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A foot in the door.

Powell K — Thu, 11 Mar 2010 00:25:08 +0000

A foot in the door.

Nature. 2010 Feb 4;463(7281):696-7

Authors: Powell K

PMID: 20213860 [PubMed - in process]

[...]

Dosage Sensitivity Shapes the Evolution of Copy-Number Varied Regions

Nicolas Guex et al. — Wed, 10 Mar 2010 08:00:00 +0000

Dosage sensitivity is an important evolutionary force which impacts on gene dispensability and duplicability. The newly available data on human copy-number variation (CNV) allow an analysis of the most recent and ongoing evolution. Provided that heterozygous gene deletions and duplications actually change gene dosage, we expect to observe negative selection against CNVs encompassing dosage sensitive genes. In this study, we make use of several sources of population genetic data to identify selection on structural variations of dosage sensitive genes. We show that CNVs can directly affect expression levels of contained genes. We find that genes encoding members of protein complexes exhibit limited expression variation and overlap significantly with a manually derived set of dosage sensitive genes. We show that complexes and other dosage sensitive genes are underrepresented in CNV regions, with a particular bias against frequent variations and duplications. These results suggest that dosage sensitivity is a significant force of negative selection on regions of copy-number variation.

FastTree 2 – Approximately Maximum-Likelihood Trees for Large Alignments

Nicolas Guex et al. — Wed, 10 Mar 2010 08:00:00 +0000

Background

We recently described FastTree, a tool for inferring phylogenies for alignments with up to hundreds of thousands of sequences. Here, we describe improvements to FastTree that improve its accuracy without sacrificing scalability.

Methodology/Principal Findings

Where FastTree 1 used nearest-neighbor interchanges (NNIs) and the minimum-evolution criterion to improve the tree, FastTree 2 adds minimum-evolution subtree-pruning-regrafting (SPRs) and maximum-likelihood NNIs. FastTree 2 uses heuristics to restrict the search for better trees and estimates a rate of evolution for each site (the “CAT” approximation). Nevertheless, for both simulated and genuine alignments, FastTree 2 is slightly more accurate than a standard implementation of maximum-likelihood NNIs (PhyML 3 with default settings). Although FastTree 2 is not quite as accurate as methods that use maximum-likelihood SPRs, most of the splits that disagree are poorly supported, and for large alignments, FastTree 2 is 100–1,000 times faster. FastTree 2 inferred a topology and likelihood-based local support values for 237,882 distinct 16S ribosomal RNAs on a desktop computer in 22 hours and 5.8 gigabytes of memory.

Conclusions/Significance

FastTree 2 allows the inference of maximum-likelihood phylogenies for huge alignments. FastTree 2 is freely available at http://www.microbesonline.org/fasttree.

The Association between Proximity to Animal Feeding Operations and Community Health: A Systematic Review

Nicolas Guex et al. — Wed, 10 Mar 2010 08:00:00 +0000

Background

A systematic review was conducted for the association between animal feeding operations (AFOs) and the health of individuals living near AFOs.

Methodology/Principal Findings

The review was restricted to studies reporting respiratory, gastrointestinal and mental health outcomes in individuals living near AFOs in North America, European Union, United Kingdom, and Scandinavia. From June to September 2008 searches were conducted in PUBMED, CAB, Web-of-Science, and Agricola with no restrictions. Hand searching of narrative reviews was also used. Two reviewers independently evaluated the role of chance, confounding, information, selection and analytic bias on the study outcome. Nine relevant studies were identified. The studies were heterogeneous with respect to outcomes and exposures assessed. Few studies reported an association between surrogate clinical outcomes and AFO proximity. A negative association was reported when odor was the measure of exposure to AFOs and self-reported disease, the measure of outcome. There was evidence of an association between self-reported disease and proximity to AFO in individuals annoyed by AFO odor.

Conclusions/Significance

There was inconsistent evidence of a weak association between self-reported disease in people with allergies or familial history of allergies. No consistent dose response relationship between exposure and disease was observable.

An Extended Nomenclature for Mammalian V-ATPase Subunit Genes and Splice Variants

Nicolas Guex et al. — Wed, 10 Mar 2010 08:00:00 +0000

The vacuolar-type H⁺-ATPase (V-ATPase) is a multisubunit proton pump that is involved in both intra- and extracellular acidification processes throughout the body. Multiple homologs and splice variants of V-ATPase subunits are thought to explain its varied spatial and temporal expression pattern in different cell types. Recently subunit nomenclature was standardized with a total of 22 subunit variants identified. However this standardization did not accommodate the existence of splice variants and is therefore incomplete. Thus, we propose here an extension of subunit nomenclature along with a literature and sequence database scan for additional V-ATPase subunits. An additional 17 variants were pulled from a literature search while 4 uncharacterized potential subunit variants were found in sequence databases. These findings have been integrated with the current V-ATPase knowledge base to create a new V-ATPase subunit catalogue. It is envisioned this catalogue will form a new platform on which future studies into tissue- and organelle-specific V-ATPase expression, localization and function can be based.

Disruption of Astrocyte STAT3 Signaling Decreases Mitochondrial Function and Increases Oxidative Stress In Vitro

Nicolas Guex et al. — Wed, 10 Mar 2010 08:00:00 +0000

Background

Astrocytes exert a wide variety of functions in health and disease and respond to a wide range of signaling pathways, including members of the Janus-kinase signal transducers and activators of transcription (Jak-STAT) family. We have recently shown that STAT3 is an important regulator of astrocyte reactivity after spinal cord injury in vivo [1].

Methodology/Principal Findings

Here, we used both a conditional gene deletion strategy that targets the deletion of STAT3 selectively to astrocytes (STAT3-CKO), and a pharmacological inhibitor of JAK-2, AG490, in cultured astrocytes in vitro, to investigate potential functions and molecules influenced by STAT3 signaling in relation to mitochondrial function and oxidative stress. Our findings show that the absence of STAT3 signaling in astrocytes leads to (i) increased production of superoxide anion and other reactive oxygen species and decreased level of glutathione, (ii) decreased mitochondrial membrane potential and decreased ATP production, and (iii) decreased rate of cell proliferation. Many of the differences observed in STAT3-CKO astrocytes were distinctly altered by exposure to rotenone, suggesting a role for complex I of the mitochondrial electron transport chain. Gene expression microarray studies identified numerous changes in STAT3-CKO cells that may have contributed to the identified deficits in cell function.

Conclusions/Significance

Taken together, these STAT3-dependent alterations in cell function and gene expression have relevance to both reactive gliosis and to the support and protection of surrounding cells in neural tissue.

Does Tropical Forest Fragmentation Increase Long-Term Variability of Butterfly Communities?

