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Expression of human protein phosphatase-1 in Saccharomyces cerevisiae highlights the role of phosphatase isoforms in regulating eukaryotic functions.

Publication ,  Journal Article
Gibbons, JA; Kozubowski, L; Tatchell, K; Shenolikar, S
Published in: J Biol Chem
July 27, 2007

Human (PP1) isoforms, PP1alpha, PP1beta, PP1gamma1, and PP1gamma2, differ in primary sequences at N and C termini that potentially bind cellular regulators and define their physiological functions. The GLC7 gene encodes the PP1 catalytic subunit with >80% sequence identity to human PP1 and is essential for viability of Saccharomyces cerevisiae. In yeast, Glc7p regulates glycogen and protein synthesis, actin cytoskeleton, gene expression, and cell division. We substituted human PP1 for Glc7p in yeast to investigate the ability of individual isoforms to catalyze Glc7p functions. S. cerevisiae expressing human PP1 isoforms were viable. PP1alpha-expressing yeast grew more rapidly than strains expressing other isoforms. On the other hand, PP1alpha-expressing yeast accumulated less glycogen than PP1beta-or PP1gamma1-expressing yeast. Yeast expressing human PP1 were indistinguishable from WT yeast in glucose derepression. However, unlike WT yeast, strains expressing human PP1 failed to sporulate. Analysis of chimeric PP1alpha/beta subunits highlighted a critical role for their unique N termini in defining PP1alpha and PP1beta functions in yeast. Biochemical studies established that the differing association of PP1 isoforms with the yeast glycogen-targeting subunit, Gac1p, accounted for their differences in glycogen synthesis. In contrast to human PP1 expressed in Escherichia coli, enzymes expressed in yeast displayed in vitro biochemical properties closely resembling PP1 from mammalian tissues. Thus, PP1 expression in yeast should facilitate future structure-function studies of this protein serine/threonine phosphatase.

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Published In

J Biol Chem

DOI

ISSN

0021-9258

Publication Date

July 27, 2007

Volume

282

Issue

30

Start / End Page

21838 / 21847

Location

United States

Related Subject Headings

  • Substrate Specificity
  • Saccharomyces cerevisiae
  • Protein Subunits
  • Protein Phosphatase 1
  • Polymerase Chain Reaction
  • Plasmids
  • Phosphoprotein Phosphatases
  • Molecular Sequence Data
  • Microscopy, Fluorescence
  • Isoenzymes
 

Citation

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Gibbons, J. A., Kozubowski, L., Tatchell, K., & Shenolikar, S. (2007). Expression of human protein phosphatase-1 in Saccharomyces cerevisiae highlights the role of phosphatase isoforms in regulating eukaryotic functions. J Biol Chem, 282(30), 21838–21847. https://doi.org/10.1074/jbc.M701272200
Gibbons, Jennifer A., Lukasz Kozubowski, Kelly Tatchell, and Shirish Shenolikar. “Expression of human protein phosphatase-1 in Saccharomyces cerevisiae highlights the role of phosphatase isoforms in regulating eukaryotic functions.J Biol Chem 282, no. 30 (July 27, 2007): 21838–47. https://doi.org/10.1074/jbc.M701272200.
Gibbons, Jennifer A., et al. “Expression of human protein phosphatase-1 in Saccharomyces cerevisiae highlights the role of phosphatase isoforms in regulating eukaryotic functions.J Biol Chem, vol. 282, no. 30, July 2007, pp. 21838–47. Pubmed, doi:10.1074/jbc.M701272200.
Gibbons JA, Kozubowski L, Tatchell K, Shenolikar S. Expression of human protein phosphatase-1 in Saccharomyces cerevisiae highlights the role of phosphatase isoforms in regulating eukaryotic functions. J Biol Chem. 2007 Jul 27;282(30):21838–21847.

Published In

J Biol Chem

DOI

ISSN

0021-9258

Publication Date

July 27, 2007

Volume

282

Issue

30

Start / End Page

21838 / 21847

Location

United States

Related Subject Headings

  • Substrate Specificity
  • Saccharomyces cerevisiae
  • Protein Subunits
  • Protein Phosphatase 1
  • Polymerase Chain Reaction
  • Plasmids
  • Phosphoprotein Phosphatases
  • Molecular Sequence Data
  • Microscopy, Fluorescence
  • Isoenzymes