Experimental simulation of environmental warming selects against pigmented morphs of land snails

In terrestrial snails, thermal selection acts on shell coloration. However, the biological relevance of small differences in the intensity of shell pigmentation and the associated thermodynamic, physiological, and evolutionary consequences for snail diversity within the course of environmental warming are still insufficiently understood. To relate temperature‐driven internal heating, protein and membrane integrity impairment, escape behavior, place of residence selection, water loss, and mortality, we used experimentally warmed open‐top chambers and field observations with a total of >11,000 naturally or experimentally colored individuals of the highly polymorphic species Theba pisana (O.F. MÜller, 1774). We show that solar radiation in their natural Mediterranean habitat in Southern France poses intensifying thermal stress on increasingly pigmented snails that cannot be compensated for by behavioral responses. Individuals of all morphs acted neither jointly nor actively competed in climbing behavior, but acted similarly regardless of neighbor pigmentation intensity. Consequently, dark morphs progressively suffered from high internal temperatures, oxidative stress, and a breakdown of the chaperone system. Concomitant with increasing water loss, mortality increased with more intense pigmentation under simulated global warming conditions. In parallel with an increase in mean ambient temperature of 1.34°C over the past 30 years, the mortality rate of pigmented individuals in the field is, currently, about 50% higher than that of white morphs. A further increase of 1.12°C, as experimentally simulated in our study, would elevate this rate by another 26%. For 34 T. pisana populations from locations that are up to 2.7°C warmer than our experimental site, we show that both the frequency of pigmented morphs and overall pigmentation intensity decrease with an increase in average summer temperatures. We therefore predict a continuing strong decline in the frequency of pigmented morphs and a decrease in overall pigmentation intensity with ongoing global change in areas with strong solar radiation.     

Selective stimulation of Hsp27 and αB-crystallin but not Hsp70 expression by p38 MAP kinase activation

The levels of Hsp27 and αB-crystallin in C6 rat glioma cells, that had been heated at 43°C for 30 min with a subsequent culture for 16 h at 37°C, were markedly increased. The exposure of the cells to a low concentration (0.1–3 µg/ml) of anisomycin for a few hours after heat stress stimulated the accumulation of the small stress proteins Hsp27 and αB-crystallin, but not that of Hsp70. The levels of mRNAs for Hsp27 and αB-crystallin but not that for Hsp70 increased in cells that had been exposed to heat and subsequently for 2 h to 0.1–3 µg/ml anisomycin. The results of a reporter assay, using an αB-crystallin promotor fused to a luciferase reporter gene, suggested that the increase in level of αB-crystallin mRNA was due to the production of new mRNA. The activation of the binding of heat shock factors to heat shock elements induced in cells that had been heat stressed was barely affected by subsequent exposure to anisomycin at 0.3 µg/ml. The stimulatory effects of anisomycin were also observed in cells that had been exposed to NaAsO₂, or CdCl₂. The active form of p38 mitogen activated protein (MAP) kinase was increased in cell that had been subjected to heat shock and subsequent exposure to 0.3 µg/ml of anisomycin. The heat-induced accumulations of Hsp27 and αB-crystallin were also stimulated by cycloheximide, another stimulator of p38 MAP kinase. SB202190, a specific inhibitor of p38 MAP kinase, suppressed the stimulation by anisomycin of the heat stress-induced expressions of Hsp27 and αB-crystallin. These results suggest that the signal transduction pathway of the stress-induced expressions of Hsp27 and αB-crystallin in C6 glioma cells includes a process that is sensitive to p38 MAP kinase.     

Heat-preconditioning confers protection from Ca2+-mediated cell toxicity in renal tubular epithelial cells (BSC-1)

A rise in intracellular calcium may mediate ischemic damage by phospholipid hydrolysis and proteolysis Heat shock proteins have been shown to provide protection from various forms of cell stress, but not from models of Ca²⁺-mediated injury. The effect of heat preconditioning in a model of ionomycin-induced injury in cultured renal tubular epitheliaal cells (BSC-1) was examined. Hsp70-mRNA expression was induced by hyperthermia (HT) (42°C, 60 min). Hsp70 protein accumulation was maximal after 12–18 h and returned to baseline levels by 96 h. Treatment of BSC-1 cells with ionomycin (7.0 µM) produced lethal cell injury characterized by LDH release. Cells examined at 18 h after HT were significantly less damaged than cells studied at 96 h after HT. Our data are the first to demonstrate that heat preconditioning confers protection from Ca²⁺-mediated cell injury. The state of increased tolerance is transient and closely parallels kinetics of Hsp70 expression.     