Nicolas Guex et al. — Wed, 10 Mar 2010 08:00:00 +0000

Habitat fragmentation is a major driver of biodiversity loss. Yet, the overall effects of fragmentation on biodiversity may be obscured by differences in responses among species. These opposing responses to fragmentation may be manifest in higher variability in species richness and abundance (termed hyperdynamism), and in predictable changes in community composition. We tested whether forest fragmentation causes long-term hyperdynamism in butterfly communities, a taxon that naturally displays large variations in species richness and community composition. Using a dataset from an experimentally fragmented landscape in the central Amazon that spanned 11 years, we evaluated the effect of fragmentation on changes in species richness and community composition through time. Overall, adjusted species richness (adjusted for survey duration) did not differ between fragmented forest and intact forest. However, spatial and temporal variation of adjusted species richness was significantly higher in fragmented forests relative to intact forest. This variation was associated with changes in butterfly community composition, specifically lower proportions of understory shade species and higher proportions of edge species in fragmented forest. Analysis of rarefied species richness, estimated using indices of butterfly abundance, showed no differences between fragmented and intact forest plots in spatial or temporal variation. These results do not contradict the results from adjusted species richness, but rather suggest that higher variability in butterfly adjusted species richness may be explained by changes in butterfly abundance. Combined, these results indicate that butterfly communities in fragmented tropical forests are more variable than in intact forest, and that the natural variability of butterflies was not a buffer against the effects of fragmentation on community dynamics.

A PubMed-Wide Associational Study of Infectious Diseases

Nicolas Guex et al. — Wed, 10 Mar 2010 08:00:00 +0000

Background

Computational discovery is playing an ever-greater role in supporting the processes of knowledge synthesis. A significant proportion of the more than 18 million manuscripts indexed in the PubMed database describe infectious disease syndromes and various infectious agents. This study is the first attempt to integrate online repositories of text-based publications and microbial genome databases in order to explore the dynamics of relationships between pathogens and infectious diseases.

Methodology/Principal Findings

Herein we demonstrate how the knowledge space of infectious diseases can be computationally represented and quantified, and tracked over time. The knowledge space is explored by mapping of the infectious disease literature, looking at dynamics of literature deposition, zooming in from pathogen to genome level and searching for new associations. Syndromic signatures for different pathogens can be created to enable a new and clinically focussed reclassification of the microbial world. Examples of syndrome and pathogen networks illustrate how multilevel network representations of the relationships between infectious syndromes, pathogens and pathogen genomes can illuminate unexpected biological similarities in disease pathogenesis and epidemiology.

Conclusions/Significance

This new approach based on text and data mining can support the discovery of previously hidden associations between diseases and microbial pathogens, clinically relevant reclassification of pathogenic microorganisms and accelerate the translational research enterprise.

Embryos in the Fast Lane: High-Temperature Heart Rates of Turtles Decline After Hatching

Nicolas Guex et al. — Wed, 10 Mar 2010 08:00:00 +0000

In ectotherms such as turtles, the relationship between cardiovascular function and temperature may be subject to different selective pressures in different life-history stages. Because embryos benefit by developing as rapidly as possible, and can “afford” to expend energy to do so (because they have access to the yolk for nutrition), they benefit from rapid heart (and thus, developmental) rates. In contrast, hatchlings do not have a guaranteed food supply, and maximal growth rates may not enhance fitness—and so, we might expect a lower heart rate, especially at high temperatures where metabolic costs are greatest. Our data on two species of emydid turtles, Chrysemys picta, and Graptemys pseudogeographica kohnii, support these predictions. Heart rates of embryos and hatchlings were similar at low temperatures, but heart rates at higher temperatures were much greater before than after hatching.

Polypyrimidine Tract Binding Protein Functions as a Negative Regulator of Feline Calicivirus Translation

Nicolas Guex et al. — Wed, 10 Mar 2010 08:00:00 +0000

Background

Positive strand RNA viruses rely heavily on host cell RNA binding proteins for various aspects of their life cycle. Such proteins interact with sequences usually present at the 5′ or 3′ extremities of the viral RNA genome, to regulate viral translation and/or replication. We have previously reported that the well characterized host RNA binding protein polypyrimidine tract binding protein (PTB) interacts with the 5′end of the feline calicivirus (FCV) genomic and subgenomic RNAs, playing a role in the FCV life cycle.

Principal Findings

We have demonstrated that PTB interacts with at least two binding sites within the 5′end of the FCV genome. In vitro translation indicated that PTB may function as a negative regulator of FCV translation and this was subsequently confirmed as the translation of the viral subgenomic RNA in PTB siRNA treated cells was stimulated under conditions in which RNA replication could not occur. We also observed that PTB redistributes from the nucleus to the cytoplasm during FCV infection, partially localizing to viral replication complexes, suggesting that PTB binding may be involved in the switch from translation to replication. Reverse genetics studies demonstrated that synonymous mutations in the PTB binding sites result in a cell-type specific defect in FCV replication.

Conclusions

Our data indicates that PTB may function to negatively regulate FCV translation initiation. To reconcile this with efficient virus replication in cells, we propose a putative model for the function of PTB in the FCV life cycle. It is possible that during the early stages of infection, viral RNA is translated in the absence of PTB, however, as the levels of viral proteins increase, the nuclear-cytoplasmic shuttling of PTB is altered, increasing the cytoplasmic levels of PTB, inhibiting viral translation. Whether PTB acts directly to repress translation initiation or via the recruitment of other factors remains to be determined but this may contribute to the stimulation of viral RNA replication via clearance of ribosomes from viral RNA.

Modeling Selective Pressures on Phytoplankton in the Global Ocean

Nicolas Guex et al. — Wed, 10 Mar 2010 08:00:00 +0000

Our view of marine microbes is transforming, as culture-independent methods facilitate rapid characterization of microbial diversity. It is difficult to assimilate this information into our understanding of marine microbe ecology and evolution, because their distributions, traits, and genomes are shaped by forces that are complex and dynamic. Here we incorporate diverse forces—physical, biogeochemical, ecological, and mutational—into a global ocean model to study selective pressures on a simple trait in a widely distributed lineage of picophytoplankton: the nitrogen use abilities of Synechococcus and Prochlorococcus cyanobacteria. Some Prochlorococcus ecotypes have lost the ability to use nitrate, whereas their close relatives, marine Synechococcus, typically retain it. We impose mutations for the loss of nitrogen use abilities in modeled picophytoplankton, and ask: in which parts of the ocean are mutants most disadvantaged by losing the ability to use nitrate, and in which parts are they least disadvantaged? Our model predicts that this selective disadvantage is smallest for picophytoplankton that live in tropical regions where Prochlorococcus are abundant in the real ocean. Conversely, the selective disadvantage of losing the ability to use nitrate is larger for modeled picophytoplankton that live at higher latitudes, where Synechococcus are abundant. In regions where we expect Prochlorococcus and Synechococcus populations to cycle seasonally in the real ocean, we find that model ecotypes with seasonal population dynamics similar to Prochlorococcus are less disadvantaged by losing the ability to use nitrate than model ecotypes with seasonal population dynamics similar to Synechococcus. The model predictions for the selective advantage associated with nitrate use are broadly consistent with the distribution of this ability among marine picocyanobacteria, and at finer scales, can provide insights into interactions between temporally varying ocean processes and selective pressures that may be difficult or impossible to study by other means. More generally, and perhaps more importantly, this study introduces an approach for testing hypotheses about the processes that underlie genetic variation among marine microbes, embedded in the dynamic physical, chemical, and biological forces that generate and shape this diversity.

How Accurate and Robust Are the Phylogenetic Estimates of Austronesian Language Relationships?

