Figure 4.
The PKA-mediated inhibition of P-body formation was suppressed by the presence of a non-phosphorylatable form of Pat1.
(A) The PKA-dependent phosphorylation of Pat1 was rapidly lost upon glucose starvation. Cells expressing a myc-tagged Pat1 were grown to mid-log phase in an SC-glucose medium and then transferred to a medium lacking glucose (SC-D) for the indicated time. The level of PKA phosphorylation was assessed by Western blotting with the anti-PKA substrate antibody.
(B, C) The presence of the non-phosphorylatable variant, Pat1-AA, resulted in an elevated number of P-body foci in bcy1Δ cells. Wild-type and bcy1Δ cells expressing the indicated GFP-tagged Pat1 proteins were grown to mid-log phase and then transferred to a medium lacking glucose for 15 min. Representative fluorescence microscopy images are shown in B and the quantification of these data in C.
(D) The presence of the non-phosphorylatable Pat1 also suppressed the inhibitory effects of elevated PKA signaling on P-body formation during an extended glucose starvation (-Glu, 90 min) or growth into early stationary phase (1d).