Heat stress induced Cup9 dependent transcriptional regulation of Sir2

Hsp90, the concertmaster: tuning transcription.

Abstract: In the last decade Hsp90 has emerged as a major regulator of cancer cell growth and proliferation In cancer cells, it assists in giving maturation to oncogenic proteins including several kinases and transcription factors Recent studies have shown that apart from its chaperone activity, it also imparts regulation of transcription machinery and thereby alters the cellular physiology Hsp90 and its co-chaperones modulate transcription at-least at three different levels In the first place, they alter the steady-state levels of certain transcription factors in response to various physiological cues Secondly, they modulate the activity of certain epigenetic modifiers, such as histone deacetylases or DNA methyl transferases, and thereby respond to the change in the environment Thirdly, they participate in the eviction of histones from the promoter region of certain genes and thereby turn on gene expression In this review, we discuss the role of Hsp90 in all the three aforementioned mechanisms of transcriptional control, taking examples from various model organisms with a special emphasis on cancer progression

Quantitative Analysis of Hsp90-Client Interactions Reveals Principles of Substrate Recognition

Abstract: National Institutes of Health (U.S.). Genomics Based Drug Discovery-Driving Medical Project (Grant UL1-DE019585)

Yeast epigenetics: the inheritance of histone modification states.

Abstract: Saccharomyces cerevisiae (budding yeast) and Schizosaccharomyces pombe (fission yeast) are two of the most recognised and well-studied model systems for epigenetic regulation and the inheritance of chromatin states. Their silent loci serve as a proxy for heterochromatic chromatin in higher eukaryotes, and as such both species have provided a wealth of information on the mechanisms behind the establishment and maintenance of epigenetic states, not only in yeast, but in higher eukaryotes. This review focuses specifically on the role of histone modifications in governing telomeric silencing in S. cerevisiae and centromeric silencing in S. pombe as examples of genetic loci that exemplify epigenetic inheritance. We discuss the recent advancements that for the first time provide a mechanistic understanding of how heterochromatin, dictated by histone modifications specifically, is preserved during S-phase. We also discuss the current state of our understanding of yeast nucleosome dynamics during DNA replication, an essential component in delineating the contribution of histone modifications to epigenetic inheritance.

Hsp90 induces increased genomic instability toward DNA-damaging agents by tuning down RAD53 transcription.

Abstract: It is well documented that elevated body temperature causes tumors to regress upon radiotherapy. However, how hyperthermia induces DNA damage sensitivity is not clear. We show that a transient heat shock and particularly the concomitant induction of Hsp90 lead to increased genomic instability under DNA-damaging conditions. Using Saccharomyces cerevisiae as a model eukaryote, we demonstrate that elevated levels of Hsp90 attenuate efficient DNA damage signaling and dictate preferential use of the potentially mutagenic double-strand break repair pathway. We show that under normal physiological conditions, Hsp90 negatively regulates RAD53 transcription to suppress DNA damage checkpoint activation. However, under DNA damaging conditions, RAD53 is derepressed, and the increased level of Rad53p triggers an efficient DNA damage response. A higher abundance of Hsp90 causes increased transcriptional repression on RAD53 in a dose-dependent manner, which could not be fully derepressed even in the presence of DNA damage. Accordingly, cells behave like a rad53 loss-of-function mutant and show reduced NHEJ efficiency, with a drastic failure to up-regulate RAD51 expression and manifestly faster accumulation of CLN1 and CLN2 in DNA-damaged G1, cells leading to premature release from checkpoint arrest. We further demonstrate that Rad53 overexpression is able to rescue all of the aforementioned deleterious effects caused by Hsp90 overproduction.

Double strand breaks may be a missing link between entropy and aging.

Abstract: It has been previously suggested that an increase in entropy production leads to aging. However, the mechanisms linking increased entropy production in living mass to aging are currently unclear. Even though entropy cannot be easily associated with any specific molecular damage, the increase of entropy in structural mass may be connected with heat stress, which is known to generate double strand breaks. Double strand breaks, which are in turn known to play an important role in process of aging, are thus connected to both aging and an increase of entropy. In view of these associations, we propose a new model where the increase of entropy leads to the formation of double strand breaks, resulting in an aging phenotype. This not only offers a new perspective on aging research and facilitates experimental validation, but could also serve as a useful explanatory tool.

Additional modules for versatile and economical PCR-based gene deletion and modification in Saccharomyces cerevisiae

Abstract: An important recent advance in the functional analysis of Saccharomyces cerevisiae genes is the development of the one-step PCR-mediated technique for deletion and modification of chromosomal genes This method allows very rapid gene manipulations without requiring plasmid clones of the gene of interest We describe here a new set of plasmids that serve as templates for the PCR synthesis of fragments that allow a variety of gene modifications Using as selectable marker the S cerevisiae TRP1 gene or modules containing the heterologous Schizosaccharomyces pombe his5 + or Escherichia coli kan r gene, these plasmids allow gene deletion, gene overexpression (using the regulatable GAL1 promoter), C- or N-terminal protein tagging [with GFP(S65T), GST, or the 3HA or 13Myc epitope], and partial N- or C-terminal deletions (with or without concomitant protein tagging) Because of the modular nature of the plasmids, they allow eYcient and economical use of a small number of PCR primers for a wide variety of gene manipulations Thus, these plasmids should further facilitate the rapid analysis of gene function in S cerevisiae ? 1998 John Wiley & Sons, Ltd

Genomic expression programs in the response of yeast cells to environmental changes.

