scholarly journals Molecular Dissection of the Human Ubiquitin C Promoter Reveals Heat Shock Element Architectures with Activating and Repressive Functions

PLoS ONE ◽  
2015 ◽  
Vol 10 (8) ◽  
pp. e0136882 ◽  
Author(s):  
Rita Crinelli ◽  
Marzia Bianchi ◽  
Lucia Radici ◽  
Elisa Carloni ◽  
Elisa Giacomini ◽  
...  
Keyword(s):  
Heat Shock ◽  
Molecular Dissection ◽  
Heat Shock Element ◽  
Human Ubiquitin

scholarly journals Activation of the DNA-binding ability of human heat shock transcription factor 1 may involve the transition from an intramolecular to an intermolecular triple-stranded coiled-coil structure.

1994 ◽  
Vol 14 (11) ◽  
pp. 7557-7568 ◽  
Author(s):  
J Zuo ◽  
R Baler ◽  
G Dahl ◽  
R Voellmy
Keyword(s):  
Transcription Factor ◽  
Heat Stress ◽  
Heat Shock ◽  
Dna Binding ◽  
Hydrophobic Interactions ◽  
Coiled Coil ◽  
Heat Shock Transcription Factor ◽  
Single Amino Acid ◽  
Binding Ability ◽  
Heat Shock Element

Heat stress regulation of human heat shock genes is mediated by human heat shock transcription factor hHSF1, which contains three 4-3 hydrophobic repeats (LZ1 to LZ3). In unstressed human cells (37 degrees C), hHSF1 appears to be in an inactive, monomeric state that may be maintained through intramolecular interactions stabilized by transient interaction with hsp70. Heat stress (39 to 42 degrees C) disrupts these interactions, and hHSF1 homotrimerizes and acquires heat shock element DNA-binding ability. hHSF1 expressed in Xenopus oocytes also assumes a monomeric, non-DNA-binding state and is converted to a trimeric, DNA-binding form upon exposure of the oocytes to heat shock (35 to 37 degrees C in this organism). Because endogenous HSF DNA-binding activity is low and anti-hHSF1 antibody does not recognize Xenopus HSF, we employed this system for mapping regions in hHSF1 that are required for the maintenance of the monomeric state. The results of mutagenesis analyses strongly suggest that the inactive hHSF1 monomer is stabilized by hydrophobic interactions involving all three leucine zippers which may form a triple-stranded coiled coil. Trimerization may enable the DNA-binding function of hHSF1 by facilitating cooperative binding of monomeric DNA-binding domains to the heat shock element motif. This view is supported by observations that several different LexA DNA-binding domain-hHSF1 chimeras bind to a LexA-binding site in a heat-regulated fashion, that single amino acid replacements disrupting the integrity of hydrophobic repeats render these chimeras constitutively trimeric and DNA binding, and that LexA itself binds stably to DNA only as a dimer but not as a monomer in our assays.

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.

scholarly journals A Constitutive Heat Shock Element-binding Factor Is Immunologically Identical to the Ku Autoantigen

1995 ◽  
Vol 270 (25) ◽  
pp. 15277-15284 ◽  
Author(s):  
Dooha Kim ◽  
Honghai Ouyang ◽  
Shao-Hua Yang ◽  
Andre Nussenzweig ◽  
Paul Burgman ◽  
...  
Keyword(s):  
Heat Shock ◽  
Heat Shock Element ◽  
Binding Factor ◽  
Ku Autoantigen

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.

Coordinate changes in heat shock element-binding activity and HSP70 gene transcription rates in human cells

1988 ◽  
Vol 8 (11) ◽  
pp. 4736-4744
Author(s):  
D D Mosser ◽  
N G Theodorakis ◽  
R I Morimoto
Keyword(s):  
Protein Synthesis ◽  
Amino Acid ◽  
Heat Shock ◽  
Gene Transcription ◽  
Elevated Temperatures ◽  
Binding Activity ◽  
Heat Shock Gene ◽  
Hsp70 Gene ◽  
Heat Shock Element ◽  
Amino Acid Analogs

Activation of human heat shock gene transcription by heat shock, heavy metal ions, and amino acid analogs required the heat shock element (HSE) in the HSP70 promoter. Both heat shock- and metal ion-induced HSP70 gene transcription occurred independently of protein synthesis, whereas induction by amino acid analogs required protein synthesis. We identified a HSE-binding activity from control cells which was easily distinguished by a gel mobility shift assay from the stress-induced HSE-binding activity which appeared following heat shock or chemically induced stress. The kinetics of HSP70 gene transcription paralleled the rapid appearance of stress-induced HSE-binding activity. During recovery from heat shock, both the rate of HSP70 gene transcription and stress-induced HSE-binding activity levels declined and the control HSE-binding activity reappeared. The DNA contacts of the control and stress-induced HSE-binding activities deduced by methylation interference were similar but not identical. While stable complexes with HSE were formed with extracts from both control and stressed cells in vitro at 25 degrees C, only the stress-induced complex was detected when binding reactions were performed at elevated temperatures.

