FIGURE 6.

Co-localization of printor and torsinA in the ER. A, SH-SY5Y cells expressing Myc-tagged printor and C-terminal HA-tagged torsinA (top) were immunostained with primary antibodies against Myc, HA, and KDEL, followed by detection with secondary antibodies conjugated to Texas Red (torsinA, red), FITC (printor, green), or Cy5 (KDEL, blue). Untransfected SH-SY5Y cells (bottom) were immunostained with primary antibodies against printor, torsinA, and KDEL, followed by detection with secondary antibodies conjugated to Texas Red (torsinA, red), FITC (printor, green), or Cy5 (KDEL, blue). Scale bars, 10 μm. B, post-nuclear supernatant from SH-SY5Y cells was fractionated on a 10–30% Opti-Prep gradient into 18 fractions, with fraction 1 corresponding to the top of the gradient. Equal volumes of each fraction were analyzed by SDS-PAGE followed by immunoblotting using anti-printor, anti-torsinA, and anti-calnexin antibodies. C, the level of the indicated protein in each fraction was quantified using NIH Scion Image and shown as a percentage of the total level of the indicated protein. Data are representative of at least three independent experiments.