Figure 9.
Proteasome degradation and decreased activity in the P23H mouse retina rescued by ablation of Atg5. (a) Cytosolic proteins from retinas of P23H and C57BL/6J mice assayed for proteasome peptidase activity over a range of ATP concentrations. Chymotrypsin-like activity was measured as the difference in fluorescence in the presence and absence of the specific inhibitor lactacystin (18 μM). N = 12, 2-way ANOVA. (b) Chymotrypsin-like activities of cytosolic retinal proteins without ATP or with ATP (7 µM) from P23H ATG5Δrod, P23H, and control C57BL/6J (C57) mice. (n = 8), unpaired t-test. (a–b) Data were normalized to chymotrypsin-like activity of the C57BL/6J sample without ATP. (c) Western blots probed for proteasome subunits 20S and PSMC5, and loading control GAPDH, in retinas of P23H ATG5Δrod, P23H, and control C57BL/6J mice. (d) Representative western blots and (e) quantification of autophagosome contents from retinas of P23H GFP-LC3 and control GFP-LC3 mice. Blots were probed for SQSTM1, 20S, and loading control GFP-LC3. (n = 4), unpaired t-test. Data are presented as mean ± SD; *, p < 0.05; **, p < 0.01. All mice were 1 month old.