Stealth proteins: in silico identification of a novel protein family rendering bacterial pathogens invisible to host immune defense

There are a variety of bacterial defense strategies to survive in a hostile environment. Generation of extracellular polysaccharides has proved to be a simple but effective strategy against the host's innate immune system. A comparative genomics approach led us to identify a new protein family termed Stealth, most likely involved in the synthesis of extracellular polysaccharides. This protein family is characterized by a series of domains conserved across phylogeny from bacteria to eukaryotes. In bacteria, Stealth (previously characterized as SacB, XcbA, or WefC) is encoded by subsets of strains mainly colonizing multicellular organisms, with evidence for a protective effect against the host innate immune defense. More specifically, integrating all the available information about Stealth proteins in bacteria, we propose that Stealth is a D-hexose-1-phosphoryl transferase involved in the synthesis of polysaccharides. In the animal kingdom, Stealth is strongly conserved across evolution from social amoebas to simple and complex multicellular organisms, such as Dictyostelium discoideum, hydra, and human. Based on the occurrence of Stealth in most Eukaryotes and a subset of Prokaryotes together with its potential role in extracellular polysaccharide synthesis, we propose that metazoan Stealth functions to regulate the innate immune system. Moreover, there is good reason to speculate that the acquisition and spread of Stealth could be responsible for future epidemic outbreaks of infectious diseases caused by a large variety of eubacterial pathogens. Our in silico identification of a homologous protein in the human host will help to elucidate the causes of Stealth-dependent virulence. At a more basic level, the characterization of the molecular and cellular function of Stealth proteins may shed light on fundamental mechanisms of innate immune defense against microbial invasion.     

Neurotransmitter release regulated by a MALS-liprin-alpha presynaptic complex

Synapses are highly specialized intercellular junctions organized by adhesive and scaffolding molecules that align presynaptic vesicular release with postsynaptic neurotransmitter receptors. The MALS/Veli-CASK-Mint-1 complex of PDZ proteins occurs on both sides of the synapse and has the potential to link transsynaptic adhesion molecules to the cytoskeleton. In this study, we purified the MALS protein complex from brain and found liprin-alpha as a major component. Liprin proteins organize the presynaptic active zone and regulate neurotransmitter release. Fittingly, mutant mice lacking all three MALS isoforms died perinatally with difficulty breathing and impaired excitatory synaptic transmission. Excitatory postsynaptic currents were dramatically reduced in autaptic cultures from MALS triple knockout mice due to a presynaptic deficit in vesicle cycling. These findings are consistent with a model whereby the MALS-CASK-liprin-alpha complex recruits components of the synaptic release machinery to adhesive proteins of the active zone.     

Microbial Community in Black Rust Exposed to Hot Ridge Flank Crustal Fluids

During Integrated Ocean Drilling Program Expedition 301, we obtained a sample of black rust from a circulation obviation retrofit kit (CORK) observatory at a borehole on the eastern flank of Juan de Fuca Ridge. Due to overpressure, the CORK had failed to seal the borehole. Hot fluids from oceanic crust had discharged to the overlying bottom seawater and resulted in the formation of black rust analogous to a hydrothermal chimney deposit. Both culture-dependent and culture-independent analyses indicated that the black-rust-associated community differed from communities reported from other microbial habitats, including hydrothermal vents at seafloor spreading centers, while it shared phylotypes with communities previously detected in crustal fluids from the same borehole. The most frequently retrieved sequences of bacterial and archaeal 16S rRNA genes were related to the genera Ammonifex and Methanothermococcus, respectively. Most phylotypes, including phylotypes previously detected in crustal fluids, were isolated in pure culture, and their metabolic traits were determined. Quantification of the dissimilatory sulfite reductase (dsrAB) genes, together with stable sulfur isotopic and electron microscopic analyses, strongly suggested the prevalence of sulfate reduction, potentially by the Ammonifex group of bacteria. Stable carbon isotopic analyses suggested that the bulk of the microbial community was trophically reliant upon photosynthesis-derived organic matter. This report provides important insights into the phylogenetic, physiological, and trophic characteristics of subseafloor microbial ecosystems in warm ridge flank crusts.     

Minimally invasive quantification of lymph flow in mice and rats by imaging depot clearance of near-infrared albumin

There is a lack of available methods to noninvasively quantify lymphatic function in small experimental animals, a necessity for studies on lymphatic system pathophysiology. We present a new method to quantify lymph flow in mice and rats, based on optically monitoring the depot clearance of near-infrared fluorescently labeled albumin and subsequent calculation of removal rate constants (k). BSA was conjugated with Alexa680 NHS ester and remained stable in protein-rich solutions without free dye dissociation. To assess lymph flow, mice or rats were imaged every 30 or 60 min during a 3- to 6-h period following an intradermal injection of 0.5 or 1 μl Alexa680-albumin. Mice were awake between measurements, whereas rats were anesthetized throughout the experiment. The k, a parameter defined as equivalent to lymph flow, was calculated from the slopes of the resultant log-linear washout curves and averaged -0.40 ± 0.03 and -0.30 ± 0.02%/min for control C57BL/6 and C3H mice, respectively. Local administration of the vasoconstrictor endothelin-1 in mice led to a significant reduction in k, whereas overhydration in rats increased k, reflecting the coupling between capillary filtration and lymph flow. Furthermore, k was 50% of wild type in lymphedema Chy mice where dermal lymphatics are absent. We conclude that lymph flow can be determined as its rate constant k by optical imaging of depot clearance of submicroliter amounts of Alexa680-albumin. The method offers a minimally invasive, reproducible, and simple alternative to assess lymphatic function in mice and rats.     

