scholarly journals Genetic and Epigenetic Determinants Establish a Continuum of Hsf1 Occupancy and Activity Across the Yeast Genome

2018 ◽  
Author(s):  
David Pincus ◽  
Jayamani Anandhakumar ◽  
Prathapan Thiru ◽  
Michael J. Guertin ◽  
Alexander M. Erkine ◽  
...  
Keyword(s):  
Heat Shock ◽  
Nucleosome Occupancy ◽  
Fold Increase ◽  
Yeast Genome ◽  
Heat Shock Factor 1 ◽  
Rna Seq ◽  
Heat Shock Element ◽  
Pioneer Factors ◽  
Accessible Chromatin

AbstractHeat Shock Factor 1 (Hsf1) is the master transcriptional regulator of molecular chaperones and binds to the same cis-acting element - Heat Shock Element (HSE) - across the eukaryotic lineage. In budding yeast, Hsf1 drives transcription of ~20 genes essential to maintain proteostasis under basal conditions, yet its specific targets and extent of inducible binding during heat shock remain unclear. Here we combine Hsf1 ChIP-seq, nascent RNA-seq and Hsf1 nuclear depletion to quantify Hsf1 binding and transcription across the yeast genome. Hsf1 binds 74 loci during acute heat shock, 46 of which are linked to genes with strong Hsf1-dependent transcription. Most of these targets show detectable Hsf1 binding under basal conditions, but basal occupancy and heat shock-inducible binding both vary over two orders of magnitude. Notably, Hsf1’s induced DNA binding leads to a disproportionate (up to 50-fold) increase in nascent transcription. While variation in basal Hsf1 occupancy poorly correlates with the strength of the HSE, promoters with high basal Hsf1 occupancy have nucleosome-depleted regions due to the presence of ‘pioneer’ factors. Such accessible chromatin may be critical for Hsf1 occupancy of its genomic sites as the activator is incapable of binding HSEs embedded within a stable nucleosome in vitro. In response to heat shock, however, Hsf1 is able to gain access to nucleosomal sites and promotes chromatin remodeling with the RSC complex playing a key role. We propose that the interplay between nucleosome occupancy, HSE strength and active Hsf1 levels allows cells to precisely tune expression of the proteostasis network.

scholarly journals Genetic and epigenetic determinants establish a continuum of Hsf1 occupancy and activity across the yeast genome

2018 ◽  
Vol 29 (26) ◽  
pp. 3168-3182 ◽  
Author(s):  
David Pincus ◽  
Jayamani Anandhakumar ◽  
Prathapan Thiru ◽  
Michael J. Guertin ◽  
Alexander M. Erkine ◽  
...  
Keyword(s):  
Heat Shock ◽  
Chromatin Immunoprecipitation ◽  
Fold Increase ◽  
Yeast Genome ◽  
Heat Shock Factor 1 ◽  
Rna Seq ◽  
Heat Shock Element ◽  
Cis Acting ◽  
Chromatin Immunoprecipitation Sequencing ◽  
Pioneer Factors

Heat shock factor 1 is the master transcriptional regulator of molecular chaperones and binds to the same cis-acting heat shock element (HSE) across the eukaryotic lineage. In budding yeast, Hsf1 drives the transcription of ∼20 genes essential to maintain proteostasis under basal conditions, yet its specific targets and extent of inducible binding during heat shock remain unclear. Here we combine Hsf1 chromatin immunoprecipitation sequencing (seq), nascent RNA-seq, and Hsf1 nuclear depletion to quantify Hsf1 binding and transcription across the yeast genome. We find that Hsf1 binds 74 loci during acute heat shock, and these are linked to 46 genes with strong Hsf1-dependent expression. Notably, Hsf1’s induced DNA binding leads to a disproportionate (∼7.5-fold) increase in nascent transcription. Promoters with high basal Hsf1 occupancy have nucleosome-depleted regions due to the presence of “pioneer factors.” These accessible sites are likely critical for Hsf1 occupancy as the activator is incapable of binding HSEs within a stably positioned, reconstituted nucleosome. In response to heat shock, however, Hsf1 accesses nucleosomal sites and promotes chromatin disassembly in concert with the Remodels Structure of Chromatin (RSC) complex. Our data suggest that the interplay between nucleosome positioning, HSE strength, and active Hsf1 levels allows cells to precisely tune expression of the proteostasis network.