Non-Lethal Heat Shock of the Asian Green Mussel,Perna viridis,Promotes Hsp70 Synthesis,Induces Thermotolerance and Protects Against Vibrio Infection

Mild heat stress promotes thermotolerance and protection against several different stresses in aquatic animals, consequences correlated with the accumulation of heat shock protein 70 (Hsp70). The purpose of this study was to determine if non-lethal heat shock (NLHS) of the Asian green mussel, Perna viridis, an aquatic species of commercial value, promoted the production of Hsp70 and enhanced its resistance to stresses. Initially, the LT₅₀ and LHT for P. viridis were determined to be 42°C and 44°C, respectively, with no heat shock induced death of mussels at 40°C or less. Immunoprobing of western blots revealed augmentation of constitutive (PvHsp70-1) and inducible (PvHsp70-2) Hsp70 in tissue from adductor muscle, foot, gill and mantel of P. viridis exposed to 38°C for 30 min followed by 6 h recovery, NLHS conditions for this organism. Characterization by liquid chromatography-tandem mass spectrometry (LC-MS/MS) revealed that PvHsp70-1 and PvHsp70-2 respectively corresponded most closely to Hsp70 from P. viridis and Mytilus galloprovincialis. Priming of adult mussels with NLHS promoted thermotolerance and increased resistance to V. alginolyticus. The induction of Hsp70 in parallel with enhanced thermotolerance and improved protection against V. alginolyticus, suggests Hsp70 functions in P. viridis as a molecular chaperone and as a stimulator of the immune system.     

COPD and levels of Hsp70 (HSPA1A) and Hsp27 (HSPB1) in plasma and lymphocytes among coal workers: a case-control study

This case-control study aimed to investigate whether the levels of Hsp70 (HSPA1A) and Hsp27 (HSPB1) in plasma and lymphocytes were associated with the risk of chronic obstructive pulmonary disease (COPD) among coal workers. A total of 76 COPD cases and 48 age-matched healthy controls from a group of coal workers were included. The case group consisted of 35 COPD patients whose condition was complicated with coal workers’ pneumoconiosis (CWP) and 41 COPD patients without CWP. Heat shock proteins (Hsps) in plasma and lymphocytes were detected by ELISA and flow cytometry, respectively. Multiple logistic regression models were applied to estimate the association between Hsp levels and COPD risk. Our results showed that plasma Hsp70 and lymphocyte Hsp27 levels were significantly higher and plasma Hsp27 levels were significantly lower in COPD cases than in controls (p < 0.01). No significant differences in lymphocyte Hsp70 levels were found between COPD cases and the matched subjects. Higher plasma Hsp70 levels (odds ratio (OR) = 13.8, 95 % confidence interval (CI) = 5.7–33.5) and lower plasma Hsp27 levels (OR = 4.6, 95 % CI = 2.0–10.5) were significantly associated with an increased risk of COPD after adjusting for confounders. Higher lymphocyte Hsp27 levels were only associated with an increased risk of COPD with CWP (OR = 6.6, 95 % CI = 2.0–22.1) but not with an increased risk of COPD without CWP (OR = 3.0, 95 % CI = 0.9–8.9). Additionally, there were strong joint effects of different Hsps on COPD risk. These results showed that higher levels of plasma Hsp70 and lower levels of plasma Hsp27 might be associated with an increased risk of COPD among coal workers. They may have the potential to serve as monitoring markers for COPD in coal workers.     

Immune defence under extreme ambient temperature

Owing to global climate change, the extreme weather conditions are predicted to become more frequent, which is suggested to have an even greater impact on ecological interactions than the gradual increase in average temperatures. Here, we examined whether exposure to high ambient temperature affects immune function of the great pond snail (Lymnaea stagnalis). We quantified the levels of several immune traits from snails maintained in a non-stressful temperature (15°C) and in an extreme temperature (30°C) that occurs in small ponds during hot summers. We found that snails exposed to high temperature had weaker immune defence, which potentially predisposes them to infections. However, while phenoloxidase and antibacterial activity of snail haemolymph were reduced at high temperature, haemocyte concentration was not affected. This suggests that the effect of high temperature on snail susceptibility to infections may vary across different pathogens because different components of invertebrate immune defence have different roles in resistance.     

