Figure 4.
BACE1 levels and activity are increased in the Hippocampi of aged Gga3−/− mice. A, Western blot analysis of hippocampal lysates from 6-month-old and 18–24-month-old Gga3−/− and Gga3+/+ mice. GGA3 (4167, Cell Signaling Technology), APP (m3.2), BACE1 (D10E5, Cell Signaling Technology), C99 (m3.2), and GAPDH (MAB374, Millipore). B, The graph represents BACE1 levels normalized to GAPDH and reported as arbitrary units. BACE1 levels are elevated 25% (p = 0.001) and 27% (p < 0.0001) in Gga3−/− mice compared with Gga3+/+ littermates at 6 months of age and 18–24 months of age, respectively. #BACE1 levels are reduced 18% (p = 0.0011) in the hippocampus of Gga3+/+ mice over aging. *BACE1 levels are reduced 17% (p < 0.0001) in the hippocampus of Gga3−/− mice over aging. C, The graph represents APP levels normalized to GAPDH and reported as arbitrary units. APP-fl levels are reduced 13% (p = 0.0061) in the hippocampus of Gga3−/− mice compared with Gga3+/+ mice at 18–24 months of age. #APP-fl levels are reduced in the hippocampus of Gga3+/+ mice 21% (p <0.0001) over aging. *APP-fl levels are reduced in the hippocampus of Gga3−/− mice 31% (p < 0.0001) over aging. D, The graph represents C99 levels normalized to APP-fl and reported as arbitrary units. C99 levels are increased 21% (p = 0.0043) in the hippocampus of Gga3−/− mice compared with their Gga3+/+ littermate controls at 18–24 months of age. E, β-Secretase activity assay measured by cleavage of the DABCYL/EDANS-conjugated β-secretase substrate IV measured in 18–24-month-old Gga3+/+ and Gga3−/− mouse hemibrain lysates. Data are reported as percentage change in Relative Fluorescence Units (RFU) of Gga3−/− mice when normalized to Gga3+/+ littermate controls. β-Secretase activity is increased 70% (p = 0.0128) in Gga3−/− mice compared with Gga3+/+ mice at 18–24 months of age. F, The graph represents Aβx-40 levels in Gga3+/+ and Gga3−/− hippocampal lysates at 6 months and 18–24 months of age. Data are reported as percentage change in Aβx-40 when normalized to Gga3+/+ mice at each age. No difference was observed in Aβx-40 between Gga3+/+ and Gga3−/− mice at either age. The data indicate mean ± SEM of 10 Gga3+/+, 9 Gga3−/− (6 month old), 16 Gga3+/+, and 13 Gga3−/− (18–24 month old) mice for APP-fl and C99 levels; 10 Gga3+/+, 9 Gga3−/− (6 month old), 15 Gga3+/+, and 13 Gga3−/− (18–24 month old) mice for BACE1 levels; 8 Gga3+/+ and 12 Gga3−/− 18–24-month-old mice for β-secretase activity; and 13 Gga3+/+, 16 Gga3−/− (6 month old), 11 Gga3+/+, and 13 Gga3−/− (18–24 month old) mice for Aβx-40. Statistical analysis of data was performed using an unpaired t test with Welch correction.