Hypothermia enhances phosphorylation of IκB kinase and prolongs nuclear localization of NF-κB in lipopolysaccharide-activated macrophages

2005 ◽  
Vol 289 (5) ◽  
pp. C1114-C1121 ◽  
Author(s):  
Karen D. Fairchild ◽  
Ishwar S. Singh ◽  
Heather C. Carter ◽  
Lisa Hester ◽  
Jeffrey D. Hasday
Keyword(s):  
Heat Shock ◽  
Fold Increase ◽  
Mononuclear Phagocytes ◽  
Nuclear Accumulation ◽  
Clinical Settings ◽  
Heat Shock Factor 1 ◽  
Iκb Kinase ◽  
Tnf Α ◽  
Nuclear Events

Hypothermia (HT) has been associated with both beneficial and detrimental consequences in various pathophysiological states. While HT is generally thought to have anti-inflammatory and cytoprotective effects, we have previously shown that moderate in vitro HT prolongs TNF-α production by LPS-stimulated mononuclear phagocytes, in part by prolonging TNF-α gene transcription and activation of the pleiotropic transcription factor NF-κB. In this study, we have further characterized the effect of moderate (32°C) and marked (28°C) HT in human monocytic THP-1 cells by showing that even short (2 h) exposure to HT followed by a return to normothermic conditions for 22 h resulted in augmented and prolonged production of TNF-α. Production of heat shock protein 72 and activation of heat shock factor 1 are not affected by HT in these studies, suggesting that the effect is not part of a generalized stress response. Using immunoblotting, we have shown that HT augments phosphorylation of IKK-β and IKK-α (up to an 8-fold increase at 28°C and a 3.6-fold increase at 32°C vs. 37°C). Furthermore, nuclear accumulation of NF-κB p65 was significantly prolonged in hypothermic cells (1.4- and 2.5-fold more nuclear p65 at 2 and 4 h at 28 vs. 37°C). Reexpression of IκB-α, which contributes to the termination of NF-κB-dependent transcription, was delayed several hours in HT-exposed cells. Thus we have shown that clinically relevant HT alters both cytosolic and nuclear events responsible for NF-κB activation and deactivation. Enhanced NF-κB activation may contribute to the immunomodulatory effects of HT in various clinical settings.

scholarly journals HSP90 Interacts with and Regulates the Activity of Heat Shock Factor 1 in Xenopus Oocytes

1998 ◽  
Vol 18 (9) ◽  
pp. 4949-4960 ◽  
Author(s):  
Adnan Ali ◽  
Steven Bharadwaj ◽  
Ruth O’Carroll ◽  
Nick Ovsenek
Keyword(s):  
Heat Shock ◽  
Transcriptional Activation ◽  
Cellular Signaling ◽  
Heat Shock Factor ◽  
Heat Shock Factor 1 ◽  
Heat Shock Element ◽  
Nuclear Extracts ◽  
Multistep Process

