Identification of a Functional Medaka Heat Shock Promoter and Characterization of Its Ability to Induce Exogenous Gene Expression in Medaka in Vitro and In Vivo

2010 ◽  
Vol 27 (5) ◽  
pp. 410-415 ◽  
Author(s):  
Shoji Oda ◽  
Sachi Mikami ◽  
Yusuke Urushihara ◽  
Yasuhiko Murata ◽  
Yasuhiro Kamei ◽  
...  
Keyword(s):  
Gene Expression ◽  
Heat Shock ◽  
Heat Shock Promoter ◽  
Exogenous Gene

scholarly journals Identification and characterization of molecular interactions between mortalin/mtHsp70 and HSP60

2005 ◽  
Vol 391 (2) ◽  
pp. 185-190 ◽  
Author(s):  
Renu Wadhwa ◽  
Syuichi Takano ◽  
Kamaljit Kaur ◽  
Satoshi Aida ◽  
Tomoko Yaguchi ◽  
...  
Keyword(s):  
Heat Shock ◽  
Molecular Interactions ◽  
Heat Shock Protein 60 ◽  
Hsp60 Expression ◽  
Mitochondrial Hsp70 ◽  
Shock Proteins ◽  
First Time

Mortalin/mtHsp70 (mitochondrial Hsp70) and HSP60 (heat-shock protein 60) are heat-shock proteins that reside in multiple subcellular compartments, with mitochondria being the predominant one. In the present study, we demonstrate that the two proteins interact both in vivo and in vitro, and that the N-terminal region of mortalin is involved in these interactions. Suppression of HSP60 expression by shRNA (short hairpin RNA) plasmids caused the growth arrest of cancer cells similar to that obtained by suppression of mortalin expression by ribozymes. An overexpression of mortalin, but not of HSP60, extended the in vitro lifespan of normal fibroblasts (TIG-1). Taken together, this study for the first time delineates: (i) molecular interactions of HSP60 with mortalin; (ii) their co- and exclusive localizations in vivo; (iii) their involvement in tumorigenesis; and (iv) their functional distinction in pathways involved in senescence.

Exertional heat injury and gene expression changes: a DNA microarray analysis study

2004 ◽  
Vol 96 (5) ◽  
pp. 1943-1953 ◽  
Author(s):  
Larry A. Sonna ◽  
C. Bruce Wenger ◽  
Scott Flinn ◽  
Holly K. Sheldon ◽  
Michael N. Sawka ◽  
...  
Keyword(s):  
Gene Expression ◽  
Stress Response ◽  
Heat Shock ◽  
Heat Shock Proteins ◽  
Molecular Evidence ◽  
Peripheral Blood Mononuclear ◽  
Heat Injury ◽  
Shock Proteins

This study examined gene expression changes associated with exertional heat injury (EHI) in vivo and compared these changes to in vitro heat shock responses previously reported by our laboratory. Peripheral blood mononuclear cell (PBMC) RNA was obtained from four male Marine recruits (ages 17-19 yr) who presented with symptoms consistent with EHI, core temperatures ranging from 39.3 to 42.5°C, and elevations in serum enzymes such as creatine kinase. Controls were age- and gender-matched Marines from whom samples were obtained before and several days after an intense field-training exercise in the heat (“The Crucible”). Expression analysis was performed on Affymetrix arrays (containing ∼12,600 sequences) from pooled samples obtained at three times for EHI group (at presentation, 2-3 h after cooling, and 1-2 days later) and compared with control values (average signals from two chips representing pre- and post-Crucible samples). After post hoc filtering, the analysis identified 361 transcripts that had twofold or greater increases in expression at one or more of the time points assayed and 331 transcripts that had twofold or greater decreases in expression. The affected transcripts included sequences previously shown to be heat-shock responsive in PBMCs in vitro (including both heat shock proteins and non-heat shock proteins), a number of sequences whose changes in expression had not previously been noted as a result of in vitro heat shock in PBMCs (including several interferon-induced sequences), and several nonspecific stress response genes (including ubiquitin C and dual-specificity phosphatase-1). We conclude that EHI produces a broad stress response that is detectable in PBMCs and that heat stress per se can only account for some of the observed changes in transcript expression. The molecular evidence from these patients is thus consistent with the hypothesis that EHI can result from cumulative effects of multiple adverse interacting stimuli.

scholarly journals Characterization of the Human Artemis Promoter by Heterologous Gene Expression In Vitro and In Vivo

2011 ◽  
Vol 30 (10) ◽  
pp. 751-761 ◽  
Author(s):  
Megan M. Multhaup ◽  
Sweta Gurram ◽  
Kelly M. Podetz-Pedersen ◽  
Andrea D. Karlen ◽  
Debra L. Swanson ◽  
...  
Keyword(s):  
Gene Expression ◽  
Heterologous Gene Expression ◽  
Heterologous Gene

scholarly journals Modulation of the expression of sirtuins and var genes by heat shock in the malaria parasite Plasmodium falciparum: a pattern emerges

