FIGURE 5:

PKA is required for Ent5 relocalization and hyperphosphorylation. (A) Prolonged inhibition of PKA is required to prevent Ent5 relocalization during glucose starvation. ENT5-GFP TPK1-as tpk2Δ tpk3Δ cells (DLY13) were incubated with 2 μM 1NM-PP1 for indicated times, and then imaged immediately after glucose removal. (B) PKA is required for starvation-induced changes in Ent5 localization. Cells were grown in glucose in the presence of DMSO (top) or 2 μM 1NM-PP1 (bottom) for 2 h. Cells were imaged just before starvation or at indicated times following glucose washout. (C) When PKA is inhibited, Ent5 is not hyperphosphorylated during glucose starvation. Cells were treated with DMSO or 2 μM 1NM-PP1 for 2 h during growth in glucose and then starved for glucose. Samples were taken from cells just before DMSO or 1NM-PP1 treatment (lane 1) or just prior to glucose starvation (lanes 2 and 6) or at indicated time-points after starvation. Cell lysates were subject to SDS–PAGE and Western blot analysis with antibodies to Ent5 or Adh1.