Accumulation of a Threonine Biosynthetic Intermediate Attenuates General Amino Acid Control by Accelerating Degradation of Gcn4 via Pho85 and Cdk8
Figure 8
Model for the roles of Pho85 and Srb10 in accelerated turnover of Gcn4 in response to ASA accumulation in hom6Δ cells.
(A) Starvation of WT (HOM6) cells for Ile/Val evokes increased synthesis of Gcn4 at the translational level and subsequent increased binding of Gcn4 to UAS elements of genes subject to GAAC. Gcn4 recruits coactivators, including Mediator (green and red shapes connecting Gcn4 to Pol II), and Pol II to the promoter (TATA) for increased transcription of target gene coding sequences (CDS). As Srb10 is associated with Mediator and recruited by UAS-bound Gcn4, we propose that Srb10 phosphorylates the bulk of UAS-bound Gcn4 molecules (orange balls labeled with “P”), stimulating their ubiquitylation and attendant degradation by the proteasome. UAS-bound Gcn4 is also sumoylated by Ubc9 (white balls labeled with “S”) and the sumoylated molecules are targeted for degradation by Srb10. Pho85 is responsible for the majority of Gcn4 turnover, and we propose it phosphorylates (green balls labeled with “P”) and triggers degradation of both active and defective non-UAS bound Gcn4 species. (B) Starvation of hom6Δ cells for Ile/Val also evokes increased synthesis of Gcn4 and attendant increased binding of Gcn4 to UAS elements. However, we hypothesize that ASA accumulation evokes damage or inactivating modifications of Gcn4. Damage/modification restricted to the activation domain, disengages UAS-bound Gcn4 from coactivators and increases its rate of phosphorylation by Srb10 at the promoter, with attendant increased degradation by the proteasome. (Gcn4 opacity is reduced to depict its decreased occupancy of the UAS.) Damage/modification that extends to the DNA binding or dimerization domain disengages Gcn4 from the UAS and makes it susceptible to phosphorylation by Pho85 and subsequent proteasomal degradation. Pho85 also targets functional Gcn4 molecules when they disengage from the UAS, just as in HOM6 cells. Hence, the absence of Srb10 in srb10Δ cells spares from degradation defective Gcn4 molecules capable of UAS-binding and thereby reduces the specific activity of UAS-bound Gcn4. The absence of Pho85 in pho85Δ cells spares both fully functional Gcn4 molecules and inactive species incapable of stable UAS-binding and thereby increases the specific activity of UAS-bound Gcn4 while simultaneously decreasing the fraction of Gcn4 capable of UAS binding.