Necrotic Cells Actively Attract Phagocytes through the Collaborative Action of Two Distinct PS-Exposure Mechanisms

Necrosis, a kind of cell death closely associated with pathogenesis and genetic programs, is distinct from apoptosis in both morphology and mechanism. Like apoptotic cells, necrotic cells are swiftly removed from animal bodies to prevent harmful inflammatory and autoimmune responses. In the nematode Caenorhabditis elegans, gain-of-function mutations in certain ion channel subunits result in the excitotoxic necrosis of six touch neurons and their subsequent engulfment and degradation inside engulfing cells. How necrotic cells are recognized by engulfing cells is unclear. Phosphatidylserine (PS) is an important apoptotic-cell surface signal that attracts engulfing cells. Here we observed PS exposure on the surface of necrotic touch neurons. In addition, the phagocytic receptor CED-1 clusters around necrotic cells and promotes their engulfment. The extracellular domain of CED-1 associates with PS in vitro. We further identified a necrotic cell-specific function of CED-7, a member of the ATP-binding cassette (ABC) transporter family, in promoting PS exposure. In addition to CED-7, anoctamin homolog-1 (ANOH-1), the C. elegans homolog of the mammalian Ca²⁺-dependent phospholipid scramblase TMEM16F, plays an independent role in promoting PS exposure on necrotic cells. The combined activities from CED-7 and ANOH-1 ensure efficient exposure of PS on necrotic cells to attract their phagocytes. In addition, CED-8, the C. elegans homolog of mammalian Xk-related protein 8 also makes a contribution to necrotic cell-removal at the first larval stage. Our work indicates that cells killed by different mechanisms (necrosis or apoptosis) expose a common “eat me” signal to attract their phagocytic receptor(s); furthermore, unlike what was previously believed, necrotic cells actively present PS on their outer surfaces through at least two distinct molecular mechanisms rather than leaking out PS passively.     

Two alternative mechanisms that regulate the presentation of apoptotic cell engulfment signal in Caenorhabditis elegans

Phosphatidylserine exposed on the surface of apoptotic mammalian cells is considered an "eat-me" signal that attracts phagocytes. The generality of using phosphatidylserine as a clearance signal for apoptotic cells in animals and the regulation of this event remain uncertain. Using ectopically expressed mouse MFG-E8, a secreted phosphatidylserine-binding protein, we detected specific exposure of phosphatidylserine on the surface of apoptotic cells in Caenorhabditis elegans. Masking the surface phosphatidylserine inhibits apoptotic cell engulfment. CED-7, an ATP-binding cassette (ABC) transporter, is necessary for the efficient exposure of phosphatidylserine on apoptotic somatic cells, and for the recognition of these cells by phagocytic receptor CED-1. Alternatively, phosphatidylserine exposure on apoptotic germ cells is not CED-7 dependent, but instead requires phospholipid scramblase PLSC-1, a homologue of mammalian phospholipid scramblases. Moreover, deleting plsc-1 results in the accumulation of apoptotic germ cells but not apoptotic somatic cells. These observations suggest that phosphatidylserine might be recognized by CED-1 and act as a conserved eat-me signal from nematodes to mammals. Furthermore, the two different biochemical activities used in somatic cells (ABC transporter) and germ cells (phospholipid scramblase) suggest an increased complexity in the regulation of phosphatidylserine presentation in response to apoptotic signals in different tissues and during different developmental stages.     

Quantum dot-based assay for Cu quantification in bacterial cell culture

A simple and sensitive method for quantification of nanomolar copper with a detection limit of 1.2 × 10⁻¹⁰ M and a linear range from 10⁻⁹ to 10⁻⁸ M is reported. For the most useful analytical concentration of quantum dots, 1160 μg/ml, a 1/K_sv value of 11 μM Cu²⁺ was determined. The method is based on the interaction of Cu²⁺ with glutathione-capped CdTe quantum dots (CdTe–GSH QDs) synthesized by a simple and economic biomimetic method. Green CdTe–GSH QDs displayed the best performance in copper quantification when QDs of different sizes/colors were tested. Cu²⁺ quantification is highly selective given that no significant interference of QDs with 19 ions was observed. No significant effects on Cu²⁺ quantification were determined when different reaction matrices such as distilled water, tap water, and different bacterial growth media were tested. The method was used to determine copper uptake kinetics on Escherichia coli cultures. QD-based quantification of copper on bacterial supernatants was compared with atomic absorption spectroscopy as a means of confirming the accuracy of the reported method. The mechanism of Cu²⁺-mediated QD fluorescence quenching was associated with nanoparticle decomposition.     

Topographical distribution of pulmonary perfusion and ventilation, assessed by PET in supine and prone humans.

Using positron emission tomography (PET) and intravenously injected (13)N(2), we assessed the topographical distribution of pulmonary perfusion (Q) and ventilation (V) in six healthy, spontaneously breathing subjects in the supine and prone position. In this technique, the intrapulmonary distribution of (13)N(2), measured during a short apnea, is proportional to regional Q. After resumption of breathing, regional specific alveolar V (sVA, ventilation per unit of alveolar gas volume) can be calculated from the tracer washout rate. The PET scanner imaged 15 contiguous, 6-mm-thick, slices of lung. Vertical gradients of Q and sVA were computed by linear regression, and spatial heterogeneity was assessed from the squared coefficient of variation (CV(2)). Both CV and CV were corrected for the estimated contribution of random imaging noise. We found that 1) both Q and V had vertical gradients favoring dependent lung regions, 2) vertical gradients were similar in the supine and prone position and explained, on average, 24% of Q heterogeneity and 8% of V heterogeneity, 3) CV was similar in the supine and prone position, and 4) CV was lower in the prone position. We conclude that, in recumbent, spontaneously breathing humans, 1) vertical gradients favoring dependent lung regions explain a significant fraction of heterogeneity, especially of Q, and 2) although Q does not seem to be systematically more homogeneous in the prone position, differences in individual behaviors may make the prone position advantageous, in terms of V-to-Q matching, in selected subjects.     

