Fig. 4: Subtle alteration of ER homeostasis in the cerebellum of SEL1LS658P KI mice.
From: SEL1L-HRD1 interaction is required to form a functional HRD1 ERAD complex
a Phos-tag-based Western blot analysis of IRE1α phosphorylation (top) and RT-PCR of Xbp1 splicing in the cerebellum of 5-week-old mice. 0/p, non-/phosphorylation; u/s, un-/spliced Xbp1. Quantitation of IRE1α and the ratio of spliced to total Xbp1 shown below the gel as mean (n = 4-5 mice per group). Livers from mice injected with tunicamycin (TM, an ER stress inducer) included as a positive control. b Western blot analysis of PERK and eIF2α phosphorylation in the cerebellum of 5-week-old mice, with quantitation of the ratio of phosphorylated to total PERK (n = 5, 4 mice for WT and KI) or eIF2α (n = 4, 5 mice for WT and KI) shown below. c RT-PCR and Western blot analyses of CHOP mRNA (n = 4, 6 mice for WT and KI) and protein levels (n = 3 mice for WT and KI) in the cerebellum of 5-week-old mice, with quantitation shown below. d Western blot analysis of ER chaperones GRP94, BiP and PDI in the cerebellum of 5-week-old mice with quantitation shown below (GRP94/BiP: n = 6, 7 mice for WT and KI; PDI: n = 4 mice for WT and KI). e, f Representative TEM images of Purkinje cells (outlined by red dotted lines) of 3- (e) and 24- (f) week-old mice. N, nucleus; M, mitochondria. n = 2 mice each genotype at each age. g Western blot analysis of pro- and cleaved Caspase 3 in the cerebellum of 5- and 24-week-old mice. Thymus, a positive control. n = 3 mice each genotype. Values, mean ± SEM. *p < 0.05, **p < 0.01, ***p < 0.001 and ****P < 0.0001 by two-tailed Student’s t test (b–d).
