Fig. 4

Inhibition of CatD slows the catabolism of tau in vitro and in cultured cells. (A) Representative Coomassie blue-stained polyacrylamide gel loaded with recombinant human tau (rTau) incubated for the indicated times with recombinant human CatD (5 nM) in the absence or presence of equal concentrations (1 µM) of Aβ40 or Aβ42. (B) Quantification of rTau levels as a function of time in 4 independent experiments. Note how rTau catabolism is unaffected by Aβ40 but markedly slowed by Aβ42, a potent competitive inhibitor of CatD. Data are mean ± SEM; n = 4. (C) Overview of the experimental approach used to quantify hTau catabolism in “Tet-Off” cultured neuroblastoma cells (see main text). (D) Representative western blot showing hTau levels (stained with antibody P44) at different time points after cessation of hTau expression in the absence or presence of the CatD inhibitor, pepstatin A (PepA; 1 µM). (E) Quantitation of hTau levels as a function of time from 6 independent experiments. Note the marked increase in the half-life of hTau in the presence of PepA (0.98 days; 95% CI 0.80 to 1.25) relative to DMSO-treated controls (0.51 days; 95% CI 0.429 to 0.627; P = 0.0012). Data are mean ± SEM, n = 6