title: 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 β 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.
authors: Department of Microbiology and Molecular Genetics, University of Pittsburgh School of Medicine, Pittsburgh, PA 15219, USA.
authors: Yuxun
authors: Y
authors: Zhang
authors: Rhonda R
authors: RR
authors: McCartney
authors: Dakshayini G
authors: DG
authors: Chandrashekarappa
authors: Simmanjeet
authors: S
authors: Mangat
authors: Martin C
authors: MC
authors: Schmidt
mesh_terms: D049452:Green Fluorescent Proteins / Q000378:metabolism; D007527:Isoenzymes / Q000378:metabolism; D019913:Nuclear Localization Signals; D010766:Phosphorylation; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs; D054645:Protein Phosphatase 1 / Q000378:metabolism / Q000502:physiology*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D021381:Protein Transport; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012097:Repressor Proteins / Q000737:chemistry / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000254:growth & development; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000378:metabolism* / Q000502:physiology*; D015534:Trans-Activators / Q000378:metabolism; D020798:Two-Hybrid System Techniques
chemical_list: C085569:GAL83 protein, S cerevisiae; D007527:Isoenzymes; D019913:Nuclear Localization Signals; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins; C091267:SIP2 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D015534:Trans-Activators; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; C071570:SNF1-related protein kinases; D017346:Protein Serine-Threonine Kinases; C092048:GLC7 protein, S cerevisiae; D054645:Protein Phosphatase 1; C070829:REG1 protein, S cerevisiae