HSP70 positively regulates translation by interacting with the IRES and stabilizes the viral structural proteins VP1 and VP3 to facilitate duck hepatitis A virus type 1 replication.

The maintenance of viral protein homeostasis depends on the interaction between host cell proteins and viral proteins. As a molecular chaperone, heat shock protein 70 (HSP70) has been shown to play an important role in viral infection. Our results showed that HSP70 can affect translation, replication, assembly, and release during the life cycle of duck hepatitis A virus type 1 (DHAV-1). We demonstrated that HSP70 can regulate viral translation by interacting with the DHAV-1 internal ribosome entry site (IRES). In addition, HSP70 interacts with the viral capsid proteins VP1 and VP3 and promotes their stability by inhibiting proteasomal degradation, thereby facilitating the assembly of DHAV-1 virions. This study demonstrates the specific role of HSP70 in regulating DHAV-1 replication, which are helpful for understanding the pathogenesis of DHAV-1 infection and provide additional information about the role of HSP70 in infection by different kinds of picornaviruses, as well as the interaction between picornaviruses and host cells.

Acclimation to higher temperature and antioxidant supplemented diets improved rainbow trout (Oncorhynchus mykiss) resilience to heatwaves.

Coldwater species are challenged with increasing water temperatures and fluctuations over their upper thermal limits. This study evaluated the potential of acclimation to higher temperature and dietary antioxidants capacity to mitigate the adverse effects of heat shocks in rainbow trout. To this end, rainbow trout fingerlings were acclimated at optimal (14 °C) and high (20 °C) temperatures and fed on selenium (5 mg/kg) and polyphenol (2 g/kg) supplemented diets for 60 days and then were exposed to heat shocks by increasing water temperature up to 30 °C. Growth performance, survival rate, haemato-immunological parameters, and expression of HSP70α, HSP70ß, HSP90ß, and IL-1ß genes were measured to evaluate the hypothesises. The rainbow trout acclimated to 20 °C and fed on antioxidants supplemented diets showed a significantly higher aftershock survival rate. Moreover, fish acclimated to higher temperature showed higher red blood cell counts as well as serum total protein and albumin during the acclimation trial and heat shocks phase. Acclimation to higher temperature and feeding on antioxidants remarkably enhanced fish immune and antioxidant capacity in comparison to fish adapted to cold water and fed on the basal diet measured by improved respiratory burst and lysozyme activities and upregulation of IL-1ß expression during exposure of fish to heat shocks. Furthermore, fish acclimated to higher temperature, especially those fed on antioxidant supplemented diets, showed lower expression levels of HSPs genes during the heat shock phase, indicating that high heat shocks were less stressful for these fish in comparison to cold water acclimated fish. This finding was also supported by lower cortisol levels during heat shocks in fish acclimated to higher temperature. In conclusion, the results of this study indicated that acclimation to higher temperature and/or fed on diets supplemented by selenium and polyphenol, can help to mitigate the adverse effects of the heat shock in rainbow trout.

The NF-κB/FXR/TonEBP pathway protects renal medullary interstitial cells against hypertonic stress.

Farnesoid X receptor (FXR), a ligand-activated transcription factor, plays an important role in maintaining water homeostasis by up-regulating aquaporin 2 (AQP2) expression in renal medullary collecting ducts; however, its role in the survival of renal medullary interstitial cells (RMICs) under hypertonic conditions remains unclear. We cultured primary mouse RMICs and found that the FXR was expressed constitutively in RMICs, and that its expression was significantly up-regulated at both mRNA and protein levels by hypertonic stress. Using luciferase and ChIP assays, we found a potential binding site of nuclear factor kappa-B (NF-κB) located in the FXR gene promoter which can be bound and activated by NF-κB. Moreover, hypertonic stress-induced cell death in RMICs was significantly attenuated by FXR activation but worsened by FXR inhibition. Furthermore, FXR increased the expression and nuclear translocation of hypertonicity-induced tonicity-responsive enhance-binding protein (TonEBP), the expressions of its downstream target gene sodium myo-inositol transporter (SMIT), and heat shock protein 70 (HSP70). The present study demonstrates that the NF-κB/FXR/TonEBP pathway protects RMICs against hypertonic stress.

