FIGURE 6.

Glucose deprivation triggers AMPKβ-Thr-148 phosphorylation. A and B, HEK293T cells transiently overexpressing Myc-tagged α1, γ1, and WT or T148D mCherry-tagged β1 were treated for 16 h either with high glucose medium containing serum (HiG), with the same medium supplemented with forskolin (100 μm; HiG-Forsk), or with medium containing serum but no glucose (NoG). A, AMPKα was immunoprecipitated using the Myc tag antibody. Immunoblot using Myc tag antibody following precipitation shows the immunoprecipitation efficiency. Immunoblot with AMPKα-Thr(P)-172 and AMPKβ-Thr(P)-148 shows AMPK activity and autophosphorylation status. B, glycogen was quantified biochemically, corrected for protein concentration, and normalized to the level of WT cells under high glucose medium containing serum. Student's t test was used; treatment with high glucose medium containing serum versus treatment within one cell type (*, p < 0.05; #, p = 0.06) (n = 2). IP, immunoprecipitation; WB, Western blot.