2021 ◽  
Author(s):  
Linda O. Anagu ◽  
David R. Hulse ◽  
Paul D. Horrocks ◽  
Srabasti J Chakravorty
Keyword(s):  
Gene Expression ◽  
Plasmodium Falciparum ◽  
Heat Shock ◽  
Malaria Parasite ◽  
Severe Disease ◽  
Expression Patterns ◽  
Stress Factors ◽  
Virulence Gene Expression

Abstract Background: In the malaria parasite Plasmodium falciparum the expression of ‘var’ virulence genes is regulated through epigenetic mechanisms. Its sirtuin epigenetic regulators have a direct effect on var gene expression patterns, are increased in a laboratory strain of P. falciparum exposed to heat shock and are positively associated with fever. A Gambia study extended this association to blood lactate and var genes commonly expressed in severe malaria, and between PfSir2A and group B var. A Kenyan study extended this association to between PfSir2A and overall var transcript level. These observations suggest a mechanism through which stress phenotypes in the human host might be sensed via a parasite sirtuin, and virulence gene expression modulated accordingly. Methods: In vitro experiments were conducted using laboratory and recently-laboratory-adapted Kenyan isolates of P. falciparum to follow up the correlative findings of the field study. To investigate a potential cause-and-effect relationship between host stress factors and parasite gene expression, qPCR was used to measure the expression of sirtuins and var genes after highly synchronous cultured parasites had been exposed to 2h or 6h of heat shock at 40°C or elevated lactate.Results: Heat shock was shown to influence the expression of PfSir2B in the trophozoites, whereas exposure to lactate was not. After the ring stages were exposed to heat shock; sirtuins, severe-disease-associated upsA and upsB var genes and var genes in general were not altered. More biological replicate experiments will be needed to confirm our observations. Conclusions: This study demonstrates that heat stress in laboratory and recently-laboratory-adapted isolates of P. falciparum results in a small increase in PfSir2B transcripts in the trophozoite stages only. By contrast, the association between hyperlactataemia and sirtuin/var gene expression that was previously observed in vivo appears to be coincidental rather than causative.

scholarly journals hsp72, a Host Determinant of Measles Virus Neurovirulence

2006 ◽  
Vol 80 (22) ◽  
pp. 11031-11039 ◽  
Author(s):  
Thomas Carsillo ◽  
Zachary Traylor ◽  
Changsun Choi ◽  
Stefan Niewiesk ◽  
Michael Oglesbee
Keyword(s):  
Gene Expression ◽  
Heat Shock ◽  
Heat Shock Protein ◽  
Protein Interactions ◽  
Measles Virus ◽  
Viral Rna ◽  
Viral Gene ◽  
Febrile Episodes

ABSTRACT Transient hyperthermia such as that experienced during febrile episodes increases expression of the major inducible 70-kDa heat shock protein (hsp72). Despite the relevance of febrile episodes to viral pathogenesis and the multiple in vitro roles of heat shock proteins in viral replication and gene expression, the in vivo significance of virus-heat shock protein interactions is unknown. The present work determined the in vivo relationship between hsp72 levels and neurovirulence of an hsp72-responsive virus using the mouse model of measles virus (MV) encephalitis. Transgenic C57BL/6 mice were created to constitutively overexpress hsp72 in neurons, and these mice were inoculated intracranially with Edmonston MV (Ed MV) at 42 h of age. The mean viral RNA burden in brain was approximately 2 orders of magnitude higher in transgenic animals than in nontransgenic animals 2 to 4 weeks postinfection, and this increased burden was associated with a fivefold increase in mortality. Mice were also challenged with an Ed MV variant exhibiting an attenuated in vitro response to hsp72-dependent stimulation of viral transcription (Ed N-522D). This virus exhibited an attenuated neuropathogenicity in transgenic mice, where mortality and viral RNA burdens were not significantly different from nontransgenic mice infected with either Ed N-522D or parent Ed MV. Collectively, these results indicate that hsp72 levels can serve as a host determinant of viral neurovirulence in C57BL/6 mice, reflecting the direct influence of hsp72 on viral gene expression.

scholarly journals Alteration of the expression of sirtuins and var genes by heat shock in the malaria parasite Plasmodium falciparum: a pattern emerges

2021 ◽  
Author(s):  
Linda O. Anagu ◽  
David R. Hulse ◽  
Paul D. Horrocks ◽  
Srabasti J Chakravorty
Keyword(s):  
Gene Expression ◽  
Plasmodium Falciparum ◽  
Heat Shock ◽  
Blood Lactate ◽  
Malaria Parasite ◽  
Severe Disease ◽  
Expression Patterns ◽  
Stress Factors

