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Vol. 19, Issue 8, 3323-3333, August 2008

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The Yeast GID Complex, a Novel Ubiquitin Ligase (E3) Involved in the Regulation of Carbohydrate Metabolism

Olivier Santt^*,, Thorsten Pfirrmann^*,,, Bernhard Braun^*, Jeannette Juretschke^*, Philipp Kimmig^*, Hartmut Scheel, Kay Hofmann, Michael Thumm^*,||, and Dieter H. Wolf^*

*Institut für Biochemie, Universität Stuttgart, 70569 Stuttgart, Germany; and Miltenyi Biotec GmbH, 50829 Köln, Germany

Submitted March 28, 2008; Revised May 13, 2008; Accepted May 19, 2008
Monitoring Editor: Thomas Sommer

Glucose-dependent regulation of carbon metabolism is a subject of intensive studies. We have previously shown that the switch from gluconeogenesis to glycolysis is associated with ubiquitin-proteasome linked elimination of the key enzyme fructose-1,6-bisphosphatase. Seven glucose induced degradation deficient (Gid)-proteins found previously in a genomic screen were shown to form a complex that binds FBPase. One of the subunits, Gid2/Rmd5, contains a degenerated RING finger domain. In an in vitro assay, heterologous expression of GST-Gid2 leads to polyubiquitination of proteins. In addition, we show that a mutation in the degenerated RING domain of Gid2/Rmd5 abolishes fructose-1,6-bisphosphatase polyubiquitination and elimination in vivo. Six Gid proteins are present in gluconeogenic cells. A seventh protein, Gid4/Vid24, occurs upon glucose addition to gluconeogenic cells and is afterwards eliminated. Forcing abnormal expression of Gid4/Vid24 in gluconeogenic cells leads to fructose-1,6-bisphosphatase degradation. This suggests that Gid4/Vid24 initiates fructose-1,6-bisphosphatase polyubiquitination by the Gid complex and its subsequent elimination by the proteasome. We also show that an additional gluconeogenic enzyme, phosphoenolpyruvate carboxykinase, is subject to Gid complex-dependent degradation. Our study uncovers a new type of ubiquitin ligase complex composed of novel subunits involved in carbohydrate metabolism and identifies Gid4/Vid24 as a major regulator of this E3.

These authors contributed equally to this work.

Present addresses: University of Stockholm, Wenner-Gren Institute, Svante Arrheniusväg 16-18, 10691 Stockholm, Sweden;

^|| Institut für Biochemie und Molekulare Zellbiologie, Universität Göttingen, Heinrich-Dueker-Weg 12, 37073 Göttingen, Germany.

Address correspondence to: Dieter H. Wolf (dieter.wolf{at}ibc.uni-stuttgart.de)

Abbreviations used: CTLH, C-terminal to LisH motif; FBPase, fructose-1,6-bisphosphatase; GST, Glutathione transferase; LisH, Lissencephaly type-1-like homology motif; ORF, Open reading frame; PEPCK, phospho-enol-pyruvate carboxykinase.