Fig 1: Phosphorylation of GluA1 MPR, Seri831, and Ser845.A, phosphorylation of the GluA1 MPR by NMDA stimulation. Cultured hippocampal neurons were treated with 50 µM NMDA for 10 min and immunoprecipitated using the anti-4.1N antibody after solubilization. The amount of immunoprecipitated 4.1N and coimmunoprecipitated GluA1 was analyzed by immunoblot analysis. The intensities of the bands corresponding to GluA1 (top) and 4.1N (bottom) in the immunoprecipitated (IPed) fraction were normalized to the intensity of the respective molecule in the input fraction. Data are presented as the mean + SEM and individual data points (Mann–Whitney U test, **p < 0.01; n = 5–6). B, Ser831 phosphorylation by CaMKII was not affected by mutations in the MPR. GST fusion proteins with C termini of GluA1wt (GluA1CTwt), GluA1AA (GluA1CTAA), and GluA1DD (GluA1CTDD) were phosphorylated by CaMKII in vitro and analyzed by the immunoblot analysis using anti-phospho-Ser831 GluA1 (top) and anti-GST (bottom) antibodies. Data are presented as the mean + SEM and individual data points (Kruskal–Wallis test; n = 5). C, Ser845 phosphorylation by PKA was not affected by mutations in the MPR. GluA1CTwt, GluA1CTAA, and GluA1CTDD were phosphorylated in vitro by PKA and analyzed by immunoblot analysis using anti-phosphor-Ser845 GluA1 and anti-GST antibodies. Data are presented as the mean + SEM and individual data points (Kruskal–Wallis test; n = 5). CaMKII, calmodulin-dependent protein kinase II; MPR, membrane-proximal region; n.s., not significant; NMDA, N-methyl-d-aspartate.