Academic journal article Genetics

Structure/Function Analysis of the Phosphatidylinositol-3-Kinase Domain of Yeast Tra1

Academic journal article Genetics

Structure/Function Analysis of the Phosphatidylinositol-3-Kinase Domain of Yeast Tra1

Article excerpt

ABSTRACT

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-P^sub 3408^A, S^sub 3463^A, and SRR^sub 3413-3415^AAA 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 ~7% of yeast genes in rich media and reduced activation of PHO5 and ADH2 promoters. Tra1-SRR^sub 3413^ associated with components of both the NuA4 and SAGA complexes and with the Gal4 transcriptional activation domain similar to wild-type protein. Tra1-SRR^sub 3413^ 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-SRR^sub 3413^ 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.

IN eukaryotic cells the post-translational modification of nucleosomes by multisubunit complexes is a key aspect of transcriptional regulation (reviewed in Berger 2002).Histone modifications including acetylation,methylation, ubiquitylation, and phosphorylation can directly alter chromatin structure or act as a recruitment signal for additional factors (Strahl and Allis 2000). As well as regulating transcriptional initiation, nucleosome modifications affect transcriptional elongation and other nuclear processes such as DNA replication, DNA repair, and RNA export (Iizuka and Smith 2003).

The Saccharomyces cerevisiae Spt-Ada-Gcn5-Acetyltransferase (SAGA) complex modifies chromatin and provides an interface between DNA-binding transcriptional regulators and the basal transcriptional machinery (reviewed in Green 2005). The structural core of SAGA is composed of a subset of the TBP-associated factors (TAFs) (Grant et al. 1998a; Wu et al. 2004), with Spt7, Ada1, and Spt20 also being required for the integrity of the complex (Horiuchi et al. 1997; Roberts and Winston 1997; Sterner et al. 1999). The histone acetyltransferase Gcn5/Ada4 activates and represses transcription by modifying histones H3 and H2B (Brownell et al. 1996; Grant et al. 1997; Kuo et al. 1998;Wang et al. 1998; Ricci et al. 2002). In turn, the Ada proteins, Ada2 and Ngg1/Ada3 regulate the activity and substrate preference of Gcn5 (Balasubramanian et al. 2001). Further regulation is provided by the interaction of Spt3 and Spt8 with the TATA-binding protein (Eisenmann et al. 1992, 1994; Dudley et al. 1999). Recruitment of SAGA to promoters is mediated by Tra1, an essential 437-kDa protein (Grant et al. 1998b; Saleh et al. 1998) that interacts directly with transcriptional activators (Brown et al. 2001; Bhaumik et al. 2004; Fishburn et al. 2005; Reeves and Hahn 2005). The mammalian ortholog of Tra1, TRRAP is required for transcriptional regulation by myc, p53, E2F, and E1A (Mcmahon et al. 1998; Bouchard et al. 2001; Deleu et al. 2001; Ard et al. 2002; Kulesza et al. 2002). Its deletion results in defects in cell cycle progression and early embryonic lethality (Herceg et al. 2001).

Tra1 is also a component of the multisubunit NuA4 complex (Allard et al. 1999) that preferentially acetylates histones H4 and H2A, the catalytic subunit being the essential protein Esa1 (Smith et al. 1998; Clarke et al. 1999). NuA4 associates with acidic activation domains, probably through Tra1-mediated interactions, and activates transcription in an acetylation-dependent manner (Vignali et al. 2000; Nourani et al. …

Search by... Author
Show... All Results Primary Sources Peer-reviewed

Oops!

An unknown error has occurred. Please click the button below to reload the page. If the problem persists, please try again in a little while.