Abstract: We explored genomic expression patterns in the yeast Saccharomyces cerevisiae responding to diverse environmental transitions. DNA microarrays were used to measure changes in transcript levels over time for almost every yeast gene, as cells responded to temperature shocks, hydrogen peroxide, the superoxide-generating drug menadione, the sulfhydryl-oxidizing agent diamide, the disulfide-reducing agent dithiothreitol, hyper- and hypo-osmotic shock, amino acid starvation, nitrogen source depletion, and progression into stationary phase. A large set of genes (approximately 900) showed a similar drastic response to almost all of these environmental changes. Additional features of the genomic responses were specialized for specific conditions. Promoter analysis and subsequent characterization of the responses of mutant strains implicated the transcription factors Yap1p, as well as Msn2p and Msn4p, in mediating specific features of the transcriptional response, while the identification of novel sequence elements provided clues to novel regulators. Physiological themes in the genomic responses to specific environmental stresses provided insights into the effects of those stresses on the cell.

Transcriptional silencing and longevity protein Sir2 is an NAD-dependent histone deacetylase

Abstract: Yeast Sir2 is a heterochromatin component that silences transcription at silent mating loci, telomeres and the ribosomal DNA, and that also suppresses recombination in the rDNA and extends replicative life span. Mutational studies indicate that lysine 16 in the amino-terminal tail of histone H4 and lysines 9, 14 and 18 in H3 are critically important in silencing, whereas lysines 5, 8 and 12 of H4 have more redundant functions. Lysines 9 and 14 of histone H3 and lysines 5, 8 and 16 of H4 are acetylated in active chromatin and hypoacetylated in silenced chromatin, and overexpression of Sir2 promotes global deacetylation of histones, indicating that Sir2 may be a histone deacetylase. Deacetylation of lysine 16 of H4 is necessary for binding the silencing protein, Sir3. Here we show that yeast and mouse Sir2 proteins are nicotinamide adenine dinucleotide (NAD)-dependent histone deacetylases, which deacetylate lysines 9 and 14 of H3 and specifically lysine 16 of H4. Our analysis of two SIR2 mutations supports the idea that this deacetylase activity accounts for silencing, recombination suppression and extension of life span in vivo. These findings provide a molecular framework of NAD-dependent histone deacetylation that connects metabolism, genomic silencing and ageing in yeast and, perhaps, in higher eukaryotes.

Histone deacetylases (HDACs): characterization of the classical HDAC family

Abstract: Transcriptional regulation in eukaryotes occurs within a chromatin setting, and is strongly influenced by the post-translational modification of histones, the building blocks of chromatin, such as methylation, phosphorylation and acetylation. Acetylation is probably the best understood of these modifications: hyperacetylation leads to an increase in the expression of particular genes, and hypoacetylation has the opposite effect. Many studies have identified several large, multisubunit enzyme complexes that are responsible for the targeted deacetylation of histones. The aim of this review is to give a comprehensive overview of the structure, function and tissue distribution of members of the classical histone deacetylase (HDAC) family, in order to gain insight into the regulation of gene expression through HDAC activity. SAGE (serial analysis of gene expression) data show that HDACs are generally expressed in almost all tissues investigated. Surprisingly, no major differences were observed between the expression pattern in normal and malignant tissues. However, significant variation in HDAC expression was observed within tissue types. HDAC inhibitors have been shown to induce specific changes in gene expression and to influence a variety of other processes, including growth arrest, differentiation, cytotoxicity and induction of apoptosis. This challenging field has generated many fascinating results which will ultimately lead to a better understanding of the mechanism of gene transcription as a whole.

TRANSFAC® and its module TRANSCompel®: transcriptional gene regulation in eukaryotes

Abstract: The TRANSFAC database on transcription factors, their binding sites, nucleotide distribution matrices and regulated genes as well as the complementing database TRANSCompel on composite elements have been further enhanced on various levels. A new web interface with different search options and integrated versions of Match and Patch provides increased functionality for TRANSFAC. The list of databases which are linked to the common GENE table of TRANSFAC and TRANSCompel has been extended by: Ensembl, UniGene, EntrezGene, HumanPSD and TRANSPRO. Standard gene names from HGNC, MGI and RGD, are included for human, mouse and rat genes, respectively. With the help of InterProScan, Pfam, SMART and PROSITE domains are assigned automatically to the protein sequences of the transcription factors. TRANSCompel contains now, in addition to the COMPEL table, a separate table for detailed information on the experimental EVIDENCE on which the composite elements are based. Finally, for TRANSFAC, in respect of data growth, in particular the gain of Drosophila transcription factor binding sites (by courtesy of the Drosophila DNase I footprint database) and of Arabidopsis factors (by courtesy of DATF, Database of Arabidopsis Transcription Factors) has to be stressed. The here described public releases, TRANSFAC 7.0 and TRANSCompel 7.0, are accessible under http://www.gene-regulation.com/pub/databases.html.