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. 2012 Jan 31;22(6):1058–1077. doi: 10.1038/cr.2012.20

Figure 2.

Figure 2

Regulatory subunit Rts1, activators Rrd1 or Rrd2 and methylation at the C-terminus of Pph21 are required for glucose activation of PP2A, whereas regulatory subunit Cdc55 inhibits activation. At time 0, 20 mM glucose was added to glucose-deprived (glycerol-grown) cells of BY-strains with different deletions and cell extracts were used to measure specific PP2A activity (A, B, C and D), or to detect C-terminal methylation of catalytic subunit Pph21 (E and F). (A) WT strain (•), tpd3Δ (○), cdc55Δ (▴), rts1Δ (△) and rts1Δ cdc55Δ (▪). (B) WT strain (•), rrd1Δ (○), rrd2Δ (▴) and rrd1Δ rrd2Δ (△). (C) WT strain (•), ppm1Δ (○) and ppe1Δ (▴). (D) pph21Δ pph22Δ + pHA-Pph21 (•), pph21Δ pph22Δ + pHA-Pph21L369A (○) and pph21Δ pph22Δ + pEMPTY (▴). (E, F) Using antibodies specifically recognizing methylated or unmethylated PP2A catalytic subunits, we monitored changes in C-terminal methylation of pHA-Pph21 and pHA-Pph21L369A in a pph21Δ pph22Δ strain (E), and of pHA-Pph21 in strains ppe1Δ pph21Δ pph22Δ and ppm1Δ ppm2Δ pph21Δ pph22Δ (F). As a control for equal loading, the membranes were incubated with anti-HA after stripping.