Regulation of apoptotic mediators reveals dynamic responses to thermal stress in the reef building coral Acropora millepora

Background

Mass coral bleaching is increasing in scale and frequency across the world''s coral reefs and is being driven primarily by increased levels of thermal stress arising from global warming. In order to understand the impacts of projected climate change upon corals reefs, it is important to elucidate the underlying cellular mechanisms that operate during coral bleaching and subsequent mortality. In this respect, increased apoptotic cell death activity is an important cellular process that is associated with the breakdown of the mutualistic symbiosis between the cnidarian host and their dinoflagellate symbionts.

Methodology/Principal Findings

The present study reports the impacts of different stressors (colchicine and heat stress) on three phases of apoptosis: (i) the potential initiation by differential expression of Bcl-2 members, (ii) the execution of apoptotic events by activation of caspase 3-like proteases and (iii) and finally, the cell disposal indicated by DNA fragmentation in the reef building coral Acropora millepora. In corals incubated with colchicine, an increase in caspase 3-like activity and DNA fragmentation was associated with a relative down-regulation of Bcl-2, suggesting that the initiation of apoptosis may be mediated by the suppression of an anti-apoptotic mechanism. In contrast, in the early steps of heat stress, the induction of caspase-dependent apoptosis was related to a relative up-regulation of Bcl-2 consecutively followed by a delayed decrease in apoptosis activity.

Conclusions/Significance

In the light of these results, we propose a model of heat stress in coral hosts whereby increasing temperatures engage activation of caspase 3-dependent apoptosis in cells designated for termination, but also the onset of a delayed protective response involving overexpression of Bcl-2 in surviving cells. This mitigating response to thermal stress could conceivably be an important regulatory mechanism for cell survival in corals exposed to sudden environmental changes.     

Nephric duct insertion is a crucial step in urinary tract maturation that is regulated by a Gata3-Raldh2-Ret molecular network in mice

Urinary tract development depends on a complex series of events in which the ureter moves from its initial branch point on the nephric duct (ND) to its final insertion site in the cloaca (the primitive bladder and urethra). Defects in this maturation process can result in malpositioned ureters and hydronephrosis, a common cause of renal disease in children. Here, we report that insertion of the ND into the cloaca is an unrecognized but crucial step that is required for proper positioning of the ureter and that depends on Ret signaling. Analysis of Ret mutant mice at birth reveals hydronephrosis and defective ureter maturation, abnormalities that our results suggest are caused, at least in part, by delayed insertion of the ND. We find a similar set of malformations in mutants lacking either Gata3 or Raldh2. We show that these factors act in parallel to regulate ND insertion via Ret. Morphological analysis of ND extension in wild-type embryos reveals elaborate cellular protrusions at ND tips that are not detected in Ret, Gata3 or Raldh2 mutant embryos, suggesting that these protrusions may normally be important for fusion with the cloaca. Together, our studies reveal a novel Ret-dependent event, ND insertion, that, when abnormal, can cause obstruction and hydronephrosis at birth; whether ND defects underlie similar types of urinary tract abnormalities in humans is an interesting possibility.     

Metagenomic exploration of antibiotic resistance in soil

The ongoing development of metagenomic approaches is providing the means to explore antibiotic resistance in nature and address questions that could not be answered previously with conventional culture-based strategies. The number of available environmental metagenomic sequence datasets is rapidly expanding and henceforth offer the ability to gain a more comprehensive understanding of antibiotic resistance at the global scale. Although there is now evidence that the environment constitutes a vast reservoir of antibiotic resistance gene determinants (ARGDs) and that the majority of ARGDs acquired by human pathogens may have an environmental origin, a better understanding of their diversity, prevalence and ecological significance may help predict the emergence and spreading of newly acquired resistances. Recent applications of metagenomic approaches to the study of ARGDs in natural environments such as soil should help overcome challenges concerning expanding antibiotic resistances.     

The multifunctional LigB adhesin binds homeostatic proteins with potential roles in cutaneous infection by pathogenic Leptospira interrogans

Leptospirosis is a potentially fatal zoonotic disease in humans and animals caused by pathogenic spirochetes, such as Leptospira interrogans. The mode of transmission is commonly limited to the exposure of mucous membrane or damaged skin to water contaminated by leptospires shed in the urine of carriers, such as rats. Infection occurs during seasonal flooding of impoverished tropical urban habitats with large rat populations, but also during recreational activity in open water, suggesting it is very efficient. LigA and LigB are surface localized proteins in pathogenic Leptospira strains with properties that could facilitate the infection of damaged skin. Their expression is rapidly induced by the increase in osmolarity encountered by leptospires upon transition from water to host. In addition, the immunoglobulin-like repeats of the Lig proteins bind proteins that mediate attachment to host tissue, such as fibronectin, fibrinogen, collagens, laminin, and elastin, some of which are important in cutaneous wound healing and repair. Hemostasis is critical in a fresh injury, where fibrinogen from damaged vasculature mediates coagulation. We show that fibrinogen binding by recombinant LigB inhibits fibrin formation, which could aid leptospiral entry into the circulation, dissemination, and further infection by impairing healing. LigB also binds fibroblast fibronectin and type III collagen, two proteins prevalent in wound repair, thus potentially enhancing leptospiral adhesion to skin openings. LigA or LigB expression by transformation of a nonpathogenic saprophyte, L. biflexa, enhances bacterial adhesion to fibrinogen. Our results suggest that by binding homeostatic proteins found in cutaneous wounds, LigB could facilitate leptospirosis transmission. Both fibronectin and fibrinogen binding have been mapped to an overlapping domain in LigB comprising repeats 9-11, with repeat 11 possibly enhancing binding by a conformational effect. Leptospirosis patient antibodies react with the LigB domain, suggesting applications in diagnosis and vaccines that are currently limited by the strain-specific leptospiral lipopolysaccharide coats.     