Capsaicin combined with stem cells improved mitochondrial dysfunction in PIG3V cells, an immortalized human vitiligo melanocyte cell line, by inhibiting the HSP70/TLR4/mTOR/FAK signaling axis.

BACKGROUND: Vitiligo is a common autoimmune skin disease. Capsaicin has been found to exert a positive effect on vitiligo treatment, and mesenchymal stem cells (MSCs) are also confirmed to be an ideal cell type. This study aimed to explore the influence of capsaicin combined with stem cells on the treatment of vitiligo and to confirm the molecular mechanism of capsaicin combined with stem cells in treating vitiligo. METHODS AND RESULTS: PIG3V cell proliferation and apoptosis were detected using CCK-8 and TUNEL assays, MitoSOX Red fluorescence staining was used to measure the mitochondrial ROS level, and JC-1 staining was used to detect the mitochondrial membrane potential. The expression of related genes and proteins was detected using RT‒qPCR and Western blotting. Coimmunoprecipitation was used to analyze the protein interactions between HSP70 and TLR4 or between TLR4 and mTOR. The results showed higher expression of HSP70 in PIG3V cells than in PIG1 cells. The overexpression of HSP70 reduced the proliferation of PIG3V cells, promoted apoptosis, and aggravated mitochondrial dysfunction and autophagy abnormalities. The expression of HSP70 could be inhibited by capsaicin combined with MSCs, which increased the levels of Tyr, Tyrp1 and DCT, promoted the proliferation of PIG3V cells, inhibited apoptosis, activated autophagy, and improved mitochondrial dysfunction. In addition, capsaicin combined with MSCs regulated the expression of TLR4 through HSP70 and subsequently affected the mTOR/FAK signaling pathway CONCLUSIONS: Capsaicin combined with MSCs inhibits TLR4 through HSP70, and the mTOR/FAK signaling pathway is inhibited to alleviate mitochondrial dysfunction and autophagy abnormalities in PIG3V cells.

Induction of heat shock protein expression in SP2/0 transgenic cells and its effect on the production of monoclonal antibodies.

The objective of the current investigation was to evaluate the induction of heat shock proteins (HSPs) in SP2/0 transgenic cells and the effect of these proteins on the production of monoclonal antibodies (mAbs). The SP2/0 cell line expressing the PSG-026 antibody, a biosimilar candidate of golimumab, the culture parameters, and the target protein expression were not justified for industrial production and were used for the experiments. Paracetamol and heat shock were used as chemical and physical inducers of HSPs, respectively. The results showed that paracetamol and heat shock increased the expression of HSP70 and HSP27 at the mRNA and protein levels. The expression of HSPs was greater in paracetamol-treated cells than in heat shock-treated cells. Paracetamol treatment at concentrations above 0.5 mM significantly reduced cell viability and mAb expression. However, treatment with 0.25 mM paracetamol results in delayed cell death and increased mAb production. Heat shock treatment at 45°C for 30 minutes after enhanced mAb expression was applied after pre-treatment with paracetamol. In bioreactor cultures, pretreatment of cells with paracetamol improved cell viability and shortened the lag phase, resulting in increased cell density. The production of mAbs in paracetamol-treated cultures was markedly greater than that in the control. Analysis of protein quality and charge variants revealed no significant differences between paracetamol-treated and control cultures, indicating that the induction of HSPs did not affect protein aggregation or charge variants. These findings suggest that inducing and manipulating HSP expression can be a valuable strategy for improving recombinant protein production in biopharmaceutical processes.

Heat acclimation improves exercise performance in hot conditions and increases heat shock protein 70 and 90 of skeletal muscles in Thoroughbred horses.