ABSTRACT Transcriptional activation of heat shock genes is a reversible and multistep process involving conversion of inactive heat shock factor 1 (HSF1) monomers into heat shock element (HSE)-binding homotrimers, hyperphosphorylation, and further modifications that induce full transcriptional competence. HSF1 is controlled by multiple regulatory mechanisms, including suppression by additional cellular factors, physical interactions with HSP70, and integration into different cellular signaling cascades. However, the signaling mechanisms by which cells respond to stress and control the HSF1 activation-deactivation pathway are not known. Here we demonstrate that HSP90, a cellular chaperone known to regulate several signal transduction molecules and transcription factors, functions in the regulation of HSF1. The existence of HSF1-HSP90 heterocomplexes was shown by coimmunoprecipitation of HSP90 with HSF1 from unshocked and heat-shocked nuclear extracts, recognition of HSF1-HSE complexes in vitro by using HSP90 antibodies (Abs), and recognition of HSF1 in vivo by HSP90 Abs microinjected directly into oocyte nuclei. The functional impact of HSP90-HSF1 interactions was analyzed by using two strategies: direct nuclear injection of HSP90 Abs and treatment of cells with geldanamycin (GA), an agent that specifically blocks the chaperoning activity of HSP90. Both HSP90 Abs and GA delayed the disassembly of HSF1 trimers during recovery from heat shock and specifically inhibited heat-induced transcription from a chloramphenicol acetyltransferase reporter construct under control of the hsp70 promoter. HSP90 Abs activated HSE binding in the absence of heat shock, an effect that could be reversed by subsequent injection of purified HSP90. GA did not activate HSE binding under nonshock conditions but increased the quantity of HSE binding induced by heat shock. On the basis of these findings and the known properties of HSP90, we propose a new regulatory model in which HSP90 participates in modulating HSF1 at different points along the activation-deactivation pathway, influencing the interconversion between monomeric and trimeric conformations as well as transcriptional activation. We also put forth the hypothesis that HSP90 links HSF1 to cellular signaling molecules coordinating the stress response.

scholarly journals Cytoprotective Effects of Taurine on Heat-Induced Bovine Mammary Epithelial Cells In Vitro

Cells ◽  
2021 ◽  
Vol 10 (2) ◽  
pp. 258
Author(s):  
Hui Bai ◽  
Tingting Li ◽  
Yan Yu ◽  
Ningcong Zhou ◽  
Huijuan Kou ◽  
...  
Keyword(s):  
T Cells ◽  
Heat Stress ◽  
Heat Shock ◽  
T Antigen ◽  
Mammary Epithelial ◽  
Epithelial Cell Line ◽  
Heat Shock Factor 1 ◽  
Alveolar Cells ◽  
Bovine Mammary Epithelial Cells

It is a widely known that heat stress induces a reduction in milk production in cows and impairs their overall health. Studies have shown that taurine protects tissues and organs under heat stress. However, there have yet to be studies showing the functions of taurine in mammary alveolar cells-large T antigen (MAC-T) (a bovine mammary epithelial cell line) cells under heat shock. Therefore, different concentrations of taurine (10 mM, 50 mM, and 100 mM) were tested to determine the effects on heat-induced MAC-T cells. The results showed that taurine protected the cells against heat-induced damage as shown by morphological observations in conjunction with suppressed the translocation and expression of heat shock factor 1 (HSF1). Moreover, taurine not only reversed the decline in antioxidase (superoxide dismutase (SOD) and glutathione peroxidase (GSH-PX)) activities but also attenuated the accumulation of malondialdehyde (MDA). Meanwhile, mitochondrial damage (morphology and complex I activity) resulting from heat exposure was mitigated. Taurine also alleviated the rates of cell apoptosis and markedly depressed the mRNA expressions of BCL2 associated X, apoptosis regulator (BAX) and caspase3. Furthermore, compared with the heat stress (HS) group, the protein levels of caspase3 and cleaved caspase3 were decreased in all taurine groups. In summary, taurine improves the antioxidant and anti-apoptosis ability of MAC-T cells thereby alleviates damage of cells due to heat insults.

scholarly journals Phosphorylation of Serine 303 Is a Prerequisite for the Stress-Inducible SUMO Modification of Heat Shock Factor 1

2003 ◽  
Vol 23 (8) ◽  
pp. 2953-2968 ◽  
Author(s):  
Ville Hietakangas ◽  
Johanna K. Ahlskog ◽  
Annika M. Jakobsson ◽  
Maria Hellesuo ◽  
Niko M. Sahlberg ◽  
...  
Keyword(s):  
Heat Shock ◽  
Phosphorylation Site ◽  
Stress Granules ◽  
Heat Shock Factor ◽  
Transcriptional Level ◽  
Heat Shock Factor 1 ◽  
Protection Mechanism ◽  
Sumo Modification

