scholarly journals Genetic Modification of the Salmonella Membrane Physical State Alters the Pattern of Heat Shock Response

2010 ◽  
Vol 192 (7) ◽  
pp. 1988-1998 ◽  
Author(s):  
Amalia Porta ◽  
Zsolt Török ◽  
Ibolya Horvath ◽  
Silvia Franceschelli ◽  
László Vígh ◽  
...  
Keyword(s):  
Heat Shock ◽  
Transcriptional Activation ◽  
Lipid Membranes ◽  
Physical State ◽  
Stress Proteins ◽  
Protein Structures ◽  
Membrane Lipid ◽  
Heat Shock Genes ◽  
Protein Ratio ◽  
Molecular Arrangement

ABSTRACT It is now recognized that membranes are not simple physical barriers but represent a complex and dynamic environment that affects membrane protein structures and their functions. Recent data emphasize the role of membranes in sensing temperature changes, and it has been shown that the physical state of the plasma membrane influences the expression of a variety of genes such as heat shock genes. It has been widely shown that minor alterations in lipid membranes are critically involved in the conversion of signals from the environment to the transcriptional activation of heat shock genes. Previously, we have proposed that the composition, molecular arrangement, and physical state of lipid membranes and their organization have crucial roles in cellular responses during stress caused by physical and chemical factors as well as in pathological states. Here, we show that transformation of Salmonella enterica serovar Typhimurium LT2 (Salmonella Typhimurium) with a heterologous Δ12-desaturase (or with its trans-membrane regions) causes major changes in the pathogen's membrane dynamic. In addition, this pathogen is strongly impaired in the synthesis of major stress proteins (heat shock proteins) under heat shock. These data support the hypothesis that the perception of temperature in Salmonella is strictly controlled by membrane order and by a specific membrane lipid/protein ratio that ultimately causes transcriptional activation of heat shock genes. These results represent a previously unrecognized mode of sensing temperature variation used by this pathogen at the onset of infection.

Transcriptional activation of heat-shock genes in eukaryotes

1988 ◽  
Vol 66 (6) ◽  
pp. 584-593 ◽  
Author(s):  
Robert M. Tanguay
Keyword(s):  
Heat Shock ◽  
Transcriptional Activation ◽  
Regulatory Elements ◽  
Tata Box ◽  
Heat Shock Gene ◽  
Regulatory Sequence ◽  
Dependent Manner ◽  
Heat Shock Genes ◽  
Isolation And Characterization ◽  
Nuclease Mapping

Prokaryotes and eukaryotes respond to thermal or various chemical stresses by the rapid induction of a group of genes collectively referred to as the heat shock genes. In eucaryotes, the expression of these genes is primarily regulated at the transcriptional level. The early observations that transfected heat shock genes were inducible in heterologous systems suggested the existence of common regulatory elements in these ubiquitous genes. Sequence analysis of cloned Drosophila heat shock genes revealed a conserved 14 base pair (bp) inverted repeat, which is essential for heat induction. This regulatory sequence, referred to as the heat shock element (HSE), is found in multiple imperfect copies upstream of the TATA box of all heat shock genes. While studies in heterologous systems indicated that a single copy of HSE was sufficient for inducibility, further analysis in homologous assays suggests that multiple HSE can act in a cooperative way and that the efficiency of transcriptional activation is related, within limits, to the number of HSE. Comparative analysis of heat shock genes reveals that HSE can be positioned at different distances from the TATA box in either orientation, a behavior reminiscent of enhancer elements. However, the presence of HSE does not necessarily confer heat inducibility, as shown by their presence in the constitutively expressed but non-heat-inducible homologous cognate genes.Footprinting and nuclease mapping have been used to show that a protein factor (HSTF: heat shock transcription factor) binds to the HSE element, activating heat shock gene transcription in a dose-dependent manner. The recent progress in the isolation and characterization of HSTF in Drosophila, yeast, and human cells is reviewed. Finally, different models suggested to account for the positive regulation of heat shock genes by the HSTF are presented.

scholarly journals Complex modes of heat shock factor activation.

1990 ◽  
Vol 10 (2) ◽  
pp. 752-759 ◽  
Author(s):  
V Zimarino ◽  
C Tsai ◽  
C Wu
Keyword(s):  
Protein Synthesis ◽  
Heat Shock ◽  
Transcriptional Activation ◽  
Heat Shock Factor ◽  
Binding Activity ◽  
Regulatory Sequence ◽  
Heat Shock Genes ◽  
Heat Shock Element ◽  
Complex Modes ◽  
Environmental Temperatures

Eucaryotic organisms respond to elevated environmental temperatures by rapidly activating the expression of heat shock genes. The transcriptional activation of heat shock genes is mediated by a conserved upstream regulatory sequence, the heat shock element (HSE). Using an HSE-binding assay, we show that a cellular factor present in a range of vertebrate species binds specifically to the HSE. This factor is presumably the transcriptional activator of heat shock genes, heat shock factor (HSF). In vertebrates, the binding of HSF to the HSE was induced when cells were subjected to heat shock at high temperatures, even in the absence of protein synthesis. Under mild heat shock conditions, HSF binding was induced to a lesser extent, but this induction required protein synthesis, suggesting that synthesis of HSF itself, or an activating factor, is necessary for response to heat shock at intermediate temperatures. The inducibility of HSF binding in higher eucaryotes is contrasted with constitutive HSF binding activity in fungi. It appears that despite conservation of the HSE in evolution, the means by which HSF is activated to bind DNA in higher and lower eucaryotes may have diverged.