This study aimed to determine whether heat acclimation could induce adaptations in exercise performance, thermoregulation, and the expression of proteins associated with heat stress in the skeletal muscles of Thoroughbreds. Thirteen trained Thoroughbreds performed 3 weeks of training protocols, consisting of cantering at 90% maximal oxygen consumption (VO2max) for 2 min 2 days/week and cantering at 7 m/s for 3 min 1 day/week, followed by a 20-min walk in either a control group (CON; Wet Bulb Globe Temperature [WBGT] 12-13°C; n = 6) or a heat acclimation group (HA; WBGT 29-30°C; n = 7). Before and after heat acclimation, standardized exercise tests (SET) were conducted, cantering at 7 m/s for 90 s and at 115% VO2max until fatigue in hot conditions. Increases in run time (p = 0.0301), peak cardiac output (p = 0.0248), and peak stroke volume (p = 0.0113) were greater in HA than in CON. Pulmonary artery temperature at 7 m/s was lower in HA than in CON (p = 0.0332). The expression of heat shock protein 70 (p = 0.0201) and 90 (p = 0.0167) increased in HA, but not in CON. These results suggest that heat acclimation elicits improvements in exercise performance and thermoregulation under hot conditions, with a protective adaptation to heat stress in equine skeletal muscles.

Extracellular Vesicles from Ecklonia cava and Phlorotannin Promote Rejuvenation in Aged Skin.

Plant-derived extracellular vesicles (EVs) elicit diverse biological effects, including promoting skin health. EVs isolated from Ecklonia cava (EV-EC) carry heat shock protein 70 (HSP70), which inhibits key regulators such as TNF-α, MAPKs, and NF-κB, consequently downregulating matrix metalloproteinases (MMPs). Aging exacerbates oxidative stress, upregulating MAPK and NF-κB signaling and worsening extracellular matrix degradation in the skin. E. cava-derived phlorotannin (PT) mitigates MAPK and NF-κB signaling. We evaluated the impact of EV-EC and PT on skin rejuvenation using an in vitro keratinocyte senescence model and an in vivo aged-mouse model. Western blotting confirmed the presence of HSP70 in EV-EC. Treatment with EV-EC and PT in senescent keratinocytes increased HSP70 expression and decreased the expression of TNF-α, MAPK, NF-κB, activator protein-1 (AP-1), and MMPs. Oxidative stress was also reduced. Sequential treatment with PT and EV-EC (PT/EV-EC) yielded more significant results compared to individual treatments. The administration of PT/EV-EC to the back skin of aged mice mirrored the in vitro findings, resulting in increased collagen fiber accumulation and improved elasticity in the aged skin. Therefore, PT/EV-EC holds promise in promoting skin rejuvenation by increasing HSP70 expression, decreasing the expression of MMPs, and reducing oxidative stress in aged skin.

Performance, HSP70 expression, antioxidant enzymes, oxidative DNA damage biomarkers, metabolic hormones, and intestinal microbiota of broiler chickens fed mistletoe leaf powder supplemented diets under tropical high ambient temperatures.

The study investigated the impact of Mistletoe Leaf Powder (MLP) supplementation on some parameters in heat-stressed broiler chickens. The standard baseline diets, comprising four different formulations, were provided during the starter and finisher stages. Chickens were randomly assigned to the 4 dietary groups: a negative control (CON) with no supplementation, a positive control (VTC) with 200 mg/kg vitamin C, and 2 experimental treatment groups with 2500 mg/kg (MLP2) and 5000 mg/kg (MLP5) MLP supplementation. The Body Weight Gain (BWG) in MLP2 and MLP5 treatment groups was comparable (P > 0.05) to those in VTC, while the CON group exhibited significantly (P < 0.05) lower BWG. Feed consumption was significantly (P < 0.05) lower broiler chickens in the CON group compared to those VTC, MLP2, and MLP5. Heat shock protein 70 (HSP70) levels were lower in broiler chickens belonging to VTC, MLP2, and MLP5 groups compared to those in CON, and MLP2 showed no difference (P > 0.05) from MLP5 and VTC. Serum glutathione peroxidase and catalase concentrations were higher (P < 0.05) in birds belonging to MLP5, MLP2, and VTC groups compared to CON. The 8-hydroxy-2'-deoxyguanosine concentration was lower (P < 0.05) in birds of VTC, MLP2, and MLP5 compared to the CON, with VTC showing the least concentration. Serum insulin levels were higher (P < 0.05) in MLP5 compared to those in CON, while serum triiodothyronine and leptin concentrations were lower (P < 0.05) in CON compared to birds in VTC, MLP2, and MLP5. Microbiota analysis revealed that the Coliform bacteria population was higher (P < 0.05) in birds belonging to CON compared to those in VTC, MLP2, and MLP5 groups, whereas lactic acid-producing bacteria were significantly (P < 0.05) lower in birds of CON and highest in MLP2 and MLP5 groups. In conclusion, dietary supplementation of MLP at 5000 mg/kg enhanced performance, oxidative status, influenced metabolic hormones, and gut microbiota in broiler chickens raised under high ambient temperature.