Nicolas Guex et al. — Wed, 10 Mar 2010 08:00:00 +0000

We recently used computational phylogenetic methods on lexical data to test between two scenarios for the peopling of the Pacific. Our analyses of lexical data supported a pulse-pause scenario of Pacific settlement in which the Austronesian speakers originated in Taiwan around 5,200 years ago and rapidly spread through the Pacific in a series of expansion pulses and settlement pauses. We claimed that there was high congruence between traditional language subgroups and those observed in the language phylogenies, and that the estimated age of the Austronesian expansion at 5,200 years ago was consistent with the archaeological evidence. However, the congruence between the language phylogenies and the evidence from historical linguistics was not quantitatively assessed using tree comparison metrics. The robustness of the divergence time estimates to different calibration points was also not investigated exhaustively. Here we address these limitations by using a systematic tree comparison metric to calculate the similarity between the Bayesian phylogenetic trees and the subgroups proposed by historical linguistics, and by re-estimating the age of the Austronesian expansion using only the most robust calibrations. The results show that the Austronesian language phylogenies are highly congruent with the traditional subgroupings, and the date estimates are robust even when calculated using a restricted set of historical calibrations.

60 kD Ro and nRNP A Frequently Initiate Human Lupus Autoimmunity

Nicolas Guex et al. — Wed, 10 Mar 2010 08:00:00 +0000

Systemic lupus erythematosus (SLE) is a clinically heterogeneous, humoral autoimmune disorder. The unifying feature among SLE patients is the production of large quantities of autoantibodies. Serum samples from 129 patients collected before the onset of SLE and while in the United States military were evaluated for early pre-clinical serologic events. The first available positive serum sample frequently already contained multiple autoantibody specificities (65%). However, in 34 SLE patients the earliest pre-clinical serum sample positive for any detectable common autoantibody bound only a single autoantigen, most commonly 60 kD Ro (29%), nRNP A (24%), anti-phospholipids (18%) or rheumatoid factor (15%). We identified several recurrent patterns of autoantibody onset using these pre-diagnostic samples. In the serum samples available, anti-nRNP A appeared before or simultaneously with anti-nRNP 70 K in 96% of the patients who had both autoantibodies at diagnosis. Anti-60 kD Ro antibodies appeared before or simultaneously with anti-La (98%) or anti-52 kD Ro (95%). The autoantibody response in SLE patients begins simply, often binding a single specific autoantigen years before disease onset, followed by epitope spreading to additional autoantigenic specificities that are accrued in recurring patterns.

Normal Bias in the Direction of Fetal Rotation Depends on Blastomere Composition during Early Cleavage in the Mouse

Nicolas Guex et al. — Wed, 10 Mar 2010 08:00:00 +0000

Interest in establishing the basis of left/right asymmetry during embryogenesis has burgeoned in recent years. Relevant studies in mammals, focused largely on the mouse, have revealed involvement of a variety of genes that are common to the process in other animals. In the mouse, lateral differences in gene expression are first evident late in gastrulation when directional rotation of nodal cilia has been implicated in effecting the normally very strong bias in handedness. Reconstructing cleavage stages with correspondingly positioned blastomeres from appropriate numbers of conceptuses with similar division planes provides a way of testing whether they differ in potency without the confounding effects of reduced cell number. In a study using this strategy, 4-cell stage conceptuses reconstructed from blastomeres produced by equatorial as opposed to meridional second cleavage were found to be compromised in their ability to support normal development. Here, in more refined reconstructions undertaken at both the 4- and 8-cell stage, no significant impairment of development to the 9^th or 12^th day of gestation was found for products of equatorial second cleavage or their 8-cell stage progeny. Most surprisingly, however, a significant increase in reversal of the direction of axial rotation was found specifically among fetuses developing from conceptuses reconstructed from 8-cell stage progeny of products of equatorial second cleavage. Hence, manipulations during early cleavage some 6 days before fetal asymmetries are first evident can perturb the normally very strong bias in specification of a facet of left-right asymmetry.

Reactivation of Latent Tuberculosis in Cynomolgus Macaques Infected with SIV Is Associated with Early Peripheral T Cell Depletion and Not Virus Load

Nicolas Guex et al. — Wed, 10 Mar 2010 08:00:00 +0000

HIV-infected individuals with latent Mycobacterium tuberculosis (Mtb) infection are at significantly greater risk of reactivation tuberculosis (TB) than HIV-negative individuals with latent TB, even while CD4 T cell numbers are well preserved. Factors underlying high rates of reactivation are poorly understood and investigative tools are limited. We used cynomolgus macaques with latent TB co-infected with SIVmac251 to develop the first animal model of reactivated TB in HIV-infected humans to better explore these factors. All latent animals developed reactivated TB following SIV infection, with a variable time to reactivation (up to 11 months post-SIV). Reactivation was independent of virus load but correlated with depletion of peripheral T cells during acute SIV infection. Animals experiencing reactivation early after SIV infection (<17 weeks) had fewer CD4 T cells in the periphery and airways than animals reactivating in later phases of SIV infection. Co-infected animals had fewer T cells in involved lungs than SIV-negative animals with active TB despite similar T cell numbers in draining lymph nodes. Granulomas from these animals demonstrated histopathologic characteristics consistent with a chronically active disease process. These results suggest initial T cell depletion may strongly influence outcomes of HIV-Mtb co-infection.

Evolutionary Divergence in Brain Size between Migratory and Resident Birds

Nicolas Guex et al. — Wed, 10 Mar 2010 08:00:00 +0000

Despite important recent progress in our understanding of brain evolution, controversy remains regarding the evolutionary forces that have driven its enormous diversification in size. Here, we report that in passerine birds, migratory species tend to have brains that are substantially smaller (relative to body size) than those of resident species, confirming and generalizing previous studies. Phylogenetic reconstructions based on Bayesian Markov chain methods suggest an evolutionary scenario in which some large brained tropical passerines that invaded more seasonal regions evolved migratory behavior and migration itself selected for smaller brain size. Selection for smaller brains in migratory birds may arise from the energetic and developmental costs associated with a highly mobile life cycle, a possibility that is supported by a path analysis. Nevertheless, an important fraction (over 68%) of the correlation between brain mass and migratory distance comes from a direct effect of migration on brain size, perhaps reflecting costs associated with cognitive functions that have become less necessary in migratory species. Overall, our results highlight the importance of retrospective analyses in identifying selective pressures that have shaped brain evolution, and indicate that when it comes to the brain, larger is not always better.

A Direct Interaction with NEDD1 Regulates γ-Tubulin Recruitment to the Centrosome

Nicolas Guex et al. — Wed, 10 Mar 2010 08:00:00 +0000

The centrosome is the primary microtubule organizing centre of the cell. γ-tubulin is a core component of the centrosome and is required for microtubule nucleation and centrosome function. The recruitment of γ-tubulin to centrosomes is mediated by its interaction with NEDD1, a WD40-repeat containing protein. Here we demonstrate that NEDD1 is likely to be oligomeric in vivo and binds directly to γ-tubulin through a small region of just 62 residues at the carboxyl-terminus of the protein. This carboxyl-terminal domain that binds γ-tubulin has a helical structure and is a stable tetramer in solution. Mutation of residues in NEDD1 that disrupt binding to γ-tubulin result in a mis-localization of γ-tubulin away from the centrosome. Hence, this study defines the binding site on NEDD1 that is required for its interaction with γ-tubulin, and shows that this interaction is required for the correct localization of γ-tubulin.

