Accumulation of aggregated htt is correlated with the decrease in synaptic proteasome activity in R6/2 mouse brain. (A) EM48 immunostaining showed more neuropil aggregates in R6/2 mice at 10 wk than at 4 wk. Bars, 10 μm. (B) Mouse brain extracts were fractionated to isolate cytosolic, nuclear, and synaptosomal fractions. Synaptosomal fraction is enriched for the synaptic protein synaptophysin but not the cytoplasmic protein GAPDH or the nuclear transcription factor TFIIB. Antibodies to proteasomal subunits LMP2, LMP7, and core subunits (a5, a7, b1, b5i, and b7) of 20S also detected the presence of these subunits in synaptosomes. (C) The chymotrypsin-like activity (mean + SEM, n = 4–5) of different fractions. (D) EM48 Western blotting showed more aggregated htt in synaptosomes (S) from younger mice at 4 wk of age. T: total lysates, C: cytosolic fraction, N: nuclear fraction. (E) The chymotrypsin-like activity (mean +SEM, n = 5–7) of total lysates (left) and synaptosomes (right) isolated from the striatum of R6/2 mice at 4 and 10 wk of age. Synaptosomal chymotrypsin-like activity in the striatum was lower than from the cortex, and synaptosomes isolated from HD brains also showed lower proteasomal activity than those from wild-type mouse brains. *, P < 0.05 as compared with wild type.