Phytomediated stress modulates antioxidant status, induces overexpression of CYP6M2, Hsp70, α-esterase, and suppresses the ABC transporter in Anopheles gambiae (sensu stricto) exposed to Ocimum tenuiflorum extracts.

The incorporation of phytoactive compounds in the management of malarial vectors holds promise for the development of innovative and efficient alternatives. Nevertheless, the molecular and physiological responses that these bioactive substances induce remain underexplored. This present study investigated the toxicity of different concentrations of aqueous and methanol extracts of Ocimum tenuiflorum against larvae of Anopheles gambiae (sensu stricto) and unraveled the possible underlying molecular pathways responsible for the observed physiological effects. FTIR and GCMS analyses of phytoactive compounds in aqueous and methanol crude extracts of O. tenuiflorum showed the presence of OH stretching vibration, C = C stretching modes of aromatics and methylene rocking vibration; ring deformation mode with high levels of trans-ß-ocimene, 3,7-dimethyl-1,3,6-octatriene in aqueous extract and 4-methoxy-benzaldehyde, 1,3,5-trimethyl-cyclohexane and o-cymene in methanol extract. The percentage mortality upon exposure to methanol and aqueous extracts of O. tenuiflorum were 21.1% and 26.1% at 24 h, 27.8% and 36.1% at 48 h and 36.1% and 45% at 72 h respectively. Using reverse transcription quantitative polymerase chain reaction (RT-qPCR), down-regulation of ABC transporter, overexpression of CYP6M2, Hsp70, and α-esterase, coupled with significantly increased levels of SOD, CAT, and GSH, were observed in An. gambiae (s.s.) exposed to aqueous and methanol extracts of O. tenuiflorum as compared to the control. Findings from this study have significant implications for our understanding of how An. gambiae (s.s.) larvae detoxify phytoactive compounds.

A native mass spectrometry approach to qualitatively elucidate interfacial epitopes of transient protein-protein interactions.

Native mass spectrometric analysis of TPR2A and GrpE with unpurified peptides derived from limited proteolysis of their respective PPI partners (HSP90 C-terminus and DnaK) facilitated efficient, qualitative identification of interfacial epitopes involved in transient PPI formation. Application of this approach can assist in elucidating interfaces of currently uncharacterised transient PPIs.

Thermotolerance in S. cerevisiae as a model to study extracellular vesicle biology.