Morphological and Genomic Characterization of Filobasidiella depauperata: A Homothallic Sibling Species of the Pathogenic Cryptococcus Species Complex

Nicolas Guex et al. — Wed, 10 Mar 2010 08:00:00 +0000

The fungal species Cryptococcus neoformans and Cryptococcus gattii cause respiratory and neurological disease in animals and humans following inhalation of basidiospores or desiccated yeast cells from the environment. Sexual reproduction in C. neoformans and C. gattii is controlled by a bipolar system in which a single mating type locus (MAT) specifies compatibility. These two species are dimorphic, growing as yeast in the asexual stage, and producing hyphae, basidia, and basidiospores during the sexual stage. In contrast, Filobasidiella depauperata, one of the closest related species, grows exclusively as hyphae and it is found in association with decaying insects. Examination of two available strains of F. depauperata showed that the life cycle of this fungal species shares features associated with the unisexual or same-sex mating cycle in C. neoformans. Therefore, F. depauperata may represent a homothallic and possibly an obligately sexual fungal species. RAPD genotyping of 39 randomly isolated progeny from isolate CBS7855 revealed a new genotype pattern in one of the isolated basidiospores progeny, therefore suggesting that the homothallic cycle in F. depauperata could lead to the emergence of new genotypes. Phylogenetic analyses of genes linked to MAT in C. neoformans indicated that two of these genes in F. depauperata, MYO2 and STE20, appear to form a monophyletic clade with the MATa alleles of C. neoformans and C. gattii, and thus these genes may have been recruited to the MAT locus before F. depauperata diverged. Furthermore, the ancestral MATa locus may have undergone accelerated evolution prior to the divergence of the pathogenic Cryptococcus species since several of the genes linked to the MATa locus appear to have a higher number of changes and substitutions than their MATα counterparts. Synteny analyses between C. neoformans and F. depauperata showed that genomic regions on other chromosomes displayed conserved gene order. In contrast, the genes linked to the MAT locus of C. neoformans showed a higher number of chromosomal translocations in the genome of F. depauperata. We therefore propose that chromosomal rearrangements appear to be a major force driving speciation and sexual divergence in these closely related pathogenic and saprobic species.

Plasmodium vivax: Induction of CD4+CD25+FoxP3+ Regulatory T Cells during Infection Are Directly Associated with Level of Circulating Parasites

Nicolas Guex et al. — Wed, 10 Mar 2010 08:00:00 +0000

Circulation CD4⁺CD25⁺FoxP3⁺ regulatory T cells (Tregs) have been associated with the delicate balancing between control of overwhelming acute malaria infection and prevention of immune pathology due to disproportionate inflammatory responses to erythrocytic stage of the parasite. While the role of Tregs has been well-documented in murine models and P. falciparum infection, the phenotype and function of Tregs in P. vivax infection is still poorly characterized. In the current study, we demonstrated that patients with acute P. vivax infection presented a significant augmentation of circulating Tregs producing anti-inflammatory (IL-10 and TGF-β) as well as pro-inflammatory (IFN-γ, IL-17) cytokines, which was further positively correlated with parasite burden. Surface expression of GITR molecule and intracellular expression of CTLA-4 were significantly upregulated in Tregs from infected donors, presenting also a positive association between either absolute numbers of CD4⁺CD25⁺FoxP3⁺GITR⁺ or CD4⁺CD25⁺FoxP3⁺CTLA-4⁺ and parasite load. Finally, we demonstrate a suppressive effect of Treg cells in specific T cell proliferative responses of P. vivax infected subjects after antigen stimulation with Pv-AMA-1. Our findings indicate that malaria vivax infection lead to an increased number of activated Treg cells that are highly associated with parasite load, which probably exert an important contribution to the modulation of immune responses during P. vivax infection.

The Most Recently Discovered Carbonic Anhydrase, CA XV, Is Expressed in the Thick Ascending Limb of Henle and in the Collecting Ducts of Mouse Kidney

Nicolas Guex et al. — Wed, 10 Mar 2010 08:00:00 +0000

Background

Carbonic anhydrases (CAs) are key enzymes for physiological pH regulation, including the process of urine acidification. Previous studies have identified seven cytosolic or membrane-bound CA isozymes in the kidney. Recently, we showed by in situ hybridization that the mRNA for the most novel CA isozyme, CA XV, is present in the renal cortex. CA XV is a unique isozyme among mammalian CAs, because it has become a pseudogene in primates even though expressed in several other species.

Methodology/Principal Findings

In the present study, we raised a polyclonal antibody against recombinant mouse CA XV that was produced in a baculovirus/insect cell expression system, and the antibody was used for immunohistochemical analysis in different mouse tissues. Positive immunoreactions were found only in the kidney, where the enzyme showed a very limited distribution pattern. Parallel immunostaining experiments with several other anti-CA sera indicated that CA XV is mainly expressed in the thick ascending limb of Henle and collecting ducts, and the reactions were most prominent in the cortex and outer medulla.

Conclusion/Significance

Although other studies have proposed a role for CA XV in cell proliferation, its tightly limited distribution may point to a specialized function in the regulation of acid-base homeostasis.

Unique Biological Properties of Catalytic Domain Directed Human Anti-CAIX Antibodies Discovered through Phage-Display Technology

Nicolas Guex et al. — Wed, 10 Mar 2010 08:00:00 +0000

Carbonic anhydrase IX (CAIX, gene G250/MN-encoded transmembrane protein) is highly expressed in various human epithelial tumors such as renal clear cell carcinoma (RCC), but absent from the corresponding normal tissues. Besides the CA signal transduction activity, CAIX may serve as a biomarker in early stages of oncogenesis and also as a reliable marker of hypoxia, which is associated with tumor resistance to chemotherapy and radiotherapy. Although results from preclinical and clinical studies have shown CAIX as a promising target for detection and therapy for RCC, only a limited number of murine monoclonal antibodies (mAbs) and one humanized mAb are available for clinical testing and development. In this study, paramagnetic proteoliposomes of CAIX (CAIX-PMPLs) were constructed and used for anti-CAIX antibody selection from our 27 billion human single-chain antibody (scFv) phage display libraries. A panel of thirteen human scFvs that specifically recognize CAIX expressed on cell surface was identified, epitope mapped primarily to the CA domain, and affinity-binding constants (KD) determined. These human anti-CAIX mAbs are diverse in their functions including induction of surface CAIX internalization into endosomes and inhibition of the carbonic anhydrase activity, the latter being a unique feature that has not been previously reported for anti-CAIX antibodies. These human anti-CAIX antibodies are important reagents for development of new immunotherapies and diagnostic tools for RCC treatment as well as extending our knowledge on the basic structure-function relationships of the CAIX molecule.