Abstract Background In the malaria parasite Plasmodium falciparum the expression of ‘var’ virulence genes is regulated through epigenetic mechanisms. P. falciparum sirtuin epigenetic regulators have a direct effect on their var gene expression patterns, are increased in the 3D7 laboratory strain on exposure to heat shock. A Gambia study showed an association with fever, blood lactate and var genes commonly expressed in severe malaria, and between PfSir2A and group B var. A Kenyan study extended this association to between PfSir2A and overall var transcript level. We investigated a causal link between heat shock or lactate levels and sirtuins or var expression. Methods In vitro experiments were conducted using laboratory and recently-laboratory-adapted Kenyan isolates of P. falciparum to follow up the correlative findings of the field study. To investigate a potential cause-and-effect relationship between host stress factors and parasite gene expression, qPCR was used to measure the expression of sirtuins and var genes after highly synchronous cultured parasites had been exposed to 2h or 6h of heat shock at 40°C or elevated lactate. Results Heat shock was shown to increase the expression of PfSir2B in the trophozoites, whereas exposure to lactate was not. After the ring stages were exposed to heat shock and lactate, there was no alteration in the expression of sirtuins and severe-disease-associated upsA and upsB var genes. The association between high blood lactate and sirtuin/var gene expression that was previously observed in vivo appears to be coincidental rather than causative. Conclusions This study demonstrates that heat stress in laboratory and recently-laboratory-adapted isolates of P. falciparum results in a small increase in PfSir2B transcripts in the trophozoite stages only. This finding adds to our understanding of how patient factors can influence the outcome of Plasmodium falciparum infections.

Characterization of a recombinant porcine follistatin in a heat shock expression system

2002 ◽  
Vol 82 (3) ◽  
pp. 295-304
Author(s):  
C. R. Christensen ◽  
J. Kowalski ◽  
P. J. Chedrese ◽  
V. Misra ◽  
B. Laarveld ◽  
...  
Keyword(s):  
Heat Shock ◽  
Ovarian Function ◽  
Expression System ◽  
Porcine Granulosa Cells ◽  
Molecular Masses ◽  
Estradiol 17Β

The role of follistatin in ovarian function has yet to be fully elucidated; it is present in low concentration in vivo and it is difficult to obtain suitable quantities of follistatin for characterization. We have cloned porcine follistatin cDNA in an expression system that uses the heat shock protein promoter BoHSP70. Recombinant follistatin with apparent molecular masses of 39, 46, 48, 50 kDa was expressed and secreted into culture medium at concentrations of 400–500 μg × 20 mL-1 × 150 cm-2 flask (4 × 107 cells). In ligand blots, the recombinant follistatin was shown to be immunologically similar to native follistatin and to bind to recombinant activin A. Porcine granulosa cells dissected from 1–3 mm follicles from immature gilts were cultured with recombinant follistatin or with a follistatin monoclonal antibody to examine the activity of the recombinant follistatin. At a physiologically relevant concentration of 1 μg mL-1 the recombinant porcine follistatin suppressed the accumulation of estradiol-17β (P < 0.05). There was a trend for estradiol-17β accumulation in the presence of 10 μg mL-1 of monoclonal anti-follistatin antibody (P = 0.1). This expression system consistently produced large quantities of recombinant porcine follistatin, which is immunologically and biologically similar to follistatin, and is capable of independently inhibiting estratiol-17βproduction in vitro. Key words: Porcine, follistatin, heat shock promoter, glycoprotein, ovary, estradiol

scholarly journals The GARDpotency Assay for Potency-Associated Subclassification of Chemical Skin Sensitizers—Rationale, Method Development, and Ring Trial Results of Predictive Performance and Reproducibility

2020 ◽  
Vol 176 (2) ◽  
pp. 423-432
Author(s):  
Robin Gradin ◽  
Angelica Johansson ◽  
Andy Forreryd ◽  
Emil Aaltonen ◽  
Anders Jerre ◽  
...  
Keyword(s):  
Gene Expression ◽  
Method Development ◽  
Expression Profiles ◽  
Hazard Identification ◽  
Predictive Toxicology ◽  
Ring Trial ◽  
Biomarker Signature

Abstract Proactive identification and characterization of hazards attributable to chemicals are central aspects of risk assessments. Current legislations and trends in predictive toxicology advocate a transition from in vivo methods to nonanimal alternatives. For skin sensitization assessment, several OECD validated alternatives exist for hazard identification, but nonanimal methods capable of accurately characterizing the risks associated with sensitizing potency are still lacking. The GARD (Genomic Allergen Rapid Detection) platform utilizes exposure-induced gene expression profiles of a dendritic-like cell line in combination with machine learning to provide hazard classifications for different immunotoxicity endpoints. Recently, a novel genomic biomarker signature displaying promising potency-associated discrimination between weak and strong skin sensitizers was proposed. Here, we present the adaptation of the defined biomarker signature on a gene expression analysis platform suited for routine acquisition, confirm the validity of the proposed biomarkers, and define the GARDpotency assay for prediction of skin sensitizer potency. The performance of GARDpotency was validated in a blinded ring trial, in accordance with OECD guidance documents. The cumulative accuracy was estimated to 88.0% across 3 laboratories and 9 independent experiments. The within-laboratory reproducibility measures ranged between 62.5% and 88.9%, and the between-laboratory reproducibility was estimated to 61.1%. Currently, no direct or systematic cause for the observed inconsistencies between the laboratories has been identified. Further investigations into the sources of introduced variability will potentially allow for increased reproducibility. In conclusion, the in vitro GARDpotency assay constitutes a step forward for development of nonanimal alternatives for hazard characterization of skin sensitizers.

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