Phosphorylation Regulates the Function of ZEB1 Transcriptional Repressor

PERRONE A; LLORENS DE LOS RIOS C; CABANILLAS, ANA MARIA

Buenos Aires

Congreso; IXL Reunión Anual Sociedad Argentina de Investigación Bioquímica y Biología Molecular (SAIB); 2013

SAIB

ZEB1 is a transcription factor important for the epithelial-mesenchymal transition by which many tumors undergo metastasis.ZEB1 is phosphorylated but the role of phosphorylation(P) is unknown.Our studies show that a decrease in ZEB1 P increases both DNA binding and transcriptional repression of ZEB1 target genes and that its ZD2 domain holds 4 phosphosites (T851,S852,S853,T867).Functional assays with ZD2 mutants show that PMA, or IGF-1 (mediated by ERK) can prevent the transcriptional role of ZD2. GFP-ZD2 clones show that IGF-1 disrupted ZEB1 nuclear localization.Our aim is to expand the significance of P to full length ZEB1 (FLZEB1) and the importance of T867 site.FLZEB1 mutated at all 4 sites repressed more the activity of gene targets than wtZEB1, which was not obtained by a single mutation T867A suggesting that the 4 sites are needed and a cooperative activation among them.Immunofluorescence of CHO cells transfected with ZEB1-T867A or ZEB1-T851A/S852A/S853A/T867A show the mutants are unresponsive to IGF-1 (they remained nuclear) and that T867 controls cellular location of ZEB1.Transfections assays with ZD2-T867A, ZD2-T867E (P-mimetic) and ZEB1-T867A show that IGF-1 induces its effect through T867 and that ZEB1 has other IGF-1 responsive phosphosites.The results confirmed that ZEB1 biological role is regulated by P. ZEB1 would serve as an integrating factor of external signals