Reg1 Protein Regulates Phosphorylation of All Three Snf1 Isoforms but Preferentially Associates with the Gal83 Isoform

Abstract

The phosphorylation status of the Snf1 activation loop threonine is determined by changes in the rate of its dephosphorylation, catalyzed by the yeast PP1 phosphatase Glc7 in complex with the Reg1 protein. Previous studies have shown that Reg1 can associate with both Snf1 and Glc7, suggesting substrate binding as a mechanism for Reg1-mediated targeting of Glc7. In this study, the association of Reg1 with the three Snf1 isoforms was measured by two-hybrid analysis and coimmunoprecipitation. We found that Reg1 association with Snf1 occurred almost exclusively with the Gal83 isoform of the Snf1 complex. Nonetheless, Reg1 plays an important role in determining the phosphorylation status of all three Snf1 isoforms. We found that the rate of dephosphorylation for isoforms of Snf1 did not correlate with the amount of associated Reg1 protein. Functional chimeric ␤ subunits containing residues from Gal83 and Sip2 were used to map the residues needed to promote Reg1 association with the N-terminal 150 residues of Gal83. The Gal83 isoform of Snf1 is the only isoform capable of nuclear localization. A Gal83-Sip2 chimera containing the first 150 residues of Gal83 was able to associate with the Reg1 protein but did not localize to the nucleus. Therefore, nuclear localization is not required for Reg1 association. Taken together, these data indicate that the ability of Reg1 to promote the dephosphorylation of Snf1 is not directly related to the strength of its association with the Snf1 complex.

The mammalian AMP‐activated protein kinase complex mediates glucose regulation of gene expression in the yeast Saccharomyces cerevisiae

“…In yeast the essential Glc7 PP1 is targeted to Snf1 by its subunits Reg1 as well as Reg2. Reg2 appears to play a minor role and generally deletion of REG2 only confers an effect in combination with deletion of REG1 [30,31,60] (and our unpublished data). The reg1 Δ reg2 Δ double mutant does not show any glucose‐induced dephosphorylation of Snf1.…”

2-Deoxyglucose Impairs Saccharomyces cerevisiae Growth by Stimulating Snf1-Regulated and α-Arrestin-Mediated Trafficking of Hexose Transporters 1 and 3

“…3A and B). Reg1 is the regulatory subunit of Glc7, the type 1 protein phosphatase (PP1) responsible for the dephosphorylation and inactivation of Snf1 (55)(56)(57). In reg1⌬ cells, Snf1 is constitutively phosphorylated and active (58,59), and therefore, one might have expected reg1⌬ cells to display the opposite phenotype from snf1⌬ cells.…”

Specific α-Arrestins Negatively Regulate Saccharomyces cerevisiae Pheromone Response by Down-Modulating the G-Protein-Coupled Receptor Ste2

“…Similarly, AMPK phosphorylates and promotes degradation of the mammalian ␣-arrestin family member TXNIP, thereby increasing glucose uptake by preventing TXNIP-mediated downregulation of the glucose transporter GLUT1 (137). Moreover, in glucose control of Rod1 action on Jen1, Reg1-bound Glc7 seems to be responsible for Rod1 dephosphorylation (64) and likely also prevents Snf1-mediated phosphorylation of Rod1 by deactivating Snf1 itself (138). However, we observed a requirement for CN-dependent dephosphorylation for Rod1 action on Ste2 on glucose-rich medium, a condition under which Snf1 is not activated (112).…”