Figure 4
Cellular localization of aminopeptidase P1 in hippocampal neurons. (a) Schematic diagram of the bicistronic expression vector. (b) Cultured hippocampal neurons were transfected with the bicistronic expression vector using a calcium phosphate method. The dendrites of transfected (EGFP-positive) and neighboring untransfected (EGFP-negative) neurons were visualized by MAP2. Exogenously expressed aminopeptidase P1 protein was mainly distributed in the soma and dendrites of cultured hippocampal neurons. Arrows indicate MAP2-negative axons of transfected neurons. Scale bars, 20 μm. (c) Distribution of aminopeptidase P1 in subcellular brain fractions. Endogenous aminopeptidase P1 was detected in all fractions, with the strongest signal observed in the cytosolic fractions (S2 and S3). Whole brain homogenates from WT and Xpnpep1–/– mice were used to determine the specificity of the aminopeptidase P1 antibody. S1, supernatant after P1 sedimentation; P1, crude nuclear fraction; S2, supernatant after P2 sedimentation; P2, crude synaptosomal pellet; S3, cytosolic; P3, light membranes; LP1, synaptosomal membranes.
