Identification and characterization of HAP4: a third component of the CCAAT-bound HAP2/HAP3 heteromer

Department of Biology, Massachusetts Institute of Technology, Cambridge 02139.
Genes & Development (Impact Factor: 10.8). 09/1989; 3(8):1166-78. DOI: 10.1101/gad.3.8.1166
Source: PubMed


The CYC1 gene of Saccharomyces cerevisiae is positively regulated by the HAP2 and HAP3 proteins, which form a heteromeric complex that binds to a CCAAT box in the upstream activation site, UAS2, and which activate transcription in a nonfermentable carbon source. We carried out a genetic analysis to identify additional trans-acting regulatory factors exerting their effects through UAS2. We present the identification and characterization of a new locus, HAP4, which is shown to encode a subunit of the DNA-binding complex at UAS2. In the hap4 mutant, the binding of HAP2 and HAP3 (HAP2/3) is not observed in vitro. The HAP4 gene is regulated transcriptionally by a carbon source, suggesting that it encodes a regulatory subunit of the bound complex. The sequence of HAP4 shows a highly acidic region, which innactivated the protein when deleted. Replacement of this region with the activation domain of GAL4 restored activity, suggesting that it provides the principal activation domain to the bound HAP2/3/4 complex.

Download full-text


Available from: Susan Forsburg
  • Source
    • "They are transcribed but then rapidly degraded (Scheffler et al., 1998). While the HAP2/3/4/5 DNA-binding complex is involved in the transcriptional activation of genes for respiration (Forsburg and Guarente, 1989), the RNA-binding protein Puf3p, which is not a canonical transcriptional factor, has also been implicated in the regulation of such genes (Gerber et al., 2004). Puf3p is a member of the PUF (PUmilio and FBF) family of RNA-binding proteins (Murata and Wharton, 1995; Zamore et al., 1997; Zhang et al., 1997) and has been shown to specifically bind to the 3 0 UTRs of mRNAs encoding mitochondrial proteins (Jackson et al., 2004; Olivas and Parker, 2000; Ulbricht and Olivas, 2008; Zhu et al., 2009). "
    [Show abstract] [Hide abstract]
    ABSTRACT: PUF proteins are post-transcriptional regulators that bind to the 3' UTRs of mRNA transcripts. Herein, we show how a yeast PUF protein, Puf3p, responds to glucose availability to switch the fate of its bound transcripts that encode proteins required for mitochondrial biogenesis. Upon glucose depletion, Puf3p becomes heavily phosphorylated within its N-terminal region of low complexity, associates with polysomes, and promotes translation of its target mRNAs. Such nutrient-responsive phosphorylation toggles the activity of Puf3p to promote either degradation or translation of these mRNAs according to the needs of the cell. Moreover, activation of translation of pre-existing mRNAs might enable rapid adjustment to environmental changes without the need for de novo transcription. Strikingly, a Puf3p phosphomutant no longer promotes translation but becomes trapped in intracellular foci in an mRNA-dependent manner. Our findings suggest that the inability to properly resolve Puf3p-containing RNA-protein granules via a phosphorylation-based mechanism might be toxic to a cell. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.
    Full-text · Article · Jun 2015 · Cell Reports
  • Source
    • "In the present study MGA2 in both strains increased by more than 3-fold in xylose-containing medium compared with glucose-containing medium (data not shown), indicating that xylose induces adaptation to oxidative stress. It should be also noted that in MA-B4, transcript levels of HAP4, encoding a transcriptional activator and global regulator of respiratory gene expression [52], were more than 6.7-fold higher with xylose than with glucose, but did not change significantly with respect to carbon sources in MA-R4 (data not shown). It is also worth mentioning that stress responses induce the expression of genes that are implicated in respiration, including CIT1 and CYC7. "
    [Show abstract] [Hide abstract]
    ABSTRACT: There has been much research on the bioconversion of xylose found in lignocellulosic biomass to ethanol by genetically engineered Saccharomyces cerevisiae. However, the rate of ethanol production from xylose in these xylose-utilizing yeast strains is quite low compared to their glucose fermentation. In this study, two diploid xylose-utilizing S. cerevisiae strains, the industrial strain MA-R4 and the laboratory strain MA-B4, were employed to investigate the differences between anaerobic fermentation of xylose and glucose, and general differences between recombinant yeast strains, through genome-wide transcription analysis. In MA-R4, many genes related to ergosterol biosynthesis were expressed more highly with glucose than with xylose. Additionally, these ergosterol-related genes had higher transcript levels in MA-R4 than in MA-B4 during glucose fermentation. During xylose fermentation, several genes related to central metabolic pathways that typically increase during growth on non-fermentable carbon sources were expressed at higher levels in both strains. Xylose did not fully repress the genes encoding enzymes of the tricarboxylic acid and respiratory pathways, even under anaerobic conditions. In addition, several genes involved in spore wall metabolism and the uptake of ammonium, which are closely related to the starvation response, and many stress-responsive genes mediated by Msn2/4p, as well as trehalose synthase genes, increased in expression when fermenting with xylose, irrespective of the yeast strain. We further observed that transcript levels of genes involved in xylose metabolism, membrane transport functions, and ATP synthesis were higher in MA-R4 than in MA-B4 when strains were fermented with glucose or xylose. Our transcriptomic approach revealed the molecular events underlying the response to xylose or glucose and differences between MA-R4 and MA-B4. Xylose-utilizing S. cerevisiae strains may recognize xylose as a non-fermentable carbon source, which induces a starvation response and adaptation to oxidative stress, resulting in the increased expression of stress-response genes.
    Full-text · Article · Jan 2014 · Microbial Cell Factories
  • Source
    • "In mammalian cells, it has been shown that NF-Y affects the histone acetylation landscape in gene promoters, thus influencing gene transcription [56]. The Hap complex binding motif CCAAT was first identified in yeast [57,58]. The yeast Hap complex plays a role as a transcriptional regulator during respiration and oxidative stress response and induces genes such as CYC1 (cytochrome c-iso-1), CYC7 (cytochrome c iso-2), CYT1 (cytochrome c1), and CTT1 (catalase). "
    [Show abstract] [Hide abstract]
    ABSTRACT: There is extensive and unequivocal evidence that secondary metabolism in filamentous fungi and plants is associated with oxidative stress. In support of this idea, transcription factors related to oxidative stress response in yeast, plants, and fungi have been shown to participate in controlling secondary metabolism. Aflatoxin biosynthesis, one model of secondary metabolism, has been demonstrated to be triggered and intensified by reactive oxygen species buildup. An oxidative stress-related bZIP transcription factor AtfB is a key player in coordinate expression of antioxidant genes and genes involved in aflatoxin biosynthesis. Recent findings from our laboratory provide strong support for a regulatory network comprised of at least four transcription factors that bind in a highly coordinated and timely manner to promoters of the target genes and regulate their expression. In this review, we will focus on transcription factors involved in co-regulation of aflatoxin biosynthesis with oxidative stress response in aspergilli, and we will discuss the relationship of known oxidative stress-associated transcription factors and secondary metabolism in other organisms. We will also talk about transcription factors that are involved in oxidative stress response, but have not yet been demonstrated to be affiliated with secondary metabolism. The data support the notion that secondary metabolism provides a secondary line of defense in cellular response to oxidative stress.
    Full-text · Article · Apr 2013 · Toxins
Show more