an increase in the expression of mesenchymal markers (vimentin (VIM) and integrin-α5) and a concomitant reduction in the expression of epithelial markers (cadherin 1 (CDH1), desmoplakin and keratin-19). Blockade of endogenous TGF-β signaling inhibited these morphological changes and the downregulation of CDH1, as elicited by ZAG silencing. In a ZAG-negative cell line, human recombinant ZAG (rZAG) specifically inhibited exogenous TGF-β-mediated tumor cell invasion and VIM expression. Furthermore, rZAG blocked TGF-β-mediated ERK2 phosphorylation. PCR array analysis revealed that ZAG-induced epithelial transdifferentiation was accompanied by a series of concerted cellular events including a shift in the energy metabolism and prosurvival signals. Thus, epigenetically regulated ZAG is a novel tumor suppressor essential for maintaining an epithelial phenotype.
    • "TritonX-100/PBS for one hour at RT, incubated with the primary antibodies, overnight at 4°C. For the TGFβ signaling cascade, the following antibodies were purchased commercially: anti-Smad2 (D43B4) rabbit mAb (1:100 dilution, Cat#5339) [19], anti-Smad2/3 (D7G7) Rabbit mAb (1:100 dilution, Cat# 8685) [20], anti-Smad3 (C67H9) rabbit mAb (1:100 dilution, Cat# 9523) [21], anti-Smad4 (cat#9515) [22], anti-Phospho-Smad2 (Ser465/467) (138D4) rabbit mAb (1:100 dilution, cat#3108) [23], and anti-phospho-Smad3 (Ser423/425) (C25A9) rabbit mAb (1:100 dilution, Cat#9520) [24] from Cell Signaling Technology, Beverly, MA. For cell proliferation activity, the following antibodies were purchased commercially: anti-Proliferating cell nuclear antigen (PCNA) PCNA (1:2400 dilution, cat# #8607) [25], anti-Histone H3 (D1H2) rabbit mAb (1:100 dilution, Cat#4499) [26], anti-phospho-Histone H3 (Thr3) (cat#9714) [ 27]; anti-phospho-Histone H3 (Thr11) (cat#9764) [ 28], anti-phospho-Histone H3 (Ser28) (1:100 dilution, cat# 9713) [29] from Cell Signaling Technology, Beverly, MA. "
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