ABSTRACT The heat shock response, which is accompanied by a rapid and robust upregulation of heat shock proteins (Hsps), is a highly conserved protection mechanism against protein-damaging stress. Hsp induction is mainly regulated at transcriptional level by stress-inducible heat shock factor 1 (HSF1). Upon activation, HSF1 trimerizes, binds to DNA, concentrates in the nuclear stress granules, and undergoes a marked multisite phosphorylation, which correlates with its transcriptional activity. In this study, we show that HSF1 is modified by SUMO-1 and SUMO-2 in a stress-inducible manner. Sumoylation is rapidly and transiently enhanced on lysine 298, located in the regulatory domain of HSF1, adjacent to several critical phosphorylation sites. Sumoylation analyses of HSF1 phosphorylation site mutants reveal that specifically the phosphorylation-deficient S303 mutant remains devoid of SUMO modification in vivo and the mutant mimicking phosphorylation of S303 promotes HSF1 sumoylation in vitro, indicating that S303 phosphorylation is required for K298 sumoylation. This finding is further supported by phosphopeptide mapping and analysis with S303/7 phosphospecific antibodies, which demonstrate that serine 303 is a target for strong heat-inducible phosphorylation, corresponding to the inducible HSF1 sumoylation. A transient phosphorylation-dependent colocalization of HSF1 and SUMO-1 in nuclear stress granules provides evidence for a strictly regulated subnuclear interplay between HSF1 and SUMO.

scholarly journals In vivo growth of a murine lymphoma cell line alters regulation of expression of HSP72.

1995 ◽  
Vol 15 (2) ◽  
pp. 1071-1078 ◽  
Author(s):  
S Davidson ◽  
P Høj ◽  
T Gabriele ◽  
R L Anderson
Keyword(s):  
Heat Shock ◽  
Cell Line ◽  
B Cell Lymphoma ◽  
Lymphoma Cell ◽  
Lymphoma Cell Line ◽  
Regulation Of Expression ◽  
Heat Shock Element ◽  
Stressed Cells

We have identified a murine B-cell lymphoma cell line, CH1, that has a much-diminished capacity to express increased levels of heat shock proteins in response to heat stress in vitro. In particular, these cells cannot synthesize the inducible 72-kDa heat shock protein (HSP72) which is normally expressed at high levels in stressed cells. We show here that CH1 fails to transcribe HSP72 mRNA after heat shock, even though the heat shock transcription factor, HSF, is activated correctly. After heat shock, HSF from CH1 is found in the nucleus and is phosphorylated, trimerized, and capable of binding the heat shock element. We propose that additional signals which CH1 cells are unable to transduce are normally required to activate hsp72 transcription in vitro. Surprisingly, we have found that when the CH1 cells are heated in situ in a mouse, they show normal expression of HSP72 mRNA and protein. Therefore, CH1 cells have a functional hsp72 gene which can be transcribed and translated when the cells are in an appropriate environment. A diffusible factor present in ascites fluid is capable of restoring normal HSP72 induction in CH1 cells. We conclude that as-yet-undefined factors are required for regulation of the hsp72 gene or, alternatively, that heat shock in vivo causes activation of hsp70 through a novel pathway which the defect in CH1 has exposed and which is distinct from that operating in vitro. This unique system offers an opportunity to study a physiologically relevant pathway of heat shock induction and to biochemically define effectors involved in the mammalian stress response.

Modular recognition of 5-base-pair DNA sequence motifs by human heat shock transcription factor

1991 ◽  
Vol 11 (7) ◽  
pp. 3504-3514
Author(s):  
N F Cunniff ◽  
J Wagner ◽  
W D Morgan
Keyword(s):  
Transcription Factor ◽  
Heat Shock ◽  
Gene Promoter ◽  
Heat Shock Transcription Factor ◽  
Heat Shock Gene ◽  
Gene Promoters ◽  
Sequence Motifs ◽  
Binding Modes ◽  
Heat Shock Element

We investigated the recognition of the conserved 5-bp repeated motif NGAAN, which occurs in heat shock gene promoters of Drosophila melanogaster and other eukaryotic organisms, by human heat shock transcription factor (HSF). Extended heat shock element mutants of the human HSP70 gene promoter, containing additional NGAAN blocks flanking the original element, showed significantly higher affinity than the wild-type promoter element for human HSF in vitro. Protein-DNA contact positions were identified by hydroxyl radical protection, diethyl pyrocarbonate interference, and DNase I footprinting. New contacts in the mutant HSE constructs corresponded to the locations of additional NGAAN motifs. The pattern of binding indicated the occurrence of multiple DNA binding modes for HSF with the various constructs and was consistent with an oligomeric, possibly trimeric, structure of the protein. In contrast to the improved binding, the extended heat shock element mutant constructs did not exhibit dramatically increased heat-inducible transcription in transient expression assays with HeLa cells.

scholarly journals Expression of a constitutively active mutant of heat shock factor 1 under the control of testis-specific hst70 gene promoter in transgenic mice induces degeneration of seminiferous epithelium.

