Contribution of [alpha]--galactopyranosyl end groups to attachment of highly and low metastatic murine fibrosarcoma cells to various substrates

There are much greater numbers of cell surface terminal, non-reducing [alpha]--galactorpyranosyl groups in highly malignant (metastatic) cells than are found in low malignant cells derived from the same murine fibrosarcoma. We have examined the contribution of these residues to attachment of the cells to various collagens and to plastic. Removal of these carbohydrate groups with [alpha]-galactosidase or blocking them with lectins from Griffonia simplicifolia seeds or with anti-blood group B antiserum all dramatically inhibited the attachment of both the highly malignant and the low malignant cells. Following removal with the enzyme, the [alpha]--galactopyranosyl end groups were rapidly resynthesized. This resynthesis was inhibited by tunicamycin, an inhibitor of de novo glycoprotein synthesis. This antibiotic also impaired cell attachment and, when used in addition to treatment with [alpha]-galactosidase, it inhibited cell attachment more than did treatment with the enzyme alone. The effects of all treatments on cell attachment were greater for the highly malignant than for the low malignant cells. With the latter cells, inhibition by lectin was seen only in the absence of serum, whereas the adhesion of highly malignant cells was affected in both the presence and the absence of serum. On their surface membrane the highly malignant cells express much more than do the low malignant cells of a glycoprotein that cross-reacts immunologically with laminin. The basement membrane glycoprotein laminin promotes cell attachment to collagen, and both glycoproteins contain terminal, non-reducing [alpha]--galactopyranosyl groups. Attachment of cells is a requirement for the formation of a metastasis, and thus the laminin-like molecule and the [alpha]--galactopyranosyl end groups (whether on the laminin-related moiety or on other cell surface molecules) may both be important for expression of the most malignant phenotype.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/24621/1/0000031.pd

From Cellular Characteristics to Disease Diagnosis: Uncovering Phenotypes with Supercells

Cell heterogeneity and the inherent complexity due to the interplay of multiple molecular processes within the cell pose difficult challenges for current single-cell biology. We introduce an approach that identifies a disease phenotype from multiparameter single-cell measurements, which is based on the concept of ‘‘supercell statistics’’, a single-cell-based averaging procedure followed by a machine learning classification scheme. We are able to assess the optimal tradeoff between the number of single cells averaged and the number of measurements needed to capture phenotypic differences between healthy and diseased patients, as well as between different diseases that are difficult to diagnose otherwise. We apply our approach to two kinds of single-cell datasets, addressing the diagnosis of a premature aging disorder using images of cell nuclei, as well as the phenotypes of two non-infectious uveitides (the ocular manifestations of Behc¸et’s disease and sarcoidosis) based on multicolor flow cytometry. In the former case, one nuclear shape measurement taken over a group of 30 cells is sufficient to classify samples as healthy or diseased, in agreement with usual laboratory practice. In the latter, our method is able to identify a minimal set of 5 markers that accurately predict Behc¸et’s disease and sarcoidosis. This is the first time that a quantitative phenotypic distinction between these two diseases has been achieved. To obtain this clear phenotypic signature, about one hundred CD8+ T cells need to be measured. Although the molecular markers identified have been reported to be important players in autoimmune disorders, this is the first report pointing out that CD8+ T cells can be used to distinguish two systemic inflammatory diseases. Beyond these specific cases, the approach proposed here is applicable to datasets generated by other kinds of state-of-the-art and forthcoming single-cell technologies, such as multidimensional mass cytometry, single-cell gene expression, and single-cell full genome sequencing techniques.Fil: Candia, Julian Marcelo. University of Maryland; Estados Unidos. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Física de Líquidos y Sistemas Biológicos. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Instituto de Física de Líquidos y Sistemas Biológicos; ArgentinaFil: Maunu, Ryan. University of Maryland; Estados UnidosFil: Driscoll, Meghan. University of Maryland; Estados UnidosFil: Biancotto, Angélique. National Institutes of Health; Estados UnidosFil: Dagur, Pradeep. National Institutes of Health; Estados UnidosFil: McCoy Jr., J Philip. National Institutes of Health; Estados UnidosFil: Nida Sen, H.. National Institutes of Health; Estados UnidosFil: Wei, Lai. National Institutes of Health; Estados UnidosFil: Maritan, Amos. Università di Padova; ItaliaFil: Cao, Kan. University of Maryland; Estados UnidosFil: Nussenblatt, Robert B. National Institutes of Health; Estados UnidosFil: Banavar, Jayanth R.. University of Maryland; Estados UnidosFil: Losert, Wolfgang. University of Maryland; Estados Unido