Complex modes of heat shock factor activation

1990 ◽  
Vol 10 (2) ◽  
pp. 752-759
Author(s):  
V Zimarino ◽  
C Tsai ◽  
C Wu
Keyword(s):  
Protein Synthesis ◽  
Heat Shock ◽  
Transcriptional Activation ◽  
Heat Shock Factor ◽  
Binding Activity ◽  
Regulatory Sequence ◽  
Heat Shock Genes ◽  
Heat Shock Element ◽  
Complex Modes ◽  
Environmental Temperatures

Eucaryotic organisms respond to elevated environmental temperatures by rapidly activating the expression of heat shock genes. The transcriptional activation of heat shock genes is mediated by a conserved upstream regulatory sequence, the heat shock element (HSE). Using an HSE-binding assay, we show that a cellular factor present in a range of vertebrate species binds specifically to the HSE. This factor is presumably the transcriptional activator of heat shock genes, heat shock factor (HSF). In vertebrates, the binding of HSF to the HSE was induced when cells were subjected to heat shock at high temperatures, even in the absence of protein synthesis. Under mild heat shock conditions, HSF binding was induced to a lesser extent, but this induction required protein synthesis, suggesting that synthesis of HSF itself, or an activating factor, is necessary for response to heat shock at intermediate temperatures. The inducibility of HSF binding in higher eucaryotes is contrasted with constitutive HSF binding activity in fungi. It appears that despite conservation of the HSE in evolution, the means by which HSF is activated to bind DNA in higher and lower eucaryotes may have diverged.

scholarly journals Protein Phosphatase 2A Activity Affects Histone H3 Phosphorylation and Transcription in Drosophila melanogaster

2003 ◽  
Vol 23 (17) ◽  
pp. 6129-6138 ◽  
Author(s):  
Scott J. Nowak ◽  
Chi-Yun Pai ◽  
Victor G. Corces
Keyword(s):  
Heat Shock ◽  
Transcriptional Activation ◽  
Protein Phosphatase ◽  
Protein A ◽  
Polytene Chromosomes ◽  
Histone H3 ◽  
Heat Shock Genes ◽  
Pp2a Activity ◽  
H3 Phosphorylation ◽  
Genome Wide

ABSTRACT Transcriptional activation of the heat shock genes during the heat shock response in Drosophila has been intimately linked to phosphorylation of histone H3 at serine 10, whereas repression of non-heat-shock genes correlates with dephosphorylation of histone H3. It is then possible that specific kinase and/or phosphatase activities may regulate histone phosphorylation and therefore transcription activation and repression, respectively. We find that treatment of cells with strong phosphatase inhibitors interferes with the genome-wide dephosphorylation of histone H3 normally observed at non-heat-shock genes during heat shock. Mutants in protein phosphatase type 2A (PP2A) also display reduced genome-wide H3 dephosphorylation, and sites of H3 phosphorylation that do not contain heat shock genes remain transcriptionally active during heat shock in PP2A mutants. Finally, the SET protein, a potent and highly selective inhibitor of PP2A activity that inhibits PP2A-mediated dephosphorylation of Ser10-phosphorylated H3, is detected at transcriptionally active regions of polytene chromosomes. These results suggest that activation and repression of gene expression during heat shock might be regulated by changes in PP2A activity controlled by the SET protein.

A Novel Reconstitution Method for Inducing the Formation of Regular 2-Dimensional Arrays of Membrane Proteins and Lipids

1988 ◽  
Vol 46 ◽  
pp. 152-153 ◽  
Author(s):  
A. Engel ◽  
A. Holzenburg ◽  
K. Stauffer ◽  
J. Rosenbusch ◽  
U. Aebi
Keyword(s):  
Membrane Proteins ◽  
Flow Rate ◽  
Lipid Membranes ◽  
Functional Characterization ◽  
Dimensional Structure ◽  
Electron Crystallography ◽  
Peltier Element ◽  
Protein Ratio ◽  
Precise Control ◽  
3 Dimensional

Reconstitution of solubilized and purified membrane proteins in the presence of phospholipids into vesicles allows their functions to be studied by simple bulk measurements (e.g. diffusion of differently sized solutes) or by conductance measurements after transformation into planar membranes. On the other hand, reconstitution into regular protein-lipid arrays, usually forming at a specific lipid-to-protein ratio, provides the basis for determining the 3-dimensional structure of membrane proteins employing the tools of electron crystallography.To refine reconstitution conditions for reproducibly inducing formation of large and highly ordered protein-lipid membranes that are suitable for both electron crystallography and patch clamping experiments aimed at their functional characterization, we built a flow-dialysis device that allows precise control of temperature and flow-rate (Fig. 1). The flow rate is generated by a peristaltic pump and can be adjusted from 1 to 500 ml/h. The dialysis buffer is brought to a preselected temperature during its travel through a meandering path before it enters the dialysis reservoir. A Z-80 based computer controls a Peltier element allowing the temperature profile to be programmed as function of time.

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