Task Shifting Routine Inpatient Pediatric HIV Testing Improves Program Outcomes in Urban Malawi: A Retrospective Observational Study

Nicolas Guex et al. — Wed, 10 Mar 2010 08:00:00 +0000

Background

This study evaluated two models of routine HIV testing of hospitalized children in a high HIV-prevalence resource-constrained African setting. Both models incorporated “task shifting,” or the allocation of tasks to the least-costly, capable health worker.

Methods and Findings

Two models were piloted for three months each within the pediatric department of a referral hospital in Lilongwe, Malawi between January 1 and June 30, 2008. Model 1 utilized lay counselors for HIV testing instead of nurses and clinicians. Model 2 further shifted program flow and advocacy responsibilities from counselors to volunteer parents of HIV-infected children, called “patient escorts.” A retrospective review of data from 6318 hospitalized children offered HIV testing between January-December 2008 was conducted. The pilot quarters of Model 1 and Model 2 were compared, with Model 2 selected to continue after the pilot period. There was a 2-fold increase in patients offered HIV testing with Model 2 compared with Model 1 (43.1% vs 19.9%, p<0.001). Furthermore, patients in Model 2 were younger (17.3 vs 26.7 months, p<0.001) and tested sooner after admission (1.77 vs 2.44 days, p<0.001). There were no differences in test acceptance or enrollment rates into HIV care, and the program trends continued 6 months after the pilot period. Overall, 10244 HIV antibody tests (4779 maternal; 5465 child) and 453 DNA-PCR tests were completed, with 97.8% accepting testing. 19.6% of all mothers (n = 1112) and 8.5% of all children (n = 525) were HIV-infected. Furthermore, 6.5% of children were HIV-exposed (n = 405). Cumulatively, 72.9% (n = 678) of eligible children were evaluated in the hospital by a HIV-trained clinician, and 68.3% (n = 387) successfully enrolled into outpatient HIV care.

Conclusions/Significance

The strategy presented here, task shifting from lay counselors alone to lay counselors and patient escorts, greatly improved program outcomes while only marginally increasing operational costs. The wider implementation of this strategy could accelerate pediatric HIV care access in high-prevalence settings.

Investigating a New Generation of Ribozymes in Order to Target HCV

Nicolas Guex et al. — Wed, 10 Mar 2010 08:00:00 +0000

For a long time nucleic acid-based approaches directed towards controlling the propagation of Hepatitis C Virus (HCV) have been considered to possess high potential. Towards this end, ribozymes (i.e. RNA enzymes) that specifically recognize and subsequently catalyze the cleavage of their RNA substrate present an attractive molecular tool. Here, the unique properties of a new generation of ribozymes are taken advantage of in order to develop an efficient and durable ribozyme-based technology with which to target HCV (+) RNA strands. These ribozymes resulted from the coupling of a specific on/off adaptor (SOFA) to the ribozyme domain derived from the Hepatitis Delta Virus (HDV). The former switches cleavage activity “on” solely in the presence of the desired RNA substrate, while the latter was the first catalytic RNA reported to function naturally in human cells, specifically in hepatocytes. In order to maximize the chances for success, a step-by-step approach was used for both the design and the selection of the ribozymes. This approach included the use of both bioinformatics and biochemical methods for the identification of the sites possessing the greatest potential for targeting, and the subsequent in vitro testing of the cleavage activities of the corresponding SOFA-HDV ribozymes. These efforts led to a significant improvement in the ribozymes’ designs. The ability of the resulting SOFA-HDV ribozymes to inhibit HCV replication was further examined using a luciferase-based replicon. Although some of the ribozymes exhibited high levels of cleavage activity in vitro, none appears to be a potential long term inhibitor in cellulo. Analysis of recent discoveries in the cellular biology of HCV might explain this failure, as well as provide some ideas on the potential limits of using nucleic acid-based drugs to control the propagation of HCV. Finally, the above conclusions received support from experiments performed using a collection of SOFA-HDV ribozymes directed against HCV (−) strands.

Economic Evaluations of Adult Male Circumcision for Prevention of Heterosexual Acquisition of HIV in Men in Sub-Saharan Africa: A Systematic Review

Nicolas Guex et al. — Wed, 10 Mar 2010 08:00:00 +0000

Background

There is conclusive evidence from observational data and three randomized controlled trials that circumcised men have a significantly lower risk of becoming infected with the human immunodeficiency virus (HIV). The aim of this study was to systematically review economic evaluations on adult male circumcision (AMC) for prevention of heterosexual acquisition of HIV in men.

Methods and Findings

Studies were identified from the following bibliographic databases: MEDLINE (Ovid), EMBASE (Ovid), Cochrane Library (Wiley’s internet version), NHS EED and DARE Office of Health Economics HEED. The searches were conducted in November 2009. The Drummond 10-point checklist was used for methodological critique of the economic evaluations. Cost data were inflated and converted to 2008 US dollars (US$). Of 264 identified papers, only five met the inclusion criteria and were included in the review. The studies were published between 2006 and 2009. Most of the studies were carried out from the perspective of government healthcare payer. The time horizon ranged from 10 to 20 years. All studies reported that AMC is cost-effective. The reported cost per HIV infection averted ranged from US$174 to US$2808. The key driver of the cost-effectiveness models was circumcision efficacy.

Conclusions

All published economic evaluations offered the same conclusion that AMC is cost-effective and potentially cost-saving for prevention of heterosexual acquisition of HIV in men. On these grounds, AMC may be seen as a promising new form of strategy for prevention of HIV and should be implemented in conjunction with other evidence-based prevention methods.

Human Leukocyte Antigens and HIV Type 1 Viral Load in Early and Chronic Infection: Predominance of Evolving Relationships

Nicolas Guex et al. — Wed, 10 Mar 2010 08:00:00 +0000

Background

During untreated, chronic HIV-1 infection, plasma viral load (VL) is a relatively stable quantitative trait that has clinical and epidemiological implications. Immunogenetic research has established various human genetic factors, especially human leukocyte antigen (HLA) variants, as independent determinants of VL set-point.

Methodology/Principal Findings

To identify and clarify HLA alleles that are associated with either transient or durable immune control of HIV-1 infection, we evaluated the relationships of HLA class I and class II alleles with VL among 563 seroprevalent Zambians (SPs) who were seropositive at enrollment and 221 seroconverters (SCs) who became seropositive during quarterly follow-up visits. After statistical adjustments for non-genetic factors (sex and age), two unfavorable alleles (A*3601 and DRB1*0102) were independently associated with high VL in SPs (p<0.01) but not in SCs. In contrast, favorable HLA variants, mainly A*74, B*13, B*57 (or Cw*18), and one HLA-A and HLA-C combination (A*30+Cw*03), dominated in SCs; their independent associations with low VL were reflected in regression beta estimates that ranged from −0.47±0.23 to −0.92±0.32 log₁₀ in SCs (p<0.05). Except for Cw*18, all favorable variants had diminishing or vanishing association with VL in SPs (p≤0.86).

Conclusions/Significance

Overall, each of the three HLA class I genes had at least one allele that might contribute to effective immune control, especially during the early course of HIV-1 infection. These observations can provide a useful framework for ongoing analyses of viral mutations induced by protective immune responses.