Sexual selection and non-random mating for shell colour in a natural population of the marine snailLittorina mariae (Gastropoda: Prosobranchia)

In order to estimate the three independent components of mating behaviour, sexual selection in females, sexual selection in males and mating pattern, we studied the distribution of shell colour morphs among mating pairs and between copulating and non-copulating snails in four subsamples of a natural population ofL. mariae. The colour of the shell, the sex and a qualitative estimate of age was recorded for every snail. We found sexual selection acting against one of the two commonest colours (yellow) among the young females. However, in males none of the eight shell colour morphs was favoured during matings. Male sexual choice or differences in female sexual activity may cause the sexual fitness disadvantage of yellow females. Moreover, individuals of different colour morphs did not mate at random, rather dissasortatively. A behavioural choice among shell colour morphs or a non-random microdistribution of the morphs may cause the departure from random mating in this population.     

Gradual Rewarming with Gradual Increase in Pressure during Machine Perfusion after Cold Static Preservation Reduces Kidney Ischemia Reperfusion Injury

In this study we evaluated whether gradual rewarming after the period of cold ischemia would improve organ quality in an Isolated Perfused Kidney Model. Left rat kidneys were statically cold stored in University of Wisconsin solution for 24 hours at 4°C. After cold storage kidneys were rewarmed in one of three ways: perfusion at body temperature (38°C), or rewarmed gradually from 10°C to 38°C with stabilization at 10°C for 30 min and rewarmed gradually from 10°C to 38°C with stabilization at 25°C for 30 min. In the gradual rewarming groups the pressure was increased stepwise to 40 mmHg at 10°C and 70 mmHg at 25°C to counteract for vasodilatation leading to low perfusate flows. Renal function parameters and injury biomarkers were measured in perfusate and urine samples. Increases in injury biomarkers such as aspartate transaminase and lactate dehydrogenase in the perfusate were lower in the gradual rewarming groups versus the control group. Sodium re-absorption was improved in the gradual rewarming groups and reached significance in the 25°C group after ninety minutes of perfusion. HSP-70, ICAM-1, VCAM-1 mRNA expressions were decreased in the 10°C and 25°C groups. Based on the data kidneys that underwent gradual rewarming suffered less renal parenchymal, tubular injury and showed better endothelial preservation. Renal function improved in the gradual rewarming groups versus the control group.     

The Hsp70 Homologue Lhs1p Is Involved in a Novel Function of the Yeast Endoplasmic Reticulum, Refolding and Stabilization of Heat-denatured Protein Aggregates

Heat stress is an obvious hazard, and mechanisms to recover from thermal damage, largely unknown as of yet, have evolved in all organisms. We have recently shown that a marker protein in the ER of Saccharomyces cerevisiae, denatured by exposure of cells to 50°C after preconditioning at 37°C, was reactivated by an ATP-dependent machinery, when the cells were returned to physiological temperature 24°C. Here we show that refolding of the marker enzyme Hsp150Δ–β-lactamase, inactivated and aggregated by the 50°C treatment, required a novel ER-located homologue of the Hsp70 family, Lhs1p. In the absence of Lhs1p, Hsp150Δ–β-lactamase failed to be solubilized and reactivated and was slowly degraded. Coimmunoprecipitation experiments suggested that Lhs1p was somehow associated with heat-denatured Hsp150Δ– β-lactamase, whereas no association with native marker protein molecules could be detected. Similar findings were obtained for a natural glycoprotein of S. cerevisiae, pro-carboxypeptidase Y (pro-CPY). Lhs1p had no significant role in folding or secretion of newly synthesized Hsp150Δ–β-lactamase or pro-CPY, suggesting that the machinery repairing heat-damaged proteins may have specific features as compared to chaperones assisting de novo folding. After preconditioning and 50°C treatment, cells lacking Lhs1p remained capable of protein synthesis and secretion for several hours at 24°C, but only 10% were able to form colonies, as compared to wild-type cells. We suggest that Lhs1p is involved in a novel function operating in the yeast ER, refolding and stabilization against proteolysis of heatdenatured protein. Lhs1p may be part of a fundamental heat-resistant survival machinery needed for recovery of yeast cells from severe heat stress.     