Integrative taxonomy and preliminary assessment of species limits in the Liolaemus walkeri complex (Squamata,Liolaemidae) with descriptions of?three?new species from Peru

Species delimitation studies based on integrative taxonomic approaches have received considerable attention in the last few years, and have provided the strongest hypotheses of species boundaries. We used three lines of evidence (molecular, morphological, and niche envelopes) to test for species boundaries in Peruvian populations of the Liolaemus walkeri complex. Our results show that different lines of evidence and analyses are congruent in different combinations, for unambiguous delimitation of three lineages that were “hidden” within known species, and now deserve species status. Our phylogenetic analysis shows that L. walkeri, L. tacnae and the three new species are strongly separated from other species assigned to the alticolor-bibronii group. Few conventional morphological characters distinguish the new species from closely related taxa and this highlights the need to integrate other sources of data to erect strong hypothesis of species limits. A taxonomic key for known Peruvian species of the subgenus Lioalemus is provided.     

Global assessment of marine biodiversity potentially threatened by offshore hydrocarbon activities

Increasing global energy demands have led to the ongoing intensification of hydrocarbon extraction from marine areas. Hydrocarbon extractive activities pose threats to native marine biodiversity, such as noise, light, and chemical pollution, physical changes to the sea floor, invasive species, and greenhouse gas emissions. Here, we assessed at a global scale the spatial overlap between offshore hydrocarbon activities and marine biodiversity (>25,000 species, nine major ecosystems, and marine protected areas), and quantify the changes over time. We discovered that two‐thirds of global offshore hydrocarbon activities occur in areas within the top 10% for species richness, range rarity, and proportional range rarity values globally. Thus, while hydrocarbon activities are undertaken in less than one percent of the ocean's area, they overlap with approximately 85% of all assessed species. Of conservation concern, 4% of species with the largest proportion of their range overlapping hydrocarbon activities are range restricted, potentially increasing their vulnerability to localized threats such as oil spills. While hydrocarbon activities have extended to greater depths since the mid‐1990s, we found that the largest overlap is with coastal ecosystems, particularly estuaries, saltmarshes and mangroves. Furthermore, in most countries where offshore hydrocarbon exploration licensing blocks have been delineated, they do not overlap with marine protected areas (MPAs). Although this is positive in principle, many countries have far more licensing block areas than protected areas, and in some instances, MPA coverage is minimal. These findings suggest the need for marine spatial prioritization to help limit future spatial overlap between marine conservation priorities and hydrocarbon activities. Such prioritization can be informed by the spatial and quantitative baseline information provided here. In increasingly shared seascapes, prioritizing management actions that set both conservation and development targets could help minimize further declines of biodiversity and environmental changes at a global scale.     

Progress towards the production of very long-chain polyunsaturated fatty acid in transgenic plants: plant metabolic engineering comes of age

Very long-chain polyunsaturated fatty acids are vital components of the human diet, playing important multiple roles in optimal health and development. Current dietary sources, in the form of fish oils, are in decline, and an alternative sustainable source is required. Recent attempts to engineer transgenic plants with the capacity to synthesize these fatty acids have clearly demonstrated the possibility of such an approach. This represents a major breakthrough in the quest for alternative sources of fish oils, as well as fulfilling the promise of transgenic plants as green factories.     

Isolation and structure of a yeast initiator tRNAmet gene.

Sixteen bacterial clones containing yeast initiator tRNAmet genes have been isolated. The size of the BamHI fragments encoding these genes ranges from 4,000 to 23,000 base pairs. The nucleotide sequence of one member of this group has been determined. It has no intervening sequences.     

The effect of pH on the structure and activity of yeast RNA polymerase I

The analysis of the effect of pH upon the rate of polymerization indicates that the activity of yeast RNA polymerase I is optimal between pH 7.5 and 9 and depends on the ionization state of two groups with apparent pK_a values of 6.5 and 10. Yeast RNA polymerase I is extremely labile at acid pH. Below pH 5 the enzyme is irreversibly inactivated by [H⁺], with a second-order rate constant of 1.6 × 10^?4m^?1 min^?1. Sucrose gradient sedimentation and gel electrophoresis analysis of the enzyme inactivated at acid pH indicates the sequential dissociation of several enzyme subunits. The polypeptides of 44,000 and 24,000 daltons dissociate first from the enzyme core followed by the dissociation of the polypeptides of 48,000 and 36,000 daltons.     

Alterations in regional ventilation, perfusion, and shunt after smoke inhalation measured by PET.

Regional changes in ventilation and perfusion occurring in the early hours after smoke inhalation injury were evaluated through the use of positron emission tomography. Five lambs were imaged before and 1, 2, and 4 h after receiving 100 breaths of cotton smoke. Utilizing a recently developed model of (13)N tracer kinetics (3), we evaluated changes in ventilation, perfusion, shunt, and regional gas content in nondependent, middle, and dependent lung zones. The data demonstrated a progressive development of regional shunt in dependent (dorsal) regions in which perfusion remained the highest throughout the study. These findings, together with decreasing regional ventilation and fractional gas content in the dependent regions, correlated with decreasing arterial Pa(O(2)) values over the course of the study. A negative correlation between regional shunt fraction and regional gas content in dependent and middle regions suggests that shunt was caused by progressive alveolar derecruitment or flooding.