Hyperforin inhibits Akt1 kinase activity and promotes caspase-mediated apoptosis involving Bad and Noxa activation in human myeloid tumor cells

Background

The natural phloroglucinol hyperforin HF displays anti-inflammatory and anti-tumoral properties of potential pharmacological interest. Acute myeloid leukemia (AML) cells abnormally proliferate and escape apoptosis. Herein, the effects and mechanisms of purified HF on AML cell dysfunction were investigated in AML cell lines defining distinct AML subfamilies and primary AML cells cultured ex vivo.

Methodology and Results

HF inhibited in a time- and concentration-dependent manner the growth of AML cell lines (U937, OCI-AML3, NB4, HL-60) by inducing apoptosis as evidenced by accumulation of sub-G1 population, phosphatidylserine externalization and DNA fragmentation. HF also induced apoptosis in primary AML blasts, whereas normal blood cells were not affected. The apoptotic process in U937 cells was accompanied by downregulation of anti-apoptotic Bcl-2, upregulation of pro-apoptotic Noxa, mitochondrial membrane depolarization, activation of procaspases and cleavage of the caspase substrate PARP-1. The general caspase inhibitor Z-VAD-fmk and the caspase-9- and -3-specific inhibitors, but not caspase-8 inhibitor, significantly attenuated apoptosis. HF-mediated apoptosis was associated with dephosphorylation of active Akt1 (at Ser⁴⁷³) and Akt1 substrate Bad (at Ser¹³⁶) which activates Bad pro-apoptotic function. HF supppressed the kinase activity of Akt1, and combined treatment with the allosteric Akt1 inhibitor Akt-I-VIII significantly enhanced apoptosis of U937 cells.

Significance

Our data provide new evidence that HF''s pro-apoptotic effect in AML cells involved inhibition of Akt1 signaling, mitochondria and Bcl-2 members dysfunctions, and activation of procaspases -9/-3. Combined interruption of mitochondrial and Akt1 pathways by HF may have implications for AML treatment.     

DPPC Bilayers in Solutions of High Sucrose Content

The properties of lipid bilayers in sucrose solutions have been intensely scrutinized over recent decades because of the importance of sugars in the field of biopreservation. However, a consensus has not yet been formed on the mechanisms of sugar-lipid interaction. Here, we present a study on the effect of sucrose on 1,2-dipalmitoyl-sn-glycero-3-phosphocholine bilayers that combines calorimetry, spectral fluorimetry, and optical microscopy. Intriguingly, our results show a significant decrease in the transition enthalpy but only a minor shift in the transition temperature. Our observations can be quantitatively accounted for by a thermodynamic model that assumes partial delayed melting induced by sucrose adsorption at the membrane interface.     

Lethal activity of miconazole in Neisseria gonorrhoeae grown in pure and mixed cultures

Abstract Low concentrations (e.g. 2 × 10⁻⁶ M) of an imidazole derivative anti-fungal agent, miconazole, were lethal for the Gram-negative, facultative aerobic pathogen Neisseria gonorrhoeae grown either alone or in mixed culture with the yeast Candida albicans . Electron microscopic observation of Neisseria cells exposed to miconazole showed the presence of blebs in the outer wall and areas of separation between the wall and the cytoplasmic membrane. Sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) of cell lysates did not reveal differences in major outer membrane proteins between the treated and the untreated cells of any one strain. Imidazole derivatives are frequently used in the treatment of candidiasis. Our in vitro results show that low concentrations of one of them, miconazole, can be bactericidal for N. gonorrhoeae , a bacterium that can colonise sites of the human body where Candida is often found.     

Growth and resilience responses of Scots pine to extreme droughts across Europe depend on predrought growth conditions

Global climate change is expected to further raise the frequency and severity of extreme events, such as droughts. The effects of extreme droughts on trees are difficult to disentangle given the inherent complexity of drought events (frequency, severity, duration, and timing during the growing season). Besides, drought effects might be modulated by trees’ phenotypic variability, which is, in turn, affected by long‐term local selective pressures and management legacies. Here we investigated the magnitude and the temporal changes of tree‐level resilience (i.e., resistance, recovery, and resilience) to extreme droughts. Moreover, we assessed the tree‐, site‐, and drought‐related factors and their interactions driving the tree‐level resilience to extreme droughts. We used a tree‐ring network of the widely distributed Scots pine (Pinus sylvestris) along a 2,800 km latitudinal gradient from southern Spain to northern Germany. We found that the resilience to extreme drought decreased in mid‐elevation and low productivity sites from 1980–1999 to 2000–2011 likely due to more frequent and severe droughts in the later period. Our study showed that the impact of drought on tree‐level resilience was not dependent on its latitudinal location, but rather on the type of sites trees were growing at and on their growth performances (i.e., magnitude and variability of growth) during the predrought period. We found significant interactive effects between drought duration and tree growth prior to drought, suggesting that Scots pine trees with higher magnitude and variability of growth in the long term are more vulnerable to long and severe droughts. Moreover, our results indicate that Scots pine trees that experienced more frequent droughts over the long‐term were less resistant to extreme droughts. We, therefore, conclude that the physiological resilience to extreme droughts might be constrained by their growth prior to drought, and that more frequent and longer drought periods may overstrain their potential for acclimation.