Figure 3

AS160 knockdown suppresses glucose consumption and inhibits cell growth by reducing PM GLUT4 content in KMS11, JJN3, and L363 but not in U266 cells. (A) Immunoblot analysis verifying AS160 knockdown with two distinct AS160 targeting shRNA’s. Image representative of three individual repeats is presented; (B, D and F) Glucose uptake rates in KMS11, JJN3, and L363 cells, respectively, containing non-target control, AS-1 and AS-2 shRNAs. Rates were determined at 0 and 5 h after incubation in 5 mM glucose; (C, E and G) Growth rates in KMS11, JJN3, and L363 cell lines containing non-target control, AS-1 and AS-2 shRNAs, show trypan blue excluded viable cell counts at 0, 3, 4, and 5 days; (H) Immunoblot analysis of plasma membrane proteins from JJN3 cells infected with non-target control and AS-1 shRNAs for GLUT4 and Na/K and α-tubulin loading controls; (I) Immunoblot analysis for AS160, GLUT4, and GAPDH loading control from whole cell lysates in JJN3 cells treated with non-target control and AS-1 shRNAs. Immunoblots are representative of two individual repeats; (J) Glucose uptake rates in U266 containing non-target control and AS-1 shRNA; (K) Growth rates in U266 containing non-target control and AS-1 shRNAs, show trypan blue excluded viable cell counts at 0, 3, and 4 days. Bars and points represent mean ± SD of three repeats; *P <0.05, **P <0.01, ***P <0.001.