Figure 4.

Coexpression of CgB and VGF in adrenal chromaffin and PC12 pheochromocytoma cells. A, B) Coimmunoprecipitation analysis was used to determine whether CgB and VGF associate in PC12 pheochromocytoma cells. PC12 cell lysates were immunoprecipitated with rabbit anti-VGF or normal rabbit IgG (A), and with goat anti-CgB or normal goat IgG (B), and complexes were collected and analyzed by SDS-PAGE and Western blotting with anti-CgB (A) and anti-VGF (B; asterisks indicate nonspecific bands). PC12 cell lysate (25 μg) was coanalyzed as a positive control and represented 5% of the input lysate immunoprecipitated. C) PC12 cell lysates were immunoprecipitated with anti-SgII, anti-CgA, or anti-SgIII, and analyzed by SDS-PAGE and Western blotting with anti-VGF; no coimmunoprecipitated VGF was identified (asterisks indicate nonspecific bands). D) Western blot analysis revealed VGF protein (arrows ∼85 and ∼90 kDa) in adrenal gland lysates from male and female wild-type mice and rat PC12 cells. E–J) VGF and CgB distributions in PC12 cells were analyzed by indirect immunofluorescence and confocal microscopy. Coexpression of VGF (red; E, H) and CgB (green; F, I) is shown in merge panels (yellow; G, J). K–Q) VGF (red; K, O) and CgB (green; L, P) were similarly coexpressed (yellow; M, N, Q) in sections of mouse adrenal gland (K–N) and, shown at lower magnification, rat adrenal gland (O–Q).