Figure 5.

Coimmunoprecipitation of AP-3 subunits. Pig brain cytosol was immunoprecipitated under nondenaturing conditions with affinity-purified polyclonal antibodies against β3B, δ, σ3 (crossreacting with both the A and B isoforms), and γ. Gels were blotted, and the appropriate region was cut out and probed with each of the above antibodies, as well as with anti-μ3 (which does not recognize the native complex). The four subunits of the AP-3 complex, δ, β3, μ3, and σ3, all coimmunoprecipitate; the γ subunit of the AP-1 complex does not coimmunoprecipitate with antibodies against the AP-3 components, and antibodies against γ do not bring down any of these components. The doublet labeled with anti-σ3 presumably corresponds to the A and B isoforms, one of which appears to be preferentially immunoprecipitated with this antibody.