Spermatozoon head size – the main differentiating feature between spermatozoa of blue and white Arctic fox (Vulpes lagopus)

Morphology and sperm morphometry, this is an important determinant of male reproductive capacity. Morphometric data may provide relevant information in studies focused on evolutionary biology, sperm quality assessment, including prediction of the potential fertility, semen cryopreservation, or the effect of reprotoxicants. The paper presents the morphometric analysis of spermatozoa from two colour morphs of Arctic fox (Vulpes lagopus), and attempts to determine the relationship between selected quality indicators and dimensions and shape of spermatozoa. The research material consisted of ejaculates collected once by manual stimulation from 20 one-year-old Arctic foxes (10 individuals of the blue morph and 10 of the white morph). Ejaculates were analysed for standard parameters (volume, sperm concentration, total number of spermatozoa in the ejaculate) and used for the preparation of microscopic specimens. It was found that, the dimensions of spermatozoa from Arctic foxes depend on the male colour morphs. Spermatozoa from white Arctic foxes were significantly longer (by 1.82 µm) and had larger heads (0.32 µm longer and 0.15 µm wider) compared to spermatozoa from blue Arctic foxes (P<0.05). The interactions between particular sperm dimensions indicated the occurrence of gametes differing in shape. The all correlation coefficients between the morphometric traits of spermatozoa were statistically significant. Our research proved that in the blue Arctic foxes, sperm dimensions (tail length and total sperm length) can be related to the percentage of spermatozoa with primary changes (respectively: r = -0.68 and r = -0.75; at P <0.05). However, in the case of white Arctic foxes, these characteristics depend on the ejaculate volume (respectively: r = 0.65 and r = 0.68; at P <0.05).     

Daily and seasonal changes in heat exposure and the Hsp70 level of individuals from a field population of Xeropicta derbentina (Krynicki 1836) (Pulmonata, Hygromiidae) in Southern France

The Mediterranean land snail Xeropicta derbentina forms huge populations in Southern France. In order to characterize heat exposure and the induction of the 70-kD heat shock protein (Hsp70) response system during the life cycle of this snail, a selected population from the Vaucluse area, Provence, was investigated encompassing the issues of morphological life cycle parameters (shell size and colouration), the daily courses of heat exposure at different heights above the ground, of shell temperature, and that of the individual Hsp70 levels. The study covered all four seasons of the year 2011. Snails were found to be annual, reaching their final size in August. The shell colouration pattern showed high variation in juveniles (spring) with a strong tendency towards becoming uniformly white at old age in autumn. In all seasons, ambient air temperature decreased with increasing distance from the ground surface during daytime while remaining constantly low in the night. Overall, the Hsp70 level of individuals followed the ambient temperature during diurnal and seasonal variations. Correlation analysis revealed a positive association of individual shell temperature and Hsp70 level for the most part of the life cycle of the snails until late summer, whereas a negative correlation was found for aged animals indicating senescence effects on the capacity of the stress response system.     

High temperature promotes amyloid β-protein production and γ-secretase complex formation via Hsp90

Alzheimer''s disease (AD) is characterized by neuronal loss and accumulation of β-amyloid-protein (Aβ) in the brain parenchyma. Sleep impairment is associated with AD and affects about 25–40% of patients in the mild-to-moderate stages of the disease. Sleep deprivation leads to increased Aβ production; however, its mechanism remains largely unknown. We hypothesized that the increase in core body temperature induced by sleep deprivation may promote Aβ production. Here, we report temperature-dependent regulation of Aβ production. We found that an increase in temperature, from 37 °C to 39 °C, significantly increased Aβ production in amyloid precursor protein-overexpressing cells. We also found that high temperature (39 °C) significantly increased the expression levels of heat shock protein 90 (Hsp90) and the C-terminal fragment of presenilin 1 (PS1-CTF) and promoted γ-secretase complex formation. Interestingly, Hsp90 was associated with the components of the premature γ-secretase complex, anterior pharynx-defective-1 (APH-1), and nicastrin (NCT) but was not associated with PS1-CTF or presenilin enhancer-2. Hsp90 knockdown abolished the increased level of Aβ production and the increased formation of the γ-secretase complex at high temperature in culture. Furthermore, with in vivo experiments, we observed increases in the levels of Hsp90, PS1-CTF, NCT, and the γ-secretase complex in the cortex of mice housed at higher room temperature (30 °C) compared with those housed at standard room temperature (23 °C). Our results suggest that high temperature regulates Aβ production by modulating γ-secretase complex formation through the binding of Hsp90 to NCT/APH-1.     

