Figure 7 : The N terminus of PrPC is required for the interaction with AMPA subunits.

From: Prion protein facilitates uptake of zinc into neuronal cells

Figure 7

(a) Zinc uptake measured using Newport Green in SH-SY5Y cells expressing wild-type PrPC exposed to 32 μM Zn2+ (blue symbols), 32 μM Zn2+ and 10 μg ml−1 antibody SAF32 (green symbols), 32 μM Zn2+ and 10 μg ml−1 antibody 8H4 (red symbols) or 32 μM Zn2+ and 10 μg ml−1 anti-transferrin receptor antibody (purple symbols). (b) Zinc uptake measured using Newport Green in rat primary hippocampal neurons exposed to 32 μM Zn2+ (blue symbols), 32 μM Zn2+ and 10 μg ml−1 antibody SAF32 (green symbols) or 32 μM Zn2+ and 10 μg ml−1 anti-transferrin receptor antibody (red symbols). Data shown as the relative Newport Green fluorescence corrected against DNA content and are plotted as mean±s.e.m. (n=3). Kruskal–Wallis, P<0.05 for SAF32 compared with no antibody. (c) Untransfected (Un) SH-SY5Y cells or SH-SY5Y cells expressing either wild-type PrPC, PrPΔOct or PrPΔN were exposed to 100 μM Zn2+ and stained using 10 μM Zinpyr-1. Data shown as mean (±s.e.m.). Zinpyr-1 fluorescence corrected against DNA content (n=8). Insert shows expression of wild-type PrPC, PrPΔOct and PrPΔN with actin as a loading control. Molecular weight markers in kDa. (d) Co-immunoprecipitation from lysates of SH-SY5Y cells expressing PrPΔOct or PrPΔN using antibodies against PrPC (6H4), GluA2, control rabbit anti-mouse IgG or no antibody for the bead-only control, and then probed with antibodies against PrPC (6D11 for PrPΔOct or SAF32 for PrPΔN) or GluA2 as indicated. (e) Co-immunoprecipitation from wild-type Ola/P2 mouse brain using antibodies against PrPC (6H4), GluA1, GluA2, control rabbit anti-mouse IgG or no antibody for the bead-only control with 20 μM TPEN included in all buffers and washes. Samples were probed with antibodies against PrPC (SAF32), GluA1 or GluA2 as indicated.