A Search for Trypanosoma brucei rhodesiense Diagnostic Antigens by Proteomic Screening and Targeted Cloning

Nicolas Guex et al. — Wed, 10 Mar 2010 08:00:00 +0000

Background

The only available diagnostic method for East African trypanosomiasis is light microscopy of blood samples. A simple immunodiagnostic would greatly aid trypanosomiasis control.

Methodology and Principal Findings

To find trypanosome proteins that are specifically recognised by sera from human sleeping sickness patients, we have screened the Trypanosoma brucei brucei proteome by Western blotting. Using cytosolic, cytoskeletal and glycosomal fractions, we found that the vast majority of abundant trypanosome proteins is not specifically recognised by patient sera. We identified phosphoglycerate kinase (PGKC), heat shock protein (HSP70), and histones H2B and H3 as possible candidate diagnostic antigens. These proteins, plus paraflagellar rod protein 1, rhodesain (a cysteine protease), and an extracellular fragment of the Trypanosoma brucei nucleoside transporter TbNT10, were expressed in E. coli and tested for reactivity with patient and control sera. Only TbHSP70 was preferentially recognized by patient sera, but the sensitivity and specificity were insufficient for use of TbHSP70 alone as a diagnostic. Immunoprecipitation using a native protein extract revealed no specifically reacting proteins.

Conclusions

No abundant T. brucei soluble, glycosomal or cytoskeletal protein is likely to be useful in diagnosis. To find useful diagnostic antigens it will therefore be necessary to use more sophisticated proteomic methods, or to test a very large panel of candidate proteins.

Localization of Canine Brachycephaly Using an Across Breed Mapping Approach

Nicolas Guex et al. — Wed, 10 Mar 2010 08:00:00 +0000

The domestic dog, Canis familiaris, exhibits profound phenotypic diversity and is an ideal model organism for the genetic dissection of simple and complex traits. However, some of the most interesting phenotypes are fixed in particular breeds and are therefore less tractable to genetic analysis using classical segregation-based mapping approaches. We implemented an across breed mapping approach using a moderately dense SNP array, a low number of animals and breeds carefully selected for the phenotypes of interest to identify genetic variants responsible for breed-defining characteristics. Using a modest number of affected (10–30) and control (20–60) samples from multiple breeds, the correct chromosomal assignment was identified in a proof of concept experiment using three previously defined loci; hyperuricosuria, white spotting and chondrodysplasia. Genome-wide association was performed in a similar manner for one of the most striking morphological traits in dogs: brachycephalic head type. Although candidate gene approaches based on comparable phenotypes in mice and humans have been utilized for this trait, the causative gene has remained elusive using this method. Samples from nine affected breeds and thirteen control breeds identified strong genome-wide associations for brachycephalic head type on Cfa 1. Two independent datasets identified the same genomic region. Levels of relative heterozygosity in the associated region indicate that it has been subjected to a selective sweep, consistent with it being a breed defining morphological characteristic. Genotyping additional dogs in the region confirmed the association. To date, the genetic structure of dog breeds has primarily been exploited for genome wide association for segregating traits. These results demonstrate that non-segregating traits under strong selection are equally tractable to genetic analysis using small sample numbers.

The RGD Domain of Human Osteopontin Promotes Tumor Growth and Metastasis through Activation of Survival Pathways

Nicolas Guex et al. — Wed, 10 Mar 2010 08:00:00 +0000

Background

Human osteopontin (OPN), a known tumor associated protein, exists in different isoforms, whose function is unclear. It also possesses a RGD domain, which has been implicated in diverse function. Here, we use genetic approaches to systematically investigate the function of the RGD domain in different OPN isoforms on tumor progression and metastasis for 2 different solid tumor models.

Methodology/Principal Findings

Using isoform-specific qRT-PCR, we found that OPN-A and B were the main isoforms overexpressed in evaluated human tumors, which included 4 soft tissue sarcomas, 24 lung and 30 head and neck carcinomas. Overexpression of either OPN-A or B in two different cell types promoted local tumor growth and lung metastasis in SCID mouse xenografts. However, expression of either isoform with the RGD domain either mutated or deleted decreased tumor growth and metastasis, and resulted in increased apoptosis by TUNEL staining. In vitro, whereas mutation of the RGD domain did not affect cell-cell adhesion, soft agar growth or cell migration, it increased apoptosis under hypoxia and serum starvation. This effect could be mitigated when the RGD mutant cells were treated with condition media containing WT OPN. Mechanistically, the RGD region of OPN inhibited apoptosis by inducing NF-κB activation and FAK phosphorylation. Inhibition of NF-κB (by siRNA to the p65 subunit) or FAK activation (by a inhibitor) significantly increased apoptosis under hypoxia in WT OPN cells, but not in RGD mutant cells.

Conclusion/Significance

Unlike prior reports, our data suggest that the RGD domain of both OPN-A and B promote tumor growth and metastasis mainly by protecting cells against apoptosis under stressed conditions and not via migration or invasion. Future inhibitors directed against OPN should target multiple isoforms and should inhibit cell survival mechanisms that involve the RGD domain, FAK phosphorylation and NF-κB activation.

Intracellular Calcium Spikes in Rat Suprachiasmatic Nucleus Neurons Induced by BAPTA-Based Calcium Dyes

Nicolas Guex et al. — Wed, 10 Mar 2010 08:00:00 +0000

Background

Circadian rhythms in spontaneous action potential (AP) firing frequencies and in cytosolic free calcium concentrations have been reported for mammalian circadian pacemaker neurons located within the hypothalamic suprachiasmatic nucleus (SCN). Also reported is the existence of “Ca²⁺ spikes” (i.e., [Ca²⁺]_c transients having a bandwidth of 10~100 seconds) in SCN neurons, but it is unclear if these SCN Ca²⁺ spikes are related to the slow circadian rhythms.

Methodology/Principal Findings

We addressed this issue based on a Ca²⁺ indicator dye (fluo-4) and a protein Ca²⁺ sensor (yellow cameleon). Using fluo-4 AM dye, we found spontaneous Ca²⁺ spikes in 18% of rat SCN cells in acute brain slices, but the Ca²⁺ spiking frequencies showed no day/night variation. We repeated the same experiments with rat (and mouse) SCN slice cultures that expressed yellow cameleon genes for a number of different circadian phases and, surprisingly, spontaneous Ca²⁺ spike was barely observed (<3%). When fluo-4 AM or BAPTA-AM was loaded in addition to the cameleon-expressing SCN cultures, however, the number of cells exhibiting Ca²⁺ spikes was increased to 13~14%.

Conclusions/Significance

Despite our extensive set of experiments, no evidence of a circadian rhythm was found in the spontaneous Ca²⁺ spiking activity of SCN. Furthermore, our study strongly suggests that the spontaneous Ca²⁺ spiking activity is caused by the Ca²⁺ chelating effect of the BAPTA-based fluo-4 dye. Therefore, this induced activity seems irrelevant to the intrinsic circadian rhythm of [Ca²⁺]_c in SCN neurons. The problems with BAPTA based dyes are widely known and our study provides a clear case for concern, in particular, for SCN Ca²⁺ spikes. On the other hand, our study neither invalidates the use of these dyes as a whole, nor undermines the potential role of SCN Ca²⁺ spikes in the function of SCN.