The molecular and metabolic program by which white adipocytes adapt to cool physiologic temperatures

Although visceral adipocytes located within the body’s central core are maintained at approximately 37°C, adipocytes within bone marrow, subcutaneous, and dermal depots are found primarily within the peripheral shell and generally exist at cooler temperatures. Responses of brown and beige/brite adipocytes to cold stress are well studied; however, comparatively little is known about mechanisms by which white adipocytes adapt to temperatures below 37°C. Here, we report that adaptation of cultured adipocytes to 31°C, the temperature at which distal marrow adipose tissues and subcutaneous adipose tissues often reside, increases anabolic and catabolic lipid metabolism, and elevates oxygen consumption. Cool adipocytes rely less on glucose and more on pyruvate, glutamine, and, especially, fatty acids as energy sources. Exposure of cultured adipocytes and gluteal white adipose tissue (WAT) to cool temperatures activates a shared program of gene expression. Cool temperatures induce stearoyl-CoA desaturase-1 (SCD1) expression and monounsaturated lipid levels in cultured adipocytes and distal bone marrow adipose tissues (BMATs), and SCD1 activity is required for acquisition of maximal oxygen consumption at 31°C.

Adipocytes in bone marrow, subcutaneous and dermal sites generally exist at temperatures below 37°C. This study identifies the molecular and metabolic program that adapts white adipocytes to these cooler environments.     

Physiological consequences of the supralittoral fringe: microhabitat temperature profiles and stress protein levels in the tropical periwinkle Cenchritis muricatus (Linneaus, 1758)

Habitat selection by marine snails is profoundly affected by variations in biotic and abiotic factors. In the supralittoral fringe of Caribbean rocky shores, the littorinid Cenchritis muricatus endures a near-terrestrial existence through a combination of active microhabitat choice and, during dry periods, repose. In this study, we sought to compare knobby periwinkle body size, thermal load, water loss, and stress protein expression among common supralittoral microhabitats to determine the physiological consequences of habitat selection. In this study, we show: (1) body temperatures in these snails exhibit daily fluctuations of more than 20°C and regularly exceed 46°C, (2) microhabitats differ in thermal stress over small spatial scales, with snails on black rocks and within crevices experiencing more extreme temperatures than snails on white rocks or grass, (3) water losses of 8.5% of total snail mass do not affect survival during 1 week, and (4) Hsp70, but not Hsp90, expression varies slightly among microhabitats but at a level much lower than physiologically possible. During arousal following hydration, snails exhibited substantially higher levels of Hsp70s than snails on dry substrates in the field. When inactive, Cenchritis appears to utilize a distinctly different physiological state consistent with aestivation metabolism and does not exhibit significant up-regulation of inducible heat shock proteins (Hsps). In summary, studies lacking detailed thermal and hydration history, and relying only upon Hsp levels, may misrepresent the true physiological consequences of microhabitat choice for high-shore tropical gastropods.     

Deleterious consequences of Hsp70 overexpression in Drosphilla melanogaster larvae

We compared transgenic Drosophila larvae varying in hsp70 copy number the consequences of Hsp70 overexpression for growth and development after heat shock. Exposure to a mildy elevated temperature (36°C) induced expression of Hsp70 (and presumably other heat shock proteins) and improved tolerance of more severe heat stress, 38.5–39.5°C. We examined this pattern in two independently derived pairs of extra-copy and excision strains that different primarily in hsp70 copy number (with 22 and 10 copies, respectively). Extra-copy larvae produced more Hsp70 in response to high temperature than did excision larvae, but surpassed the excision strain in survival only immediately after thermal stress. Excision larvae survived to adulthood at higher proportions than did extra-copy larvae and grew more rapidly after thermal stress. Furthermore, multiple pretreatment reduced survival of 1st-instar extra-copy larvae, but did not affect the corresponding excision strain. While extra Hsp70 provides additional protection against the immediate damage from heat stress, abnormally high concentrations can decrease growth, development and survival to adulthood.