Fig. 4.

Effects of protein kinase inhibitors on insulin-stimulated proteolytic processing of transgenic SREBP-1c in rat hepatocytes. Hepatocytes from nonfasted TgHA-hSREBP-1c rats were prepared and plated on day 0. On day 1, cells were left untreated or were pretreated for 3 h with the indicated protein kinase inhibitor: 0.2 μM wortmannin (lane 3), 0.1 μM rapamycin (lane 4), or 3 μM LYS6K2 (lane 5). After this preincubation, the cells were left untreated or were treated with 10 nM insulin for 30 min, harvested, and then pooled (three dishes of cells per sample). Each sample was subjected to immunoblot analyses of the following proteins: precursor (P) and nuclear (N) forms of transgenic HA-hSREBP-1c, phosphorylated Akt (P-Akt), total Akt, phosphorylated ribosomal protein S6 (P-S6), and total S6. Gels were exposed to film for 3–25 s.