Effects of genetic variants in the TSPO gene on protein structure and stability
Fig 5
Analysis of protein stability by the cycloheximide protein degradation assay of wt and mutant TSPO proteins.
(A) TSPO is localized to mitochondria in primary dermal fibroblasts. (B) Lysates of human primary fibroblast cells were subjected to Western blotting to detect endogenous TSPO. (C) Fibroblasts from carriers of wt and homozygous A147T and R162H mutations of TSPO were treated with cycloheximide (25 μg/ml) for the indicated times, and endogenous TSPO was then detected by specific rabbit anti-TSPO antibody. Equal amounts of protein were subjected to Western blot analyses, as determined by comparing the amount of beta 1 tubulin. (D) Densitometry results for endogenous TSPO after treatment with cycloheximide were quantified as percentage of the initial TSPO protein level (0h of CHX treatment) and normalized to the intensity of beta 1 tubulin, and then plotted. Data are shown as mean ± SEM from three independent experiments.