The budding yeast Saccharomyces cerevisiae is a proven model organism for elucidating conserved eukaryotic biology, but to date its extracellular vesicle (EV) biology is understudied. Here, we show yeast transmit information through the extracellular medium that increases survival when confronted with heat stress and demonstrate the EV-enriched samples mediate this thermotolerance transfer. These samples contain vesicle-like particles that are exosome-sized and disrupting exosome biogenesis by targeting endosomal sorting complexes required for transport (ESCRT) machinery inhibits thermotolerance transfer. We find that Bro1, the yeast ortholog of the human exosome biomarker ALIX, is present in EV samples, and use Bro1 tagged with green fluorescent protein (GFP) to track EV release and uptake by endocytosis. Proteomics analysis reveals that heat shock protein 70 (HSP70) family proteins are enriched in EV samples that provide thermotolerance. We confirm the presence of the HSP70 ortholog stress-seventy subunit A2 (Ssa2) in EV samples and find that mutant yeast cells lacking SSA2 produce EVs but they fail to transfer thermotolerance. We conclude that Ssa2 within exosomes shared between yeast cells contributes to thermotolerance. Through this work, we advance Saccharomyces cerevisiae as an emerging model organism for elucidating molecular details of eukaryotic EV biology and establish a role for exosomes in heat stress and proteostasis that seems to be evolutionarily conserved.

Circulating mortalin in blood and activation of the alternative complement pathway as risk indicators in COVID-19 infection.

Background: Mortalin/GRP75 is a ubiquitous mitochondrial chaperone related to the cytosolic heat shock protein 70. It protects cells from various types of damages and from senescence. Our goal was to determine whether COVID-19 patients have circulating mortalin in their blood and to assess its prognostic value in anticipating disease severity. Methods: Mortalin was determined by ELISA in the sera of 83 COVID-19 patients enrolled in the study. Patients were categorized into 4 groups: critical patients who died (FATAL) or required intensive care and survived (ICU), patients of mild severity (hospitalized but not critical) who required nasal oxygen support (HOSP+O2), and patients who did not need oxygen therapy (HOSP). Results: The mortalin concentration in the serum of all COVID-19 patients in the cohort was 194-2324 pg/mL. A comparison of the mortalin levels by peak severity among the various patient groups showed a highly significant difference between the HOSP and FATAL groups and a significant difference between the HOSP and the ICU groups. COVID-19 patients who eventually failed to survive had at hospitalization a markedly higher level of mortalin in their sera. Cox regression analysis revealed a high mortality hazard (HR=3.96, p<0.01) in patients with high mortalin circulating levels (above the median, ≥651 pg/mL). This was confirmed in survival curve analysis (Kaplan-Meier; p=0.0032, log-rank test). Mortalin remained an independent predictor of mortality even after adjusting for age and sex or various complement activation products. Complement activation data collected in an earlier study in the same cohort was compared regarding the mortalin levels. Patients with higher circulating mortalin levels also had higher levels of complement C3a but reduced levels of properdin. Discussion: This is the first report on circulating mortalin in COVID-19 patients. Higher mortalin levels were associated with more severe illnesses and a higher risk of death. We claim that quantifying the blood levels of mortalin and activated complement proteins will provide important information on the prognosis of COVID-19 patients and will serve as a useful tool for guiding their clinical management and treatment.

Morphological and molecular characteristics of a Trypanosoma sp. from triatomines (Triatoma rubrofasciata) in China.

