Role of signaling pathways in N-ZEB1 phosphorilation and in Epitelial-Mesenchymal Transition (EMT)

LLORENS DE LOS RIOS C; CAVALLO N; CABANILLAS, AM

Rosario, Santa Fe

Congreso; L Reunión Anual de la Sociedad Argentina de Investigación en Bioquímica y Biología Molecular SAIB; 2014

SAIB

ZEB1 (Zn Finger E-box Binding Homeobox) is a transcription factor that mediates the EMT and invasion. It contains 2 Zn finger domains (ZD1-ZD2) which bind DNA independently. ZEB1 expresses as two phosphorylated (P) forms important for its biological role. We had characterized the role of phosphorylated C-term ZD2. Our goal is to uncover the role of signaling pathway/s on N-term ZD1 (N-ZEB1) biological function on EMT. IP assays followed by WB with antiP-substrates Ab revealed that N-ZEB1 is modified by PI3K(AKT), MAPK and PKC pathways. IGF-1 induced relocation of a nuclear N-ZEB1 GFP clone harboring first 490aa (eGFPZ1) into the cytosol of CHO-K1 cells; a 2nd clone, eGFPZ2 (included in the former) was unresponsive to IGF-1. PMA/IONO treatment relocated both clones in the cytosol. NMuMG cells were used to investigate the role of N-ZEB1 in EMT.  These epithelial cells, that can turn into mesenchymal under various treatments, were stably transfected with eGFPZ2 or empty vector (control). The epithelial markers E-cadherin and actin assessed by WB and confocal microscopy were diminished or absent in eGFPZ2 cells. Also, cell proliferation and cell migration were significantly increased (P<0.001 and  P<0.05, respectively) in eGFPZ2 cells vs. controls. We have identified a small fragment in ZEB1 that induces EMT by itself. The activity of N-ZEB1 could be regulated by IGF-1 and PKC effectors.