Laminin supports neurite outgrowth from explants of axotomized adult rat retinal neurons

The influence of laminin on neurite outgrowth from explants of adult rat retina and its distribution in normal and lesioned rat optic nerves were examined. Neurite outgrowth required the presence of laminin in the substratum, and as with a goldfish retinal explant system, was markedly stimulated by prior axotomy. Except for blood vessels and the nerve sheath, normal rat optic nerve was devoid of laminin immunoreactivity. Unlike results seen in the goldfish optic nerve, injury to the rat optic nerve induced no observable increase in laminin content or change in its distribution. The differences in the in vivo regenerative capacities of these two species may in part be related to the differences in their abilities to provide a proper substratum for axon regrowth.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/26113/1/0000189.pd

Binding of Griffonia simplicifolia I lectin to rat pulmonary alveolar macrophages and its use in purifying type II alveolar epithelial cells

We report that the isolectin Griffonia simplicifolia I-B4 isolated from G. simplicifolia seeds binds to rat alveolar macrophages present in frozen sections of lung tissue or bronchoalveolar lavage fluid. G. simplicifolia I-B4 does not bind to alveolar epithelial cells. We established that G. simplicifolia I-B4 binds to the macrophages via interaction with terminal [alpha]--galactopyranosyl residues present on these cells. This was substantiated by demonstrating that binding is inhibited either by the haptenic sugar [alpha]--galactopyranoside or by treating the cells with coffee bean [alpha]-galactosidase. Because murine laminin is known to contain terminal [alpha]--galactopyranosyl end-groups, and because we found that an anti-laminin antiserum binds to rat alveolar macrophages, we suspect that G. simplicifolia I-B4 may be binding to laminin present on the macrophages. To isolate alveolar type II epithelial cells from rat lungs, we developed a method that utilizes the lectin G. simplicifolia I. When proteinase-derived suspensions of pulmonary cells are incubated with G. simplicifolia I, the macrophages agglutinate and can be removed by filtration through nylon mesh. After incubating the resulting cellular suspension in tissue culture, the adherent cells are 94 +/- 2% (S.D.) type II cells. When compared to cells isolated by repeated differential adherence, the lectin-prepared type II cells have similar morphology and staining characteristics, form domes in monolayers and incorporate similar amounts of palmitate into disaturated phosphatidylcholine. We believe that the procedure outlined in this report provides a simple and effective method to isolate type II alveolar epithelial cells from rat lungs.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/26290/1/0000375.pd

Isolation and purification of a squamous cell carcinoma of the head and neck-associated antigen identified by autologous antibody

We have previously shown that detection of autologous antibody activity to squamous cell carninoma of the head and neck many be augmented by dissociation in immune complexes. Western blot analysis with autologous antibody has identified a 60 kDa squamous cell carcinoma of the head and neck-associated antigen in spent media and immune complex-dissociated serum ultrafiltrate not recognized by normal human area. Antigen-containing fractions of spent media were eluted from anion exchange columns immediately after serum albumin indicating that the antigen has an acidic PI Triticum vulgaris column. A single 60 kDa band was detected by silver stain and Western blot in antigen-containing fractions eluted following lectin affinity chromotography and SDS-PAGE. Final concentration of the antigen was determined to be 1 [mu]m/ml of protein with relative activity increased 1600 x over unfractionated spent media. We conclude that a squamous cell carcinoma of the head and neck-associated antigen, detected by autologous antibody, is an acidic kDa glycoprotein.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/30831/1/0000493.pd

A transient homotypic interaction model for the influenza A virus NS1 protein effector domain

Influenza A virus NS1 protein is a multifunctional virulence factor consisting of an RNA binding domain (RBD), a short linker, an effector domain (ED), and a C-terminal 'tail'. Although poorly understood, NS1 multimerization may autoregulate its actions. While RBD dimerization seems functionally conserved, two possible apo ED dimers have been proposed (helix-helix and strand-strand). Here, we analyze all available RBD, ED, and full-length NS1 structures, including four novel crystal structures obtained using EDs from divergent human and avian viruses, as well as two forms of a monomeric ED mutant. The data reveal the helix-helix interface as the only strictly conserved ED homodimeric contact. Furthermore, a mutant NS1 unable to form the helix-helix dimer is compromised in its ability to bind dsRNA efficiently, implying that ED multimerization influences RBD activity. Our bioinformatical work also suggests that the helix-helix interface is variable and transient, thereby allowing two ED monomers to twist relative to one another and possibly separate. In this regard, we found a mAb that recognizes NS1 via a residue completely buried within the ED helix-helix interface, and which may help highlight potential different conformational populations of NS1 (putatively termed 'helix-closed' and 'helix-open') in virus-infected cells. 'Helix-closed' conformations appear to enhance dsRNA binding, and 'helix-open' conformations allow otherwise inaccessible interactions with host factors. Our data support a new model of NS1 regulation in which the RBD remains dimeric throughout infection, while the ED switches between several quaternary states in order to expand its functional space. Such a concept may be applicable to other small multifunctional proteins