BACKGROUND: Triatomines (kissing bugs) are natural vectors of trypanosomes, which are single-celled parasitic protozoans, such as Trypanosoma cruzi, T. conorhini and T. rangeli. The understanding of the transmission cycle of T. conorhini and Triatoma rubrofasciata in China is not fully known. METHODS: The parasites in the faeces and intestinal contents of the Tr. rubrofasciata were collected, and morphology indices were measured under a microscope to determine the species. DNA was extracted from the samples, and fragments of 18S rRNA, heat shock protein 70 (HSP70) and glycosomal glyceraldehyde-3-phosphate dehydrogenase (gGAPDH) were amplified and sequenced. The obtained sequences were then identified using the BLAST search engine, followed by several phylogenetic analyses. Finally, laboratory infections were conducted to test whether Tr. rubrofasciata transmit the parasite to rats (or mice) through bites. Moreover, 135 Tr. rubrofasciata samples were collected from the Guangxi region and were used in assays to investigate the prevalence of trypanosome infection. RESULTS: Trypanosoma sp. were found in the faeces and intestinal contents of Tr. rubrofasciata, which were collected in the Guangxi region of southern China and mostly exhibited characteristics typical of epimastigotes, such as the presence of a nucleus, a free flagellum and a kinetoplast. The body length ranged from 6.3 to 33.9 µm, the flagellum length ranged from 8.7 to 29.8 µm, the nucleus index was 0.6 and the kinetoplast length was -4.6. BLAST analysis revealed that the 18S rRNA, HSP70 and gGAPDH sequences of Trypanosoma sp. exhibited the highest degree of similarity with those of T. conorhini (99.7%, 99.0% and 99.0%, respectively) and formed a well-supported clade close to T. conorhini and T. vespertilionis but were distinct from those of T. rangeli and T. cruzi. Laboratory experiments revealed that both rats and mice developed low parasitaemia after inoculation with Trypanosoma sp. and laboratory-fed Tr. rubrofasciata became infected after feeding on trypanosome-positive rats and mice. However, the infected Tr. rubrofasciata did not transmit Trypanosoma sp. to their offspring. Moreover, our investigation revealed a high prevalence of Trypanosoma sp. infection in Tr. rubrofasciata, with up to 36.3% of specimens tested in the field being infected. CONCLUSIONS: Our study is the first to provide a solid record of T. conorhini from Tr. rubrofasciata in China with morphological and molecular evidence. This Chinese T. conorhini is unlikely to have spread through transovarial transmission in Tr. rubrofasciata, but instead, it is more likely that the parasite is transmitted between Tr. rubrofasciata and mice (or rats). However, there was a high prevalence of T. conorhini in the Tr. rubrofasciata from our collection sites and numerous human cases of Tr. rubrofasciata bites were recorded. Moreover, whether these T. conorhini strains are pathogenic to humans has not been investigated.

[Identification of HSP70 gene family members in Fritillaria cirrhosa and expression analysis in different tissues under high temperature stress].

The heat shock protein 70 family contains the stress proteins ubiquitous in plants. These proteins are involved in the responses to different abiotic stress conditions and have highly conserved gene sequences. However, little is known about the molecular mechanisms of Fritillaria cirrhosa in response to high-temperature stress. Here, 26 HSP70s, FcHSP70-1 to FcHSP70-26, were identified from the transcriptome data of root, bulb, stem, leaf, and fruit samples of F. cirrhosa. The proteins encoded by FcHSP70s had the lengths ranging from 560 aa to 944 aa, with the molecular weight of 61.64-100.01 kDa and the theoretical isoelectric point between 5.00 and 6.59. The secondary structural elements of HSP70s were mainly random coils and α-helixes. Subcellular localization prediction revealed that FcHSP70s were distributed in mitochondria, chloroplasts, nuclei, endoplasmic reticulum, and cytoplasm. The phylogenetic tree showed that 7 members of the HSP70 family belonged to the Dnak subfamily and 19 members belonged to the HSP110/SSE subfamily. In addition, the qRT-PCR results showed that the expression of FcHSP70-5, FcHSP70-8, FcHSP70-17, FcHSP70-18, and FcHSP70-23 in F. cirrhosa was significantly up-regulated at 35 ℃, which indicated that these genes might play a role in the response to high temperature stress. In addition, compared with other tissues, stems and leaves were sensitive to high temperature stress, with the expression of 18 genes up-regulated by 18.18 and 8.03 folds on average, respectively. These findings provide valuable information about the molecular mechanism of HSP70s of F. cirrhosa in response to high temperature stress.

Synergistic Effect of Dietary Supplementation with Sodium Butyrate, ß-Glucan and Vitamins on Growth Performance, Cortisol Level, Intestinal Microbiome and Expression of Immune-Related Genes in Juvenile African Catfish (Clarias gariepinus).

