The DegU Orphan Response Regulator Contributes to Heat Stress Resistance in Listeria monocytogenes

Listeria monocytogenes is more heat-resistant than most other non-spore-forming foodborne pathogens, posing a severe threat to food safety and human health, particularly during chilled food processing. The DegU orphan response regulator is known to control heat resistance in L. monocytogenes; however, the underlying regulatory mechanism is poorly understood. Here, we show that DegU contributes to L. monocytogenes exponential growth under mild heat-shock stress. We further demonstrate that DegU directly senses heat stress through autoregulation and upregulates the hrcA-grpE-dnaK-dnaJ operon, leading to increased production of heat-shock proteins. We also show that DegU can directly regulate the expression of the hrcA-grpE-dnaK-dnaJ operon. In conclusion, our results shed light on the regulatory mechanisms underlying how DegU directly activates the hrcA-grpE-dnaK-dnaJ operon, thereby regulating heat resistance in L. monocytogenes.

The CRZ-1 Transcription Factor Regulates Hsp 80 Involves in the Acquisition of Thermotolerance, and NCA-2 for Calcium Stress Tolerance in Neurospora Crassa

Heat shock proteins (Hsps) are molecular chaperones and required for survival of organisms under heat stress conditions. In this study, we studied Hsp80, a member of the Hsp90 family, in Neurospora crassa. The expression of hsp80 was severely reduced in the N. crassa calcineurin B subunit RIP-mutant (cnb-1RIP) strains under the heat shock conditions. Furthermore, the expression levels of cnb-1, hsp60, hsp80, and the calcineurin-regulated transcription factor crz-1 were increased, but expression levels were reduced in the presence of the calcineurin inhibitor FK506 under the heat shock stress in the N. crassa wild type. Therefore, the calcineurin-crz-1 signaling pathway transcriptionally regulates hsp60 and hsp80 under the heat shock stress condition in N. crassa. In addition, the transcript levels of trm-9 and nca-2, a Ca2+ sensor and a Ca2+ ATPase, respectively, were increased under the heat shock stress condition. Moreover, the expression of the hsp80, but not the hsp60, was reduced in the Δtrm-9, Δnca-2, and the Δtrm-9 Δnca-2 double mutants. These results suggested that hsp80, trm-9, and nca-2 play a role in coping the heat shock stress in N. crassa. We found that CRZ-1 binds to 5ʹ-CCTTCACA-3ʹ and 5ʹ-AGCGGAGC-3ʹ 8 bp nucleotide sequences, located about 1075 bp and 679 bp upstream of the ATG start codon, respectively, of hsp80. We also found that CRZ-1 binds to an 8 bp nucleotide sequence 5ʹ-ACCGCGCC-3ʹ, located 234 bp upstream of the ATG start codon of nca-2 under Ca2+ stress condition. Thus, cnb-1, hsp60, hsp80, and crz-1 are involved in the heat shock stress response in N. crassa. Moreover, CRZ-1 upregulates the expressions of hsp80 and nca-2 under the heat shock stress and Ca2+ stress conditions, respectively, in N. crassa.

EPIGENETIC REGULATION OF ADAPTOGENESIS BY PATHOLOGY AND AGING

В обзоре с точки зрения эпигенетики рассмотрены адаптационные возможности организма при патологии и старении. Апаптация организма к внутренним и внешним факторам осуществляется единой гуморальной защитной системой организма, включающей гипоталамо-гипофизарно-эпифизарную и гипоталамогипофизарно-тимусную оси. Короткие пептиды AEDG, AEDP, EDR, KED, EW, KE являются эпигенетическими регуляторами экспрессии генов и синтеза белков, которые могут быть вовлечены в адаптацию при стрессе и активацию гипоталамо-гипофизарно-эпифизарной и гипоталамогипофизарно-тимусной осей. Указанные короткие пептиды регулируют синтез белков теплового шока, стресспротекторных белков, цитокинов, факторов фибринолиза и гемостаза. Эти пептиды могут участвовать в первичной и отсроченной эпигенетической регуляции адаптивного ответа при стрессе, патологии и старении. Ранняя функциональная диагностика нарушения сопряжения звеньев единой гуморальной защитной системы организма при возраст-ассоциированных заболеваниях позволит выявить недостаточную синхронность эпигенетических механизмов, при которой наступает истощение и снижение резервных возможностей организма. Применение пептидов может нивелировать проявления адаптационного синдрома при стрессе и возрастной патологии. The organism adaptive possibilities by pathology and aging are discussed in account of the epigenetic. The organism adaptation to inner and external factors is carried out by organism unite humoral protective system, inclusive hypothalamus-hypophysis-pineal and hypothalamus-hypophysis-thymus axises. AEDG, AEDP, EDR, KED, EW, KE short peptides are the epigenetic regulators of gene expression and protein synthesis, which can be involve to the adaptation by stress and in the activation of hypothalamus-hypophysispineal and hypothalamus-hypophysis-thymus axises. These short peptides regulate the synthesis of proteins of heat shock, stress-protective proteins, cytocines, fibrinolysis and hemostasis factors and can participate in primary and tardive epigenetic regulation of adaptive response by stress, pathology and aging. The early functional diagnostic of element disturbances of organism unite humoral protective system by age-associative pathology can be usefull for the detection of deficient synchronization of epigenetic mechanisms, by wich the depletion and decrease of organism reserve possibilities occurs. The use of peptide can grade the adaptive syndrome manifestation by the stress and age pathology.

