Fig. 1
From: Reduced chronic restraint stress in mice overexpressing hyperactive proteasomes in the forebrain
Enhancing proteasome activity in the brain may lessen chronic restraint stress (CRS). a A genetic scheme of neuron-specific expression of open-gated proteasomes. A flag-tagged form of α3 mutant, with a deleted N-terminal 9-residue tail (α3ΔN), was overexpressed by CaMKIIα promoter-driven transcriptional activator (tTA), in the absence of doxycycline (dox). b mRNA levels of α3ΔN from CaMKIIα-tTA/TRE-α3ΔN double transgenic (α3ΔN Tg) mice were measured with quantitative RT-PCR. Different brain regions were analyzed, which revealed little expression of α3ΔN in the cerebellum. Liver tissue was used as a negative control. Transgene expression was virtually completely abrogated with the administration of dox in drinking water. β-Actin was used as the loading control. c As in (b), except that α3ΔN proteins were analyzed with SDS-PAGE/immunoblotting (IB), using whole cell lysates from the indicated regions of the brain. The level of α3ΔNflag protein was largely proportional to that of α3ΔN mRNA. d Proteasome activity in whole brain extracts, from wild-type (WT) and α3ΔN Tg mice, was measured using the fluorogenic suc-LLVY-AMC reporter substrate (mean ± SD from three independent experiments). **, p < 0.01 (two-tailed Student’s t test). e Immobility times in the forced swimming test before CRS and post-CRS day 14 (2 h per day). N = 5 for WT, N = 9 mice for Tg. **, p < 0.01 (two-tailed Student’s t test) N.S., not significant. f Less accumulation of polyubiquitin conjugates and phosphorylated IRE1α in the hippocampus of α3ΔN Tg mice than in WT controls. Cortex and hippocampus were isolated after 14 d of CRS and subjected to SDS-PAGE/IB analysis, using the indicated antibodies