Category and Perceptual Learning in Subjects with Treated Wilson’s Disease

Nicolas Guex et al. — Wed, 10 Mar 2010 08:00:00 +0000

To explore the relationship between category and perceptual learning, we examined both category and perceptual learning in patients with treated Wilson’s disease (WD), whose basal ganglia, known to be important in category learning, were damaged by the disease. We measured their learning rate and accuracy in rule-based and information-integration category learning, and magnitudes of perceptual learning in a wide range of external noise conditions, and compared the results with those of normal controls. The WD subjects exhibited deficits in both forms of category learning and in perceptual learning in high external noise. However, their perceptual learning in low external noise was relatively spared. There was no significant correlation between the two forms of category learning, nor between perceptual learning in low external noise and either form of category learning. Perceptual learning in high external noise was, however, significantly correlated with information-integration but not with rule-based category learning. The results suggest that there may be a strong link between information-integration category learning and perceptual learning in high external noise. Damage to brain structures that are important for information-integration category learning may lead to poor perceptual learning in high external noise, yet spare perceptual learning in low external noise. Perceptual learning in high and low external noise conditions may involve separate neural substrates.

Human Mammary Epithelial Cells Exhibit a Bimodal Correlated Random Walk Pattern

Nicolas Guex et al. — Wed, 10 Mar 2010 08:00:00 +0000

Background

Organisms, at scales ranging from unicellular to mammals, have been known to exhibit foraging behavior described by random walks whose segments confirm to Lévy or exponential distributions. For the first time, we present evidence that single cells (mammary epithelial cells) that exist in multi-cellular organisms (humans) follow a bimodal correlated random walk (BCRW).

Methodology/Principal Findings

Cellular tracks of MCF-10A pBabe, neuN and neuT random migration on 2-D plastic substrates, analyzed using bimodal analysis, were found to reveal the BCRW pattern. We find two types of exponentially distributed correlated flights (corresponding to what we refer to as the directional and re-orientation phases) each having its own correlation between move step-lengths within flights. The exponential distribution of flight lengths was confirmed using different analysis methods (logarithmic binning with normalization, survival frequency plots and maximum likelihood estimation).

Conclusions/Significance

Because of the presence of non-uniform turn angle distribution of move step-lengths within a flight and two different types of flights, we propose that the epithelial random walk is a BCRW comprising of two alternating modes with varying degree of correlations, rather than a simple persistent random walk. A BCRW model rather than a simple persistent random walk correctly matches the super-diffusivity in the cell migration paths as indicated by simulations based on the BCRW model.

Discovery of Novel MicroRNAs in Female Reproductive Tract Using Next Generation Sequencing

Nicolas Guex et al. — Wed, 10 Mar 2010 08:00:00 +0000

MicroRNAs (miRNAs) are small non-coding RNAs that mediate post-transcriptional gene silencing. Over 700 human miRNAs have currently been identified, many of which are mutated or de-regulated in diseases. Here we report the identification of novel miRNAs through deep sequencing the small RNAome (<30 nt) of over 100 tissues or cell lines derived from human female reproductive organs in both normal and disease states. These specimens include ovarian epithelium and ovarian cancer, endometrium and endometriomas, and uterine myometrium and uterine smooth muscle tumors. Sequence reads not aligning with known miRNAs were each mapped to the genome to extract flanking sequences. These extended sequence regions were folded in silico to identify RNA hairpins. Sequences demonstrating the ability to form a stem loop structure with low minimum free energy (<−25 kcal) and predicted Drosha and Dicer cut sites yielding a mature miRNA sequence matching the actual sequence were considered putative novel miRNAs. Additional confidence was achieved when putative novel hairpins assembled a collection of sequences highly similar to the putative mature miRNA but with heterogeneous 3′-ends. A confirmed novel miRNA fulfilled these criteria and had its “star” sequence in our collection. We found 7 distinct confirmed novel miRNAs, and 51 additional novel miRNAs that represented highly confident predictions but without detectable star sequences. Our novel miRNAs were detectable in multiple samples, but expressed at low levels and not specific to any one tissue or cell type. To date, this study represents the largest set of samples analyzed together to identify novel miRNAs.

Characterization of the Activation of Protein Tyrosine Phosphatase, Receptor-Type, Z Polypeptide 1 (PTPRZ1) by Hypoxia Inducible Factor-2 Alpha

Nicolas Guex et al. — Wed, 10 Mar 2010 08:00:00 +0000

Background

Hypoxia inducible factors (HIFs) are the principal means by which cells upregulate genes in response to hypoxia and certain other stresses. There are two major HIFs, HIF-1 and HIF-2. We previously found that certain genes are preferentially activated by HIF-2. One was protein tyrosine phosphatase, receptor-type, Z polypeptide 1 (PTPRZ1). PTPRZ1 is overexpressed in a number of tumors and has been implicated in glioblastoma pathogenesis.

Methodology/Principal Findings

To understand the preferential activation of PTPRZ1 by HIF-2, we studied the PTPRZ1 promoter in HEK293T cells and Hep3B cells. Through deletion and mutational analysis, we identified the principal hypoxia response element. This element bound to both HIF-1 and HIF-2. We further identified a role for ELK1, an E26 transformation-specific (Ets) factor that can bind to HIF-2α but not HIF-1α, in the HIF-2 responsiveness. Knock-down experiments using siRNA to ELK1 decreased HIF-2 activation by over 50%. Also, a deletion mutation of one of the two Ets binding motifs located near the principal hypoxia response element similarly decreased activation of the PTPRZ1 promoter by HIF-2. Finally, chromatin immunoprecipitation assays showed binding of HIF and ELK1 to the PTPRZ1 promoter region.

Conclusions/Significance

These results identify HIF-binding and Ets-binding motifs on the PTPRZ1 promoter and provide evidence that preferential activation of PTPRZ1 by HIF-2 results at least in part from cooperative binding of HIF-2 and ELK1 to nearby sites on the PTPRZ1 promoter region. These results may have implications in tumor pathogenesis and in understanding neurobiology, and may help inform the development of novel tumor therapy.

Pathogenic Polyglutamine Tracts Are Potent Inducers of Spontaneous Sup35 and Rnq1 Amyloidogenesis

Nicolas Guex et al. — Wed, 10 Mar 2010 08:00:00 +0000

The glutamine/asparagine (Q/N)-rich yeast prion protein Sup35 has a low intrinsic propensity to spontaneously self-assemble into ordered, β-sheet-rich amyloid fibrils. In yeast cells, de novo formation of Sup35 aggregates is greatly facilitated by high protein concentrations and the presence of preformed Q/N-rich protein aggregates that template Sup35 polymerization. Here, we have investigated whether aggregation-promoting polyglutamine (polyQ) tracts can stimulate the de novo formation of ordered Sup35 protein aggregates in the absence of Q/N-rich yeast prions. Fusion proteins with polyQ tracts of different lengths were produced and their ability to spontaneously self-assemble into amlyloid structures was analyzed using in vitro and in vivo model systems. We found that Sup35 fusions with pathogenic (≥54 glutamines), as opposed to non-pathogenic (19 glutamines) polyQ tracts efficiently form seeding-competent protein aggregates. Strikingly, polyQ-mediated de novo assembly of Sup35 protein aggregates in yeast cells was independent of pre-existing Q/N-rich protein aggregates. This indicates that increasing the content of aggregation-promoting sequences enhances the tendency of Sup35 to spontaneously self-assemble into insoluble protein aggregates. A similar result was obtained when pathogenic polyQ tracts were linked to the yeast prion protein Rnq1, demonstrating that polyQ sequences are generic inducers of amyloidogenesis. In conclusion, long polyQ sequences are powerful molecular tools that allow the efficient production of seeding-competent amyloid structures.