A heat-shock 20 protein isolated from watermelon (ClHSP22.8) negatively regulates the response of Arabidopsis to salt stress via multiple signaling pathways

Heat-shock protein 20s (HSP20) were initially shown to play a role during heat shock stress; however, recent data indicated that HSP20 proteins are also involved in abiotic stress in plants. Watermelon is known to be vulnerable to various stressors; however, HSP20 proteins have yet to be investigated and characterized in the watermelon. In a previous study, we identified a negative regulator of salt stress response from watermelon: ClHSP22.8, a member of the HSP20 family. Quantitative real-time PCR (qRT-PCR) and promoter::β-glucuronidase (GUS) analysis revealed that ClHSP22.8 was expressed widely in a range of different tissues from the watermelon, but particularly in the roots of 7-day-old seedlings and flowers. Furthermore, qRT-PCR and GUS staining showed that the expression of ClHSP22.8 was significantly repressed by exogenous abscisic acid (ABA) and salt stress. The over-expression of ClHSP22.8 in Arabidopsis lines resulted in hypersensitivity to ABA and reduced tolerance to salt stress. Furthermore, the expression patterns of key regulators associated with ABA-dependent and independent pathways, and other stress-responsive signaling pathways, were also repressed in transgenic lines that over-expressed ClHSP22.8. These results indicated that ClHSP22.8 is a negative regulator in plant response to salt stress and occurs via ABA-dependent and independent, and other stress-responsive signaling pathways.

Exogenous Salicylic Acid Modulates the Response to Combined Salinity-Temperature Stress in Pepper Plants (Capsicum annuum L. var. Tamarin)

Growers in the cultivated areas where the climate change threatens the agricultural productivity and livelihoods are aware that the current constraints for good quality water are being worsened by heatwaves. We studied the combination of salinity (60 mM NaCl) and heat shock stress (43 °C) in pepper plants (Capsicum annuum L. var. Tamarin) since this can affect physiological and biochemical processes distinctly when compared to separate effects. Moreover, the exogenous application of 0.5 mM salicylic acid (SA) was studied to determine its impacts and the SA-mediated processes that confer tolerance of the combined or stand-alone stresses. Plant growth, leaf Cl− and NO3− concentrations, carbohydrates, and polyamines were analyzed. Our results show that both salinity stress (SS) and heat stress (HS) reduced plant fresh weight, and SA only increased it for HS, with no effect for the combined stress (CS). While SA increased the concentration of Cl− for SS or CS, it had no effect on NO3−. The carbohydrates concentrations were, in general, increased by HS, and were decreased by CS, and for glucose and fructose, by SA. Additionally, when CS was imposed, SA significantly increased the spermine and spermidine concentrations. Thus, SA did not always alleviate the CS and the plant response to CS cannot be directly attributed to the full or partial sum of the individual responses to each stress.

Identification of novel heat shock-induced long non-coding RNA in human cells

The heat-shock response is a crucial system for survival of organisms under heat stress. During heat-shock stress, gene expression is globally suppressed, but expression of some genes, such as chaperone genes, is selectively promoted. These selectively activated genes have critical roles in the heat-shock response, so it is necessary to discover heat-inducible genes to reveal the overall heat-shock response picture. The expression profiling of heat-inducible protein-coding genes has been well-studied, but that of non-coding genes remains unclear in mammalian systems. Here, we used RNA-seq analysis of heat shock-treated A549 cells to identify seven novel long non-coding RNAs that responded to heat shock. We focussed on CTD-2377D24.6 RNA, which is most significantly induced by heat shock, and found that the promoter region of CTD-2377D24.6 contains the binding site for transcription factor HSF1 (heat shock factor 1), which plays a central role in the heat-shock response. We confirmed that HSF1 knockdown cancelled the induction of CTD-2377D24.6 RNA upon heat shock. These results suggest that CTD-2377D24.6 RNA is a novel heat shock-inducible transcript that is transcribed by HSF1.

Clarithromycin Exerts an Antibiofilm Effect against Salmonella enterica Serovar Typhimurium rdar Biofilm Formation and Transforms the Physiology towards an Apparent Oxygen-Depleted Energy and Carbon Metabolism

ABSTRACT Upon biofilm formation, production of extracellular matrix components and alteration in physiology and metabolism allows bacteria to build up multicellular communities which can facilitate nutrient acquisition during unfavorable conditions and provide protection toward various forms of environmental stresses to individual cells. Thus, bacterial cells within biofilms become tolerant against antimicrobials and the immune system. In the present study, we evaluated the antibiofilm activity of the macrolides clarithromycin and azithromycin. Clarithromycin showed antibiofilm activity against rdar (red, dry, and rough) biofilm formation of the gastrointestinal pathogen Salmonella enterica serovar Typhimurium ATCC 14028 (Nalr) at a 1.56 μM subinhibitory concentration in standing culture and dissolved cell aggregates at 15 μM in a microaerophilic environment, suggesting that the oxygen level affects the activity of the drug. Treatment with clarithromycin significantly decreased transcription and production of the rdar biofilm activator CsgD, with biofilm genes such as csgB and adrA to be concomitantly downregulated. Although fliA and other flagellar regulon genes were upregulated, apparent motility was downregulated. RNA sequencing showed a holistic cell response upon clarithromycin exposure, whereby not only genes involved in the biofilm-related regulatory pathways but also genes that likely contribute to intrinsic antimicrobial resistance, and the heat shock stress response were differentially regulated. Most significantly, clarithromycin exposure shifted the cells toward an apparent oxygen- and energy-depleted status, whereby the metabolism that channels into oxidative phosphorylation was downregulated, and energy gain by degradation of propane 1,2-diol, ethanolamine and l-arginine catabolism, potentially also to prevent cytosolic acidification, was upregulated. This analysis will allow the subsequent identification of novel intrinsic antimicrobial resistance determinants.

Clarithromycin Exerts an Antibiofilm Effect against Salmonella typhimurium rdar Biofilm Formation, and Transforms the Physiology towards an Apparent Oxygen-depleted Energy and Carbon Metabolism

ABSTRACTUpon biofilm formation, production of extracellular matrix components and alteration in physiology and metabolism allows bacteria to build up multicellular communities which can facilitate nutrient acquisition during unfavorable conditions and provide protection towards various forms of environmental stresses to individual cells. Thus, bacterial cells become tolerant against antimicrobials and the immune system within biofilms. In the current study, we evaluated the antibiofilm activity of the macrolides clarithromycin and azithromycin. Clarithromycin showed antibiofilm activity against rdar (red, dry and rough) biofilm formation of the gastrointestinal pathogen Salmonella typhimurium ATCC14028 Nalr at 1.56 μM subinhibitory concentration in standing culture and dissolved cell aggregates at 15 μM in a microaerophilic environment suggesting that the oxygen level affects the activity of the drug. Treatment with clarithromycin significantly decreased transcription and production of the rdar biofilm activator CsgD, with biofilm genes such as csgB and adrA to be consistently downregulated. While fliA and other flagellar regulon genes were upregulated, apparent motility was downregulated. RNA sequencing showed a holistic cell response upon clarithromycin exposure, whereby not only genes involved in the biofilm-related regulatory pathways, but also genes that likely contribute to intrinsic antimicrobial resistance, and the heat shock stress response were differentially regulated. Most significantly, clarithromycin exposure shifts the cells towards an apparent oxygen- and energy-depleted status, whereby the metabolism that channels into oxidative phosphorylation is downregulated, and energy gain by degradation of propane 1,2-diol, ethanolamine and L-arginine catabolism is upregulated. This initial analysis will allow the subsequent identification of novel intrinsic antimicrobial resistance determinants.