Extended Data Fig. 7: Expression of HMGCR(K248R) by AAV or knock-in reverses the phenotypes of L-Usp20−/− mice.
From: Feeding induces cholesterol biosynthesis via the mTORC1–USP20–HMGCR axis
a, HMGCR(K248R) was resistant to sterol-induced degradation. +: 0.3 μg ml-1 25-HC and 3 mM Mev; ++: 1 μg ml-1 25-HC and 10 mM Mev. The experiments were carried out as described in Fig. 1d. b, USP20 did not increase the level of HMGCR(K248R). 1 μg ml-1 25-HC and 10 mM Mev were used. c-j, Eight-week-old male L-Usp20−/− mice and their male WT littermates were injected with 1 × 1010 viral genome (v.g.) of AAV-HMGCR(K248R) or AAV-control (Ctr). After HFHS diet for 8 weeks, the mice were analysed. c, Immunoblotting analysis of liver samples. d, Body weight (n = 3 for WT injected with AAV-Ctr, n = 4 for other groups). e, Total cholesterol in serum (n = 3 for WT injected with AAV-Ctr, n = 4 for other groups). f, Triglyceride in serum (n = 3 for WT injected with AAV-Ctr, n = 4 for other groups). g, h, Whole-body oxygen consumption of mice over dark and light cycles (n = 3 for WT injected with AAV-Ctr, n = 4 for other groups). i, GTT (n = 3 for WT injected with AAV-Ctr, n = 4 for other groups). j, Strategy to generate HmgcrKI/KI mice expressing HMGCR(K248R). k–p, Eight-week-old male mice were fed the HFHS diet for 7 weeks and then subjected to fasting-refeeding treatment. k, western blotting analysis of liver samples. l, Total cholesterol in serum (n = 3 per group). m, Triglyceride in serum (n = 3 per group). n, o, Whole-body oxygen consumption of mice (n = 4 per group). p, GTT (n = 5 per group). Each value represents mean ± SEM. Experiments in (a–c, k) were performed as indicated twice with similar results. Statistical significance was determined using unpaired two-tailed Student’s t-test (d–f, h, i (right), l, m, o, p (right)); or two-way ANOVA (g, i (left), n, p (left)). F: fasted; R: refed.