The effect of dietary supplementation with sodium butyrate, ß-glucan and vitamins (A, D3, E, K, C) on breeding indicators and immune parameters of juvenile African catfish was examined. The fish were fed with unenriched (group C) and enriched feed with a variable proportion of sodium butyrate/ß-glucan, and constant content of vitamins (W1-W3). After the experiment, blood and the middle gut were collected. The microbiome of the gut was determined using Next Generation Sequencing (NGS). Liver tissue was collected for determination of expression of immune-related genes (HSP70, IL-1ß, TNFα). W2 and W3 were characterized by the most favorable values of breeding indicators (p < 0.05). The highest blood cortisol concentration was in group C (71.25 ± 10.45 ng/mL), and significantly the lowest in W1 (46.03 ± 7.01 ng/ mL) (p < 0.05). The dominance of Cetobacterium was observed in all study groups, with the largest share in W3 (65.25%) and W1 (61.44%). Gene expression showed an increased number of HSP70 genes in W1. IL-1ß and TNFα genes peaked at W3. The W3 variant turns out to be the most beneficial supplementation, due to the improvement of breeding and immunological parameters. The data obtained can be used to create a preparation for commercial use in the breeding of this species.

The Listeria monocytogenes persistence factor ClpL is a potent stand-alone disaggregase.

Heat stress can cause cell death by triggering the aggregation of essential proteins. In bacteria, aggregated proteins are rescued by the canonical Hsp70/AAA+ (ClpB) bi-chaperone disaggregase. Man-made, severe stress conditions applied during, e.g., food processing represent a novel threat for bacteria by exceeding the capacity of the Hsp70/ClpB system. Here, we report on the potent autonomous AAA+ disaggregase ClpL from Listeria monocytogenes that provides enhanced heat resistance to the food-borne pathogen enabling persistence in adverse environments. ClpL shows increased thermal stability and enhanced disaggregation power compared to Hsp70/ClpB, enabling it to withstand severe heat stress and to solubilize tight aggregates. ClpL binds to protein aggregates via aromatic residues present in its N-terminal domain (NTD) that adopts a partially folded and dynamic conformation. Target specificity is achieved by simultaneous interactions of multiple NTDs with the aggregate surface. ClpL shows remarkable structural plasticity by forming diverse higher assembly states through interacting ClpL rings. NTDs become largely sequestered upon ClpL ring interactions. Stabilizing ring assemblies by engineered disulfide bonds strongly reduces disaggregation activity, suggesting that they represent storage states.

Challenges for heat stress: Intestinal culturable bacteria of Lohmann Brown chickens.

This study aimed to assess how heat stress, specifically within the range of 35-38 °C, affects the populations of culturable intestinal lactobacilli, enterococci, and Escherichia coli, as well as the expression of Heat Shock Proteins (HSP70), in Lohmann Brown chickens. It also explored the influence of the chickens' blood transferrin and ceruloplasmin genotypes on these responses. Thirty chickens underwent eight hours of heat stress, maintained at an average temperature of 37 °C and a relative humidity of 75-80%, with continuous access to food and water. Behavioral monitoring was conducted throughout to prevent excessive heat-related mortality. The Lohmann Brown chickens from the Yerevan "Arax" poultry farm were initially classified based on their blood transferrin and ceruloplasmin genotypes to investigate potential correlations between intestinal bacterial composition and variations in these polymorphisms. A significant correlation was found between heat stress and the abundance of culturable enterococci within the intestinal microbiota, regardless of chicken TfAB, TfBC, CpAB, CpCC and TfAB, TfBC, CpAB, CpCD genotypes. Heat stress led to nearly double the HSP70 levels in chicken blood, along with a reduction in the culturable enterococci population by at least 10,000-fold in the intestinal microbiota. These findings are significant for targeted management strategies to mitigate heat stress in chicken populations.

Tertiary structure and conformational dynamics of the anti-amyloidogenic chaperone DNAJB6b at atomistic resolution.

DNAJB6b is a molecular chaperone of the heat shock protein network, shown to play a crucial role in preventing aggregation of several disease-related intrinsically disordered proteins. Using homology modeling and microsecond-long all-atom molecular dynamics (MD) simulations, we show that monomeric DNAJB6b is a transiently interconverting protein cycling between three states: a closed state, an open state (both abundant), and a less abundant extended state. Interestingly, the reported regulatory autoinhibitory anchor between helix V in the G/F1 region and helices II/III of the J-domain, which obstructs the access of Hsp70 to the J-domain remains present in all three states. This possibly suggests a mechanistically intriguing regulation in which DNAJB6b only becomes exposed when loaded with substrates that require Hsp70 processing. Our MD results of DNAJB6b carrying mutations in the G/F1 region that are linked to limb-girdle muscular dystrophy type D1 (LGMDD1) show that this G/F1 region becomes highly dynamic, pointing towards a spontaneous release of the autoinhibitory helix V from helices II/III. This would increase the probability of non-functional Hsp70 interactions to DNAJB6b without substrates. Our cellular data indeed confirm that non-substrate loaded LGMDD1 mutants have aberrant interactions with Hsp70.

[Identification and expression analysis of the HSP70 gene family under abiotic stresses in Litchi chinensis].

HSP70 protein, as an important member of the heat shock protein (HSP) family, plays an important role in plant growth, development, and response to biotic and abiotic stresses. In order to explore the role of HSP70 gene family members in Litchi chinensis under low temperature, high temperature, drought, and salt stress, bioinformatics methods were used to identify the HSP70 gene family members within the entire L. chinensis genome. The expression of these genes under various abiotic stresses was then detected using quantitative real-time PCR (qRT-PCR). The results showed that the LcHSP70 gene family consisted of 18 members, which were unevenly distributed across ten L. chinensis chromosomes. The LcHSP70 protein contained 479-851 amino acids, with isoelectric points ranging from 5.07 to 6.95, and molecular weights from 52.44 kDa to 94.07 kDa. The predicted subcellular localization showed that LcHSP70 protein was present in the nucleus, cytoplasm, endoplasmic reticulum, mitochondria, and chloroplast. Phylogenetic analysis divided the LcHSP70 proteins into five subgroups, namely Ⅰ, Ⅱ, Ⅲ, Ⅳ, and Ⅵ. The promoter regions of the LcHSP70 genes contained various cis-acting elements related to plant growth, development, hormone response, and stress response. Moreover, the expression of LcHSP70 genes displayed distint tissue-specific expression level, categorized into universal expression and specific expression. From the selected 6 LcHSP70 genes (i.e., LcHSP70-1, LcHSP70-5, LcHSP70-10, LcHSP70-14, LcHSP70-16, and LcHSP70-18), their relative expression levels were assessed under different abiotic stresses using qRT-PCR. The results indicated that the gene family members exhibited diverse responses to low temperature, high temperature, drought, and salt stress, with significant variations in their expression levels across different time periods. These results provide a foundation for further exploration of the function of the LcHSP70 gene family.

Chaperone Hsp70 helps Salmonella survive infection-relevant stress by reducing protein synthesis.

In all domains of life, Hsp70 chaperones preserve protein homeostasis by promoting protein folding and degradation and preventing protein aggregation. We now report that the Hsp70 from the bacterial pathogen Salmonella enterica serovar Typhimurium-termed DnaK-independently reduces protein synthesis in vitro and in S. Typhimurium facing cytoplasmic Mg2+ starvation, a condition encountered during infection. This reduction reflects a 3-fold increase in ribosome association with DnaK and a 30-fold decrease in ribosome association with trigger factor, the chaperone normally associated with translating ribosomes. Surprisingly, this reduction does not involve J-domain cochaperones, unlike previously known functions of DnaK. Removing the 74 C-terminal amino acids of the 638-residue long DnaK impeded DnaK association with ribosomes and reduction of protein synthesis, rendering S. Typhimurium defective in protein homeostasis during cytoplasmic Mg2+ starvation. DnaK-dependent reduction in protein synthesis is critical for survival against Mg2+ starvation because inhibiting protein synthesis in a dnaK-independent manner overcame the 10,000-fold loss in viability resulting from DnaK truncation. Our results indicate that DnaK protects bacteria from infection-relevant stresses by coordinating protein synthesis with protein folding capacity.