Dietary supplementation with acetyl-l-carnitine in seizure treatment of pentylenetetrazole kindled mice

In spite of the availability of new antiepileptic drugs a considerable number of epilepsy patients still have pharmacoresistant seizures, and thus there is a need for novel approaches. Acetyl-l-carnitine (ALCAR), which delivers acetyl units to mitochondria for acetyl-CoA production, has been shown to improve brain energy homeostasis and protects against various neurotoxic insults. To our knowledge, this is the first study of ALCAR's effect on metabolism in pentylenetetrazole (PTZ) kindled mice. ALCAR or the commonly used antiepileptic drug valproate, was added to the drinking water of mice for 25days, and animals were injected with PTZ or saline three times a week during the last 21days. In order to investigate ALCAR's effects on glucose metabolism, mice were injected with [1-(13)C]glucose 15min prior to microwave fixation. Brain extracts from cortex and the hippocampal formation (HF) were studied using (1)H and (13)C NMR spectroscopy and HPLC. PTZ kindling caused glucose hypometabolism, evidenced by a reduction in both glycolysis and TCA cycle turnover in both brain regions investigated. Glutamatergic and GABAergic neurons were affected in cortex and HF, but the amount of glutamate was only reduced in HF. Slight astrocytic involvement could be detected in the cortex. Interestingly, the dopamine content was increased in the HF. ALCAR attenuated the PTZ induced reduction in [3-(13)C]alanine and the increase in dopamine in the HF. However, TCA cycle metabolism was not different from that seen in PTZ kindled animals. In conclusion, even though ALCAR did not delay the kindling process, it did show some promising ameliorative effects, worthy of further investigation.     

A review on self-destructive defense behaviors in social insects

Colony defense is a necessary but dangerous task for social insects, and nest defensive behaviors often lead to a premature death of the actor. As an extreme form of colony defense, self-sacrificial behaviors have evolved by kin selection in various social insects. Most self-sacrificial defensive mechanisms occur in response to an acute threat to the colony, but some behaviors are preemptive actions that avert harm to the colony. Self-sacrifice has also been observed as a form of preemptive defense against parasites and pathogens where individuals will abandon their normal colony function and die in self-exile to reduce the risk of infecting nestmates. Here, we provide an overview of the self-destructive defense mechanisms that eusocial insects have evolved and discuss avenues for future research into this form of altruism.     

Temperature responses of dark respiration in relation to leaf sugar concentration

Changes in leaf sugar concentrations are a possible mechanism of short‐term adaptation to temperature changes, with natural fluctuations in sugar concentrations in the field expected to modify the heat sensitivity of respiration. We studied temperature‐response curves of leaf dark respiration in the temperate tree Populus tremula (L.) in relation to leaf sugar concentration (1) under natural conditions or (2) leaves with artificially enhanced sugar concentration. Temperature‐response curves were obtained by increasing the leaf temperature at a rate of 1°C min^?1. We demonstrate that respiration, similarly to chlorophyll fluorescence, has a break‐point at high temperature, where respiration starts to increase with a faster rate. The average break‐point temperature (T_RD) was 48.6 ± 0.7°C at natural sugar concentration. Pulse‐chase experiments with ¹⁴CO₂ demonstrated that substrates of respiration were derived mainly from the products of starch degradation. Starch degradation exhibited a similar temperature‐response curve as respiration with a break‐point at high temperatures. Acceleration of starch breakdown may be one of the reasons for the observed high‐temperature rise in respiration. We also demonstrate that enhanced leaf sugar concentrations or enhanced osmotic potential may protect leaf cells from heat stress, i.e. higher sugar concentrations significantly modify the temperature‐response curve of respiration, abolishing the fast increase of respiration. Sugars or enhanced osmotic potential may non‐specifically protect respiratory membranes or may block the high‐temperature increase in starch degradation and consumption in respiratory processes, thus eliminating the break‐points in temperature curves of respiration in sugar‐fed leaves.     

Population dynamics in changing environments: the case of an eruptive forest pest species

In recent decades we have seen rapid and co‐occurring changes in landscape structure, species distributions and even climate as consequences of human activity. Such changes affect the dynamics of the interaction between major forest pest species, such as bark beetles (Coleoptera: Curculionidae, Scolytinae), and their host trees. Normally breeding mostly in broken or severely stressed spruce; at high population densities some bark beetle species can colonise and kill healthy trees on scales ranging from single trees in a stand to multi‐annual landscape‐wide outbreaks. In Eurasia, the largest outbreaks are caused by the spruce bark beetle, Ips typographus (Linnaeus), which is common and shares a wide distribution with its main host, Norway spruce (Picea abies Karst.). A large literature is now available, from which this review aims to synthesize research relevant for the population dynamics of I. typographus and co‐occurring species under changing conditions. We find that spruce bark beetle population dynamics tend to be metastable, but that mixed‐species and age‐heterogeneous forests with good site‐matching tend to be less susceptible to large‐scale outbreaks. While large accumulations of logs should be removed and/or debarked before the next swarming period, intensive removal of all coarse dead wood may be counterproductive, as it reduces the diversity of predators that in some areas may play a role in keeping I. typographus populations below the outbreak threshold, and sanitary logging frequently causes edge effects and root damage, reducing the resistance of remaining trees. It is very hard to predict the outcome of interspecific interactions due to invading beetle species or I. typographus establishing outside its current range, as they can be of varying sign and strength and may fluctuate depending on environmental factors and population phase. Most research indicates that beetle outbreaks will increase in frequency and magnitude as temperature, wind speed and precipitation variability increases, and that mitigating forestry practices should be adopted as soon as possible considering the time lags involved.