Figure 5.

No UPS dysfunction in presenilin-null cells. A, No differences in ubiquitinated proteins by immunoblot were apparent between wild-type [control (Con)] and presenilin-null [double knock-out (DKO)] fibroblasts (100 ± 11.88 vs 76.88 ± 17.56). Quantification of Western blots was performed by densiometric analysis and is presented as percentage of control, normalized to actin. n = 4; unpaired Student's t test. B, C, Confocal immunofluorescent microscopy reveals no difference in ubiquitinated proteins in wild-type (control) and PSDKO fibroblasts. Scale bar, 29 μm. D, E, Transfection with the proteasome reporter ZsProsensor shows no difference in fluorescence intensity between wild-type and presenilin-null cells by confocal microscopy. F, To assess proteasome activity, lysates of wild-type (control) and presenilin-null fibroblasts were exposed to three proteasome substrates (LLVY-AMC for the chymotrypsin-like, VGR-AMC for the trypsin-like, and LLE-AMC for the PGPH/caspase-like activity). AMC release was measured and rate of release was quantified as AFU/90 s, revealing no significant differences in proteasome activities between the two cell types. Error bars represent ± SEM; n = 3; unpaired Student's t test.