Fig. 3.
Expression profile of the anti-apoptotic Bcl2 members upon withdrawal of trophic factors. (a) Constitutive expression of Nex1 induces expression of Bcl-XL and Bcl-XΔ™ proteins in serum-deprived PC12-Nex1 cells. PC12-Nex1 and PC12-bsd cells were grown in the absence of serum for periods of time indicated above each lane. Whole-cell extracts were prepared and equal amounts of total proteins (40 μg) were electrophoresed and subjected to immunoblot analysis with a polyclonal antibody raised against the 63-amino acid domain specific to the splice variant Bcl-XL to avoid cross-reaction with Bcl-XS. Equal loading was verified by staining the membrane with Ponceau S after transfer and by probing with the β-III tubulin antibody. Molecular weight markers are indicated on the left side of each figure. The asterisk indicates a non-specific anti-Bcl-XL immunoreactive polypeptide migrating more slowly than the Bcl-XL protein, and serves as an internal control for equal loading. (b) Constitutive expression of Nex1 results in increased expression of Bcl-w upon serum deprivation. Immunoblot analyses were performed as described above after stripping the membrane before probing with an anti-Bcl-w polyclonal antibody that does not cross-react with other Bcl2 proteins. Data shown are representative of at least three independent experiments.