Skip to main content
. 2007 Sep 5;27(36):9801–9815. doi: 10.1523/JNEUROSCI.2661-07.2007

Figure 5.

Figure 5.

VAPB-P56S recruits both wild-type VAPA and VAPB. A, B, HeLa cells transfected with HA-VAPB-P56S (A) or HA-VAPA-P56S (B) and stained with anti-HA (green) and anti-VAPA (red; A) or anti-VAPB (red; B) antibodies. Endog., Endogenous. C, D, HeLa cells double transfected with HA-VAPB-P56S and myc-VAPB-wt (C) or myc-VAPA-wt (D), fixed and stained with anti-HA (green) and anti-Myc antibodies (red) to visualize the expressing VAP protein. E, Lysates (L) of COS-1 cells transfected with HA-VAPB-wt and HA-VAPB-P56S were solubilized with Triton X-100, fractionated in supernatant (S) and pellet (P), and analyzed by immunoblotting using anti-VAPB (#1006-03) antibodies. F, COS-1 cells cotransfected with HA-VAPB-P56S and myc-VAPA-wt or myc-VAPB-wt were immunoprecipitated with control IgG or HA antibodies. The cells were incubated for 2 h at 20°C before lysis to solubilize VAPB-P56S. Each immunoprecipitation reaction is shown in three lanes: I, input to IP reaction; S, supernatant remaining after IP; P, precipitated pellet. The L, I, S, and P samples were immunoblotted for the indicated proteins. G, COS-1 cells cotransfected with HA-VAPB-P56S and GFP-VAPB-wt-N or GFP-VAPB-TMD were immunoprecipitated and immunoblotted as in F. H–K, HeLa cells single or double transfected with GFP-VAPB-TMD (H, J) or GFP-VAPB-wt-N (I, K) and HA-VAPB-P56S (J, K) and stained with anti-calreticulin to reveal the ER (H, I), or anti-HA (J, K) antibodies to visualize the expressing VAP protein. L–N, HeLa cells single or double transfected with GFP-VAPB-P56S-N and HA-VAPB-wt (M) or HA-VAPB-P56S (N) and stained with anti-calreticulin (L) or anti-HA (M, N) antibodies. Scale bar, 10 μm.