FIGURE 3.

As₂O₃ does not increase cardiac PI3K activity. Freshly isolated guinea pig ventricular myocytes were incubated for 4 h under control conditions or in the presence of 10 μm As₂O₃. Catalytic PI3K subunits p110α, p110β, and p110γ were immunoprecipitated from whole cell lysates. PI3K activity of immunocomplexes was assayed as a function of PI-3P production. A, shown is a typical autoradiogram of ³²P-labeled PI-3P produced on immunoprecipitation (IP) of p110γ or p100α and resolved by TLC. Ori, origin of TLC migration. Incubation with As₂O₃ does not alter PI-3P production, whereas co-incubation with 200 nm wortmannin inhibits PI-3P production. PI-3P production is not observed upon the omission of substrate from the assay reaction (no PI). Note that the basal activity of PI3K-γ/p110γ is very low under control conditions. B, quantitative analysis of PI3K/p110 activities. Shown is normalized PI-3P production under control conditions or in the presence of 10 μm As₂O₃ for PI-3K/p110α, -β, and -γ. PI-3K/p110α activity is significantly inhibited by 200 nm wortmannin. Assays without the addition of PI were used as negative controls (n = 1–5). *, significant difference from PI-3K/p100α activity measured under control conditions (Dunnett's test, p < 0.05). Error bars, S.E.