Psy2 Targets the PP4 Family Phosphatase Pph3 To Dephosphorylate Mth1 and Repress Glucose Transporter Gene Expression

Abstract

The reversible nature of protein phosphorylation dictates that any protein kinase activity must be counteracted by protein phosphatase activity. How phosphatases target specific phosphoprotein substrates and reverse the action of kinases, however, is poorly understood in a biological context. We address this question by elucidating a novel function of the conserved PP4 family phosphatase Pph3-Psy2, the yeast counterpart of the mammalian PP4c-R3 complex, in the glucose-signaling pathway. Our studies show that Pph3-Psy2 specifically targets the glucose signal transducer protein Mth1 via direct binding of the EVH1 domain of the Psy2 regulatory subunit to the polyproline motif of Mth1. This activity is required for the timely dephosphorylation of the downstream transcriptional repressor Rgt1 upon glucose withdrawal, a critical event in the repression of HXT genes, which encode glucose transporters. Pph3-Psy2 dephosphorylates Mth1, an Rgt1 associated corepressor, but does not dephosphorylate Rgt1 at sites associated with inactivation, in vitro. We show that Pph3-Psy2 phosphatase antagonizes Mth1 phosphorylation by protein kinase A (PKA), the major protein kinase activated in response to glucose, in vitro and regulates Mth1 function via putative PKA phosphorylation sites in vivo. We conclude that the Pph3-Psy2 phosphatase modulates Mth1 activity to facilitate precise regulation of HXT gene expression by glucose.

Yeast PP4 Interacts with ATR Homolog Ddc2-Mec1 and Regulates Checkpoint Signaling

“…Several PP4 targets interact stably with the phosphatase (Keogh et al, 2006; Lee et al, 2010; Ma et al, 2014; O’Neill et al, 2007). Thus, we hypothesized that Pph3 might regulate Mec1, and not only counteract the enzyme by dephosphorylating its targets.…”

Cross-Talk between Carbon Metabolism and the DNA Damage Response in S. cerevisiae

“…Protein kinase A (PKA) affects glucose sensing at multiple levels [inhibition of Snf1 (Barrett et al, 2012), Rgt1 (Kim and Johnston, 2006, Roy et al, 2014) and Mth1 (Ma et al, 2014)], and is activated by Mec1 inhibition of the PKA regulatory subunit Bcy1 (Searle et al, 2011). We observed that phosphorylation of Mms21 in response to glucose (but not MMS) is reduced by overexpression of the PDE2 phosphodiesterase ( Figures 1b, c ) and that phosphorylation of Mms21 on a PKA consensus motif ( RxxS, Figure 1e ) is affected by changes in PKA activity.…”

Centromeric binding and activity of Protein Phosphatase 4

“…Furthermore, yeast R3, Psy2, specifically interacts with the glucose signal transducer protein Mth1 as a prerequisite for the PP4c–R3-mediated dephosphorylation of both Mth1 and a transcriptional repressor, Rgt1 (see ref. 18 ). These studies suggest that R3s are substrate-targeting subunits of PP4 but how substrate specificity is achieved remains unknown.…”