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. 2021 Jun 10;17(6):e1009616. doi: 10.1371/journal.ppat.1009616

Fig 2. NEDD4 can be acetylated by P300.

Fig 2

(A)-(C) NEDD4 acetylation in HEK293T cells was measured following overexpression of the indicated acetyltransferases targeted by “*” (A), treatment with the P300 activator CTB and inhibitors C646 and Curcumin (B), or overexpressing P300-HA and the acetyltransferase-inactive P300-WY-HA in P300 knockout (KO) cells (C). Cell lysates were immunoprecipitated with an anti-Myc antibody and analyzed via immunoblotting with an anti-Acetyl-K antibody to detect the acetylation of NEDD4. (D) NEDD4 acetylation was measured in HEK293T WT and P300 KO cells. (E) In vitro acetylation assays were measured by incubating purified His-NEDD4 with purified GST-P300-HAT followed by immunoblotting with an anti-Acetyl-K antibody to detect the acetylation of NEDD4. (F) Interaction between endogenous P300 and NEDD4. (G) Acetylation of Myc-NEDD4 and mutants was measured in HEK293T cells. (H) HEK293T cells were transfected with the indicated plasmid combinations to detect the influence of P300 on the acetylation of NEDD4 mutants. Error bars, mean ± SD of three experiments. Student’s t test; *p < 0.05; **p < 0.01; ***p < 0.001.