Glucose-induced posttranslational activation of protein phosphatases PP2A and PP1 in yeast

Abstract

The protein phosphatases PP2A and PP1 are major regulators of a variety of cellular processes in yeast and other eukaryotes. Here, we reveal that both enzymes are direct targets of glucose sensing. Addition of glucose to glucose-deprived yeast cells triggered rapid posttranslational activation of both PP2A and PP1. Glucose activation of PP2A is controlled by regulatory subunits Rts1, Cdc55, Rrd1 and Rrd2. It is associated with rapid carboxymethylation of the catalytic subunits, which is necessary but not sufficient for activation. Glucose activation of PP1 was fully dependent on regulatory subunits Reg1 and Shp1. Absence of Gac1, Glc8, Reg2 or Red1 partially reduced activation while Pig1 and Pig2 inhibited activation. Full activation of PP2A and PP1 was also dependent on subunits classically considered to belong to the other phosphatase. PP2A activation was dependent on PP1 subunits Reg1 and Shp1 while PP1 activation was dependent on PP2A subunit Rts1. Rts1 interacted with both Pph21 and Glc7 under different conditions and these interactions were Reg1 dependent. Reg1-Glc7 interaction is responsible for PP1 involvement in the main glucose repression pathway and we show that deletion of Shp1 also causes strong derepression of the invertase gene SUC2. Deletion of the PP2A subunits Pph21 and Pph22, Rrd1 and Rrd2, specifically enhanced the derepression level of SUC2, indicating that PP2A counteracts SUC2 derepression. Interestingly, the effect of the regulatory subunit Rts1 was consistent with its role as a subunit of both PP2A and PP1, affecting derepression and repression of SUC2, respectively. We also show that abolished phosphatase activation, except by reg1Δ, does not completely block Snf1 dephosphorylation after addition of glucose. Finally, we show that glucose activation of the cAMP-PKA (protein kinase A) pathway is required for glucose activation of both PP2A and PP1. Our results provide novel insight into the complex regulatory role of these two major protein phosphatases in glucose regulation.

Modulation of TORC2 Signaling by a Conserved Lkb1 Signaling Axis in Budding Yeast

“…Since ceramide production is controlled by TORC2-dependent signals, this observation strengthens previous genetic data that suggested a role for Rts1 in a ceramide-dependent feedback loop that negatively regulates TORC2 signaling (Lucena et al 2018). Rts1 hyperphosphorylation in poor carbon is correlated with a decrease in Rts1-associated phosphatase activity, consistent with a previous study that suggested that a fraction of Rts1 dissociates from PP2A in poor carbon (Castermans et al 2012). A previous study suggested that the Rts1 that dissociates from PP2A in poor carbon could have PP2A-independent functions.…”

“…Phosphatase activity associated with Rts1 was reduced in cells growing in poor carbon, when Rts1 was hyperphosphorylated ( Figure 3E). A previous study found that PP2A dissociates from Rts1 when cells are grown in poor carbon, which could explain the reduction in Rts1-associated phosphatase activity in those conditions (Castermans et al 2012). Further evidence for dissociation of the PP2A Rts1 complex in poor carbon came from analysis of Rts1 phosphorylation in response to inactivation of PP2A catalytic subunits.…”

“…A potential explanation is that Rts1 dissociates from PP2A in poor carbon and then carries out PP2A-independent functions. A previous study suggested that Rts1 associates with Glc7, the budding yeast homolog of the PP1 phosphatase, and that association of Rts1 with Glc7 increases in poor carbon (Castermans et al 2012). Thus, the PP2A Rts1 complex could dominate in rich carbon, while a Glc7 Rts1 complex could play a role in the response to poor carbon, including a role in reducing cell size.…”

“…Feedback signals are partially dependent upon Rts1, a conserved regulatory subunit of PP2A (Lucena et al 2018). There is evidence that Rts1 may also be a regulatory subunit for Glc7, which is the budding yeast homolog of the PP1 phosphatase (Castermans et al 2012). …”

The AMP-activated Protein Kinase Snf1 Regulates Transcription Factor Binding, RNA Polymerase II Activity, and mRNA Stability of Glucose-repressed Genes in Saccharomyces cerevisiae

“…The addition of glucose to a yeast culture growing in its absence activates the GLC7-and SIT4-encoded protein phosphatases (54). The effect of glucose on their activation is rapid, occurring in less than 1 min and acts through a G-proteincoupled receptor.…”