Role of pyrophosphatase activity on PPA1-induced inhibition of neuronal differentiation in N1E115 cells.

<p>(A) Recombinant wild-type PPA1 (his-PPA1) and PPA1 Asp117Ala (his-PPA1 D117A) proteins. Western blot using anti-PPA1 antibody showed that the his-PPA1 and his-PPA1 D117A protein amount is similar. (B) Recombinant protein pyrophosphatase activity. Pyrophosphatase activity was seen in his-PPA1 while no activity was detected in his-PPA1 D117A (n = 5). (C) Effect of either green fluorescence protein (AD-GFP), wild-type PPA1 (AD-PPA1), or PPA1 D117A (AD-PPA1 D117A) overexpression using adenoviral vector, on PPA1 protein levels in N1E115 cells treated with 1 mM valproic acid (VPA). (D) Typical morphological changes in N1E115 cells treated with VPA, using AD-GFP overexpression as a control (VPA+GFP), AD-PPA1 (VPA+AD-PPA1) or D117A (VPA+AD-PPA1 D117A). (E) Quantitative analysis of the neurite growth ratio is shown. Neurite growth is determined by morphological analysis as described in the Experimental procedure (n = 100). *p<0.05 vs. his-PPA1, #p<0.05 vs. N1E115 cells treated with VPA+GFP, a<0.05 vs. VPA+Ad-PPA1.