Fig. 4: Lysine 403 acetylation is regulated by Sirt1 and Sirt2.
From: Acetylation-dependent coupling between G6PD activity and apoptotic signaling
a The effect of acetylation on G6PD activity is maintained following the inhibition of endogenous KDACs. Bars represent relative Vmax of acetylated G6PD variants expressed in HEK293T cells cultured in the presence or absence of indicated KDACi. The activity was measured in cleared total cell lysates, corrected to G6PD expression level, and normalized to the activity of pWT expressed without KDACi treatment. Data were analyzed using one-way ANOVA followed by Tukey’s post hoc test and are presented as mean values ± SD; n = 4 (AcK171), 5 or 6 (AcK403), 6 (AcK89), or 9 (pWT) biologically independent samples. b Lysine 403 acetylation is stabilized upon inhibition of endogenous NAD+-dependent deacetylases. Representative Western blots showing the acetylation level of immunoprecipitated AcK89 and AcK403 G6PD, expressed in HEK293T cells cultured in the presence or absence of indicated KDACi. c The inhibitory effect of K403 acetylation is alleviated by NAD+-dependent deacetylases in vitro. Bars represent the relative Vmax of recombinant G6PD incubated with cleared lysates of HEK293T cells pretreated with indicated KDACi [mean ± SD; n = 4 (AcK89), or 5 (AcK403) independent experiments]. Western blots show the amount of G6PD (anti 6×His) and acetylation level following incubation with cleared cell lysates. d Lysine 403-acetylated G6PD is a substrate of Sirt1 and Sirt2. Western blots show the acetylation level of recombinant WT, AcK89, and AcK403 G6PD, following incubation with immunoprecipitated Sirt1, Sirt2, or HDAC6, in the presence or absence of indicated KDACi. e Sirt1 and Sirt2 reactivate AcK403 G6PD. Bars represent the relative Vmax of recombinant WT and AcK403 G6PD, following incubation with deacetylases as described in d. Data are the mean ± SD; n = 3 (Sirt1 treatment), or 7 (Sirt2, HDAC6 treatment) independent experiments. f Sirt1 and Sirt2 deacetylate AcK403 in a time-dependent manner. Representative Western blots showing K403 acetylation level as a function of incubation time with immunoprecipitated Sirt1, Sirt2, or HDAC6. Source data are provided as a Source Data file.
