Figure 2.

Insulin stimulates the association of IR with Munc18c. (A) Munc18c was immunoprecipitated (IP) from cleared detergent lysates prepared from fully differentiated 3T3-L1 adipocytes that were incubated in serum-free medium for 2 h and then stimulated with 100 nM insulin for 5 min. Proteins were resolved by 10% SDS-PAGE and immunoblotted (IB) with anti-Munc18c and anti–insulin receptor (IR) antibodies. Data are from three independent sets of lysates quantified as the ratio of IR–Munc18c, with each set normalized to basal = 1.0; *, P < 0.05. (B) Munc18c was immunoprecipitated from hind-limb muscle extracts prepared from mice injected with vehicle (saline) or insulin (10 U/kg of body weight) for 5 min and processed for analysis as described in Fig. 1 B; *, P < 0.05 versus saline-injected controls. Equivalent IR abundance in the corresponding starting lysates was confirmed by immunoblotting (Lysate). n = 6 muscle extracts per data point. Data represent means ± SEM. Black lines indicate that intervening lanes have been spliced out.