Fig. 4

HDAC7 promotes β-catenin deacetylation, dephosphorylation and nuclear transport, whereas melatonin interrupts above actions. a and b Co-IP analysis was performed to determine the interaction between HDAC7 and β-catenin in both EC109 and HEK-293T cells. HDAC7 or β-catenin proteins were respectively immunoprecipitated by the anti-Flag antibody followed by target proteins detection using Western blotting analyses. c Cytosolic fractionation and immunoblot assays were applied to detect β-catenin nuclear import after overexpressing HDAC7 by infecting LV-Flag-HDAC7 or melatonin treatment in EC109 cells; GAPDH and Lamin B1 were respectively cytoplasmic and nucleus loading controls. d and e Melatonin treatment for 48 h enhanced β-catenin phosphorylation (Ser675) and acetylation (Lys49) without affecting total β-catenin and TCF4 levels, whereas HDAC7 overexpression or downregulation could partially hinder or enhance the above melatonin’s actions. Representative Western blotting results of phosphorylated β-catenin (Ser675), acetylated β-catenin (Lys49), β-catenin and TCF4 were shown. HDAC7 histone deacetylase 7, LV lentivirus, MEL melatonin, TCF4 transcription factor 4, OE overexpression