Structure/function analysis of the phosphatidylinositol-3-kinase domain of yeast Tra1

• Published in: Genetics (Austin) Vol. 177; no. 1; pp. 151 - 166
• Main Authors: Mutui, A.I, Hoke, S.M.T, Genereaux, J, Hannam, C, MacKenzie, K, Jobin-Robitaille, O, Guzzo, J, Cote, J, Andrews, B, Haniford, D.B
• Format: Journal Article
• Language: English
• Published: United States Genetics Society of America 01.09.2007; Copyright © 2007 by the Genetics Society of America
• Subjects: Alleles; Amino acids; beta-Galactosidase - metabolism; Blotting, Western; Chromatin Immunoprecipitation; DNA, Fungal - genetics; DNA, Fungal - metabolism; DNA-binding proteins; Enzymes; Gene Expression Profiling; gene expression regulation; Genetics; Histone Acetyltransferases; histones; Investigations; Kinases; mutants; Mutation - genetics; Phenotype; phosphatidylinositol 3-kinase; Phosphatidylinositol 3-Kinases - genetics; Phosphatidylinositol 3-Kinases - metabolism; promoter regions; Proteins; Saccharomyces cerevisiae; Saccharomyces cerevisiae - genetics; Saccharomyces cerevisiae - growth & development; Saccharomyces cerevisiae - metabolism; Saccharomyces cerevisiae Proteins - chemistry; Saccharomyces cerevisiae Proteins - genetics; Saccharomyces cerevisiae Proteins - metabolism; SAGA complex; Telomere - physiology; tra1 gene; transcription (genetics); transcriptional activation; yeasts
• ISSN: 0016-6731; 1943-2631; 1943-2631
• DOI: 10.1534/genetics.107.074476

Tra1 is an essential component of the Saccharomyces cerevisiae SAGA and NuA4 complexes. Using targeted mutagenesis, we identified residues within its C-terminal phosphatidylinositol-3-kinase (PI3K) domain that are required for function. The phenotypes of tra1-P3408A, S3463A, and SRR3413-3415AAA included temperature sensitivity and reduced growth in media containing 6% ethanol or calcofluor white or depleted of phosphate. These alleles resulted in a twofold or greater change in expression of approximately 7% of yeast genes in rich media and reduced activation of PHO5 and ADH2 promoters. Tra1-SRR3413 associated with components of both the NuA4 and SAGA complexes and with the Gal4 transcriptional activation domain similar to wild-type protein. Tra1-SRR3413 was recruited to the PHO5 promoter in vivo but gave rise to decreased relative amounts of acetylated histone H3 and histone H4 at SAGA and NuA4 regulated promoters. Distinct from other components of these complexes, tra1-SRR3413 resulted in generation-dependent telomere shortening and synthetic slow growth in combination with deletions of a number of genes with roles in membrane-related processes. While the tra1 alleles have some phenotypic similarities with deletions of SAGA and NuA4 components, their distinct nature may arise from the simultaneous alteration of SAGA and NuA4 functions.