2003 ◽  
Vol 50 (2) ◽  
pp. 535-541 ◽  
Author(s):  
Wiesława Widłak ◽  
Konrad Benedyk ◽  
Natallia Vydra ◽  
Magdalena Głowala ◽  
Dorota Scieglińska ◽  
...  
Keyword(s):  
Heat Shock ◽  
Transgenic Mice ◽  
Target Genes ◽  
Gene Promoter ◽  
Basic Mechanism ◽  
Seminiferous Epithelium ◽  
High Rate ◽  
Heat Shock Factor 1 ◽  
Heat Shock Element ◽  
Constitutively Active

Heat shock activates in somatic cells a set of genes encoding heat shock proteins which function as molecular chaperones. The basic mechanism by which these genes are activated is the interaction of the specific transcription factor HSF1 with a regulatory DNA sequence called heat shock element (HSE). In higher eukaryotes HSF1 is present in unstressed cells as inactive monomers which, in response to cellular stress, aggregate into transcriptionally competent homotrimers. In the present paper we showed that the expression of a transgene encoding mutated constitutively active HSF1 placed under the control of a spermatocyte-specific promoter derived from the hst70 gene severely affects spermatogenesis. We found the testes of transgenic mice to be significantly smaller than those of wild-type males and histological analysis showed massive degeneration of the seminiferous epithelium. The lumen of tubules was devoid of spermatids and spermatozoa and using the TUNEL method we demonstrated a high rate of spermatocyte apoptosis. The molecular mechanism by which constitutively active HSF1 arrests spermatogenesis is not known so far. One can assume that HSF1 can either induce or repress so far unknown target genes involved in germ cell apoptosis.

Activation of heat shock factor 2 during hemin-induced differentiation of human erythroleukemia cells

1992 ◽  
Vol 12 (9) ◽  
pp. 4104-4111
Author(s):  
L Sistonen ◽  
K D Sarge ◽  
B Phillips ◽  
K Abravaya ◽  
R I Morimoto
Keyword(s):  
Heat Shock ◽  
Dna Binding ◽  
Binding Activity ◽  
Erythroid Cell ◽  
Hsp70 Gene ◽  
Mobility Shift ◽  
Heat Shock Element ◽  
Dna Binding Activity ◽  
Erythroleukemia Cells

Hemin induces nonterminal differentiation of human K562 erythroleukemia cells, which is accompanied by the expression of certain erythroid cell-specific genes, such as the embryonic and fetal globins, and elevated expression of the stress genes hsp70, hsp90, and grp78/BiP. Previous studies revealed that, as during heat shock, transcriptional induction of hsp70 in hemin-treated cells is mediated by activation of heat shock transcription factor (HSF), which binds to the heat shock element (HSE). We report here that hemin activates the DNA-binding activity of HSF2, whereas heat shock induces predominantly the DNA-binding activity of a distinct factor, HSF1. This constitutes the first example of HSF2 activation in vivo. Both hemin and heat shock treatments resulted in equivalent levels of HSF-HSE complexes as analyzed in vitro by gel mobility shift assay, yet transcription of the hsp70 gene was stimulated much less by hemin-induced HSF than by heat shock-induced HSF. Genomic footprinting experiments revealed that hemin-induced HSF and heat shock-induced HSF, HSF2, and HSF1, respectively, occupy the HSE of the human hsp70 promoter in a similar yet not identical manner. We speculate that the difference in occupancy and/or in the transcriptional abilities of HSF1 and HSF2 accounts for the observed differences in the stimulation of hsp70 gene transcription.

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