Fig 8.

Us11 does not block autophagy by inhibiting Beclin 1 or mTOR. (A) HeLa cells were transiently transfected to express Beclin 1 and empty vector pSG5-HA, HA-Us11, Flag-ICP34.5, or Bcl-2. The Us11-Beclin 1 interaction was tested in complete medium or after stimulating autophagy by poly(I · C) (top panel). Beclin 1 was immunoprecipitated as described in Materials and Methods. Coimmunoprecipitations of Beclin 1 and Bcl-2 or of Beclin 1 and Flag-ICP34.5 in HeLa cells were performed as positive controls (bottom panels). Each immunoblot was representative of three independent experiments. IP, immunoprecipitation; IB, immunoblot. (B) PKR^+/+ and PKR^−/− MEFs were transfected with empty vector or Us11 for 24 h and grown in starvation medium (EBSS) for 4 h. Cells were analyzed for mTOR activity by immunoblotting for levels of phosphorylated 4E-BP1 (4E-BP1-P), total 4E-BP1, phosphorylated p70S6K (p70S6K-P), and total p70S6K. NT, not transfected.