Munc18 is required for APP–syntaxin 1 copatching and regulates β cleavage in N2a cells. (a) The effect of RNAi-mediated knockdown of endogenous Munc18 on APP–syntaxin 1 copatching. RNAi reduced APP–syntaxin 1 copatching, indicating a requirement of Munc18 for APP–syntaxin 1 copatching. Results are means + SD of 32–38 measurements. (b) The effect of Munc18 knockdown on β cleavage. N2a cells were transfected with pHVenus-APP and RNAi vector. Released sAPP in medium was captured with anti-GFP and quantified with anti-sAPP end-specific antibodies. Munc18 knockdown promoted β cleavage, which is consistent with an inhibitory role of X11–Munc18–syntaxin 1 on β cleavage. Data are means + SD based on four independent experiments (n = 8). (c) The effect of Munc18 knockdown on the interaction between APP and syntaxin 1. N2a cells were lyzed with Lubrol and used for IP with anti-APPex. In the lysate of N2a cells transfected with RNAi vector, syntaxin 1 association with APP was reduced. ***, P < 0.001.