Exploration of Victoria Crater by the Mars Rover Opportunity

The Mars rover Opportunity has explored Victoria crater, a ~750-meter eroded impact crater formed in sulfate-rich sedimentary rocks. Impact-related stratigraphy is preserved in the crater walls, and meteoritic debris is present near the crater rim. The size of hematite-rich concretions decreases up-section, documenting variation in the intensity of groundwater processes. Layering in the crater walls preserves evidence of ancient wind-blown dunes. Compositional variations with depth mimic those ~6 kilometers to the north and demonstrate that water-induced alteration at Meridiani Planum was regional in scope.Organismic and Evolutionary Biolog

Nestedness of Ectoparasite-Vertebrate Host Networks

Determining the structure of ectoparasite-host networks will enable disease ecologists to better understand and predict the spread of vector-borne diseases. If these networks have consistent properties, then studying the structure of well-understood networks could lead to extrapolation of these properties to others, including those that support emerging pathogens. Borrowing a quantitative measure of network structure from studies of mutualistic relationships between plants and their pollinators, we analyzed 29 ectoparasite-vertebrate host networks—including three derived from molecular bloodmeal analysis of mosquito feeding patterns—using measures of nestedness to identify non-random interactions among species. We found significant nestedness in ectoparasite-vertebrate host lists for habitats ranging from tropical rainforests to polar environments. These networks showed non-random patterns of nesting, and did not differ significantly from published estimates of nestedness from mutualistic networks. Mutualistic and antagonistic networks appear to be organized similarly, with generalized ectoparasites interacting with hosts that attract many ectoparasites and more specialized ectoparasites usually interacting with these same “generalized” hosts. This finding has implications for understanding the network dynamics of vector-born pathogens. We suggest that nestedness (rather than random ectoparasite-host associations) can allow rapid transfer of pathogens throughout a network, and expand upon such concepts as the dilution effect, bridge vectors, and host switching in the context of nested ectoparasite-vertebrate host networks

The James Webb Space Telescope Mission

Twenty-six years ago a small committee report, building on earlier studies, expounded a compelling and poetic vision for the future of astronomy, calling for an infrared-optimized space telescope with an aperture of at least $4m$. With the support of their governments in the US, Europe, and Canada, 20,000 people realized that vision as the $6.5m$ James Webb Space Telescope. A generation of astronomers will celebrate their accomplishments for the life of the mission, potentially as long as 20 years, and beyond. This report and the scientific discoveries that follow are extended thank-you notes to the 20,000 team members. The telescope is working perfectly, with much better image quality than expected. In this and accompanying papers, we give a brief history, describe the observatory, outline its objectives and current observing program, and discuss the inventions and people who made it possible. We cite detailed reports on the design and the measured performance on orbit.Comment: Accepted by PASP for the special issue on The James Webb Space Telescope Overview, 29 pages, 4 figure

Climate Change, Coral Reef Ecosystems, and Management Options for Marine Protected Areas

Marine protected areas (MPAs) provide place-based management of marine ecosystems through various degrees and types of protective actions. Habitats such as coral reefs are especially susceptible to degradation resulting from climate change, as evidenced by mass bleaching events over the past two decades. Marine ecosystems are being altered by direct effects of climate change including ocean warming, ocean acidification, rising sea level, changing circulation patterns, increasing severity of storms, and changing freshwater influxes. As impacts of climate change strengthen they may exacerbate effects of existing stressors and require new or modified management approaches; MPA networks are generally accepted as an improvement over individual MPAs to address multiple threats to the marine environment. While MPA networks are considered a potentially effective management approach for conserving marine biodiversity, they should be established in conjunction with other management strategies, such as fisheries regulations and reductions of nutrients and other forms of land-based pollution. Information about interactions between climate change and more “traditional” stressors is limited. MPA managers are faced with high levels of uncertainty about likely outcomes of management actions because climate change impacts have strong interactions with existing stressors, such as land-based sources of pollution, overfishing and destructive fishing practices, invasive species, and diseases. Management options include ameliorating existing stressors, protecting potentially resilient areas, developing networks of MPAs, and integrating climate change into MPA planning, management, and evaluation