The novel EhHSTF7 transcription factor displays an oligomer state and recognizes a heat shock element in the Entamoeba histolytica parasite

2022 ◽  
Vol 162 ◽  
pp. 105349
Author(s):  
Fabiola Bello ◽  
Esther Orozco ◽  
Claudia G. Benítez-Cardoza ◽  
Absalom Zamorano-Carrillo ◽  
César A. Reyes-López ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Heat Shock ◽  
Entamoeba Histolytica ◽  
The Novel ◽  
Heat Shock Element

Heat shock factor can activate transcription while bound to nucleosomal DNA in Saccharomyces cerevisiae

1994 ◽  
Vol 14 (1) ◽  
pp. 189-199
Author(s):  
D S Pederson ◽  
T Fidrych
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Heat Shock ◽  
Transcription Initiation ◽  
Heat Shock Factor ◽  
Micrococcal Nuclease ◽  
Nucleosome Assembly ◽  
Heat Shock Element ◽  
Yeast Saccharomyces Cerevisiae ◽  
Nucleosomal Dna

After each round of replication, new transcription initiation complexes must assemble on promoter DNA. This process may compete with packaging of the same promoter sequences into nucleosomes. To elucidate interactions between regulatory transcription factors and nucleosomes on newly replicated DNA, we asked whether heat shock factor (HSF) could be made to bind to nucleosomal DNA in vivo. A heat shock element (HSE) was embedded at either of two different sites within a DNA segment that directs the formation of a stable, positioned nucleosome. The resulting DNA segments were coupled to a reporter gene and transfected into the yeast Saccharomyces cerevisiae. Transcription from these two plasmid constructions after induction by heat shock was similar in amount to that from a control plasmid in which HSF binds to nucleosome-free DNA. High-resolution genomic footprint mapping of DNase I and micrococcal nuclease cleavage sites indicated that the HSE in these two plasmids was, nevertheless, packaged in a nucleosome. The inclusion of HSE sequences within (but relatively close to the edge of) the nucleosome did not alter the position of the nucleosome which formed with the parental DNA fragment. Genomic footprint analyses also suggested that the HSE-containing nucleosome was unchanged by the induction of transcription. Quantitative comparisons with control plasmids ruled out the possibility that HSF was bound only to a small fraction of molecules that might have escaped nucleosome assembly. Analysis of the helical orientation of HSE DNA in the nucleosome indicated that HSF contacted DNA residues that faced outward from the histone octamer. We discuss the significance of these results with regard to the role of nucleosomes in inhibiting transcription and the normal occurrence of nucleosome-free regions in promoters.

Identification of cis and trans components of a novel heat shock stress regulatory pathway in Saccharomyces cerevisiae

1993 ◽  
Vol 13 (1) ◽  
pp. 248-256
Author(s):  
N Kobayashi ◽  
K McEntee
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Stress Response ◽  
Heat Shock ◽  
Reporter Gene ◽  
Heat Shock Element ◽  
Cis Acting ◽  
Heat Shock Stress ◽  
Specific Complex ◽  
Cis And Trans ◽  
Shock Stress

The stress-responsive DDR2 gene (previously called DDRA2) of Saccharomyces cerevisiae is transcribed at elevated levels following stress caused by heat shock or DNA damage. Previously, we identified a 51-bp promoter fragment, oligo31/32, which conferred heat shock inducibility on the heterologous CYC1-lacZ reporter gene in S. cerevisiae (N. Kobayashi and K. McEntee, Proc. Natl. Acad. Sci. USA 87:6550-6554, 1990). Using a series of synthetic oligonucleotides, we have identified a pentanucleotide, CCCCT (C4T), as an essential component of this stress response sequence. This element is not a binding site for the well-characterized heat shock transcription factor which recognizes a distinct cis-acting heat shock element in the promoters of many heat shock genes. Here we demonstrate the ability of oligonucleotides containing the C4T sequence to confer heat shock inducibility on the reporter gene and show that the presence of two such elements produces more than additive effects on induction. Gel retardation experiments have been used to demonstrate specific complex formation between C4T-containing fragments and one or more yeast proteins. Formation of these complexes was not competed by fragments containing mutations in the C4T sequence nor by heat shock element-containing competitor DNAs. Fragments containing the C4T element bound to a single 140-kDa polypeptide, distinct from heat shock transcription factors in yeast crude extracts. These experiments identify key cis- and trans-acting components of a novel heat shock stress response pathway in S. cerevisiae.

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