title: Sumoylation of the yeast Gcn5 protein.
abstract: Sumoylation, the process by which the ubiquitin-related SUMO protein is covalently attached to lysine side chains in other proteins, is involved in numerous processes in the eukaryotic cell, including transcriptional repression. In this study, we identify Gcn5, the histone-modifying subunit of the transcriptional regulatory complex SAGA, as a sumoylation substrate in yeast. In vitro, multiple sumoylation of recombinant Gcn5 alone or as a trimer with its interacting proteins Ada2 and Ada3 did not affect Gcn5's histone acetyltransferase (HAT) activity, suggesting that modification of Gcn5 with yeast SUMO (Smt3) may not directly regulate its HAT function. Through site-directed mutagenesis, the primary in vivo sumoylation site was identified as lysine-25, although an unsumoylatable K-to-R mutation of this residue led to no obvious in vivo effects. However, fusion of SUMO to the N-terminus of Gcn5 to mimic constitutive sumoylation resulted in defective growth on 3-aminotriazole media and reduced basal and activated transcription of the SAGA-dependent gene TRP3. Taken together with recent identification of multiple additional subunits of SAGA as sumoylated proteins in vivo, these data suggest that Gcn5 sumoylation may have an inhibitory role in transcriptional regulation.
authors.affiliation: Gene Expression and Regulation Program, The Wistar Institute, Philadelphia, Pennsylvania 19104, USA.
authors.forename: David E
authors.initials: DE
authors.lastname: Sterner
authors.forename: Dafna
authors.initials: D
authors.lastname: Nathan
authors.forename: Alison
authors.initials: A
authors.lastname: Reindle
authors.forename: Erica S
authors.initials: ES
authors.lastname: Johnson
authors.forename: Shelley L
authors.initials: SL
authors.lastname: Berger
mesh_terms: D001665:Binding Sites; D004926:Escherichia coli / Q000235:genetics; D051548:Histone Acetyltransferases / Q000235:genetics / Q000378:metabolism*; D008239:Lysine / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D025842:SUMO-1 Protein / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D000099224:p300-CBP-Associated Factor
chemical_list: D051548:Histone Acetyltransferases; D008239:Lysine; D011994:Recombinant Proteins; D025842:SUMO-1 Protein; D029701:Saccharomyces cerevisiae Proteins; C501392:GCN5 protein, S cerevisiae; D000099224:p300-CBP-Associated Factor