Decreased NK Cell FcRγ in HIV-1 Infected Individuals Receiving Combination Antiretroviral Therapy: a Cross Sectional Study

Nicolas Guex et al. — Wed, 10 Mar 2010 08:00:00 +0000

Background

FcRγ is an immunoreceptor tyrosine-based activation motif (ITAM)-signalling protein essential for immunoreceptor signaling and monocyte, macrophage and NK cell function. Previous study from our laboratory showed that FcRγ is down-regulated in HIV-infected macrophages in vitro. FcRγ expression in immune cells present in HIV-infected individuals is unknown.

Methodology/Principal Findings

We compared FcRγ expression in peripheral blood mononuclear cells isolated from HIV-1-infected individuals receiving combination antiretroviral therapy and healthy, HIV-1-uninfected individuals. FcRγ mRNA and protein levels were measured using quantitative real-time PCR and immunoblotting, respectively. CD56⁺ CD94⁺ lymphocytes isolated from blood of HIV-1 infected individuals had reduced FcRγ protein expression compared to HIV-uninfected individuals (decrease = 76.8%, n = 18 and n = 12 respectively, p = 0.0036). In a second group of patients, highly purified NK cells had reduced FcRγ protein expression compared to uninfected controls (decrease = 50.2%, n = 9 and n = 8 respectively, p = 0.021). Decreased FcRγ expression in CD56+CD94+ lymphocytes was associated with reduced mRNA (51.7%, p = 0.021) but this was not observed for the smaller group of patients analysed for NK cell expression (p = 0.36).

Conclusion/Significance

These data suggest biochemical defects in ITAM-dependent signalling within NK cells in HIV-infected individuals which is present in the context of treatment with combination antiretroviral therapy.

Large–Scale Movement and Reef Fidelity of Grey Reef Sharks

Nicolas Guex et al. — Wed, 10 Mar 2010 08:00:00 +0000

Despite an Indo-Pacific wide distribution, the movement patterns of grey reef sharks (Carcharhinus amblyrhynchos) and fidelity to individual reef platforms has gone largely unstudied. Their wide distribution implies that some individuals have dispersed throughout tropical waters of the Indo-Pacific, but data on large-scale movements do not exist. We present data from nine C. amblyrhynchos monitored within the Great Barrier Reef and Coral Sea off the coast of Australia. Shark presence and movements were monitored via an array of acoustic receivers for a period of six months in 2008. During the course of this monitoring few individuals showed fidelity to an individual reef suggesting that current protective areas have limited utility for this species. One individual undertook a large-scale movement (134 km) between the Coral Sea and Great Barrier Reef, providing the first evidence of direct linkage of C. amblyrhynchos populations between these two regions. Results indicate limited reef fidelity and evidence of large-scale movements within northern Australian waters.

Spatial control of EGF receptor activation by reversible dimerization on living cells.

Chung I, Akita R, Vandlen R, Toomre D, Schlessinger J, Mellman I — Wed, 10 Mar 2010 00:25:06 +0000

Spatial control of EGF receptor activation by reversible dimerization on living cells.

Nature. 2010 Mar 7;

Authors: Chung I, Akita R, Vandlen R, Toomre D, Schlessinger J, Mellman I

Epidermal growth factor receptor (EGFR) is a type I receptor tyrosine kinase, the deregulation of which has been implicated in a variety of human carcinomas. EGFR signalling is preceded by receptor dimerization, typically thought to result from a ligand-induced conformational change in the ectodomain that exposes a loop (dimerization arm) required for receptor association. Ligand binding may also trigger allosteric changes in the cytoplasmic domain of the receptor that is crucial for signalling. Despite these insights, ensemble-averaging approaches have not determined the precise mechanism of receptor activation in situ. Using quantum-dot-based optical tracking of single molecules combined with a novel time-dependent diffusivity analysis, here we present the dimerization dynamics of individual EGFRs on living cells. Before ligand addition, EGFRs spontaneously formed finite-lifetime dimers kinetically stabilized by their dimerization arms. The dimers were primed both for ligand binding and for signalling, such that after EGF addition they rapidly showed a very slow diffusivity state that correlated with activation. Although the kinetic stability of unliganded dimers was in principle sufficient for EGF-independent activation, ligand binding was still required for signalling. Interestingly, dimers were enriched in the cell periphery in an actin- and receptor-expression-dependent fashion, resulting in a peripheral enhancement of EGF-induced signalling that may enable polarized responses to growth factors.

PMID: 20208517 [PubMed - as supplied by publisher]

[...]

Control of female gamete formation by a small RNA pathway in Arabidopsis.

Wed, 10 Mar 2010 00:25:05 +0000

Control of female gamete formation by a small RNA pathway in Arabidopsis.

Nature. 2010 Mar 7;

Authors: Olmedo-Monfil V, Durán-Figueroa N, Arteaga-Vázquez M, Demesa-Arévalo E, Autran D, Grimanelli D, Slotkin RK, Martienssen RA, Vielle-Calzada JP

In the ovules of most sexual flowering plants female gametogenesis is initiated from a single surviving gametic cell, the functional megaspore, formed after meiosis of the somatically derived megaspore mother cell (MMC). Because some mutants and certain sexual species exhibit more than one MMC, and many others are able to form gametes without meiosis (by apomixis), it has been suggested that somatic cells in the ovule are competent to respond to a local signal likely to have an important function in determination. Here we show that the Arabidopsis protein ARGONAUTE 9 (AGO9) controls female gamete formation by restricting the specification of gametophyte precursors in a dosage-dependent, non-cell-autonomous manner. Mutations in AGO9 lead to the differentiation of multiple gametic cells that are able to initiate gametogenesis. The AGO9 protein is not expressed in the gamete lineage; instead, it is expressed in cytoplasmic foci of somatic companion cells. Mutations in SUPPRESSOR OF GENE SILENCING 3 and RNA-DEPENDENT RNA POLYMERASE 6 exhibit an identical defect to ago9 mutants, indicating that the movement of small RNA (sRNAs) silencing out of somatic companion cells is necessary for controlling the specification of gametic cells. AGO9 preferentially interacts with 24-nucleotide sRNAs derived from transposable elements (TEs), and its activity is necessary to silence TEs in female gametes and their accessory cells. Our results show that AGO9-dependent sRNA silencing is crucial to specify cell fate in the Arabidopsis ovule, and that epigenetic reprogramming in companion cells is necessary for sRNA-dependent silencing in plant gametes.

PMID: 20208518 [PubMed - as supplied by publisher]

[...]

Transcription-independent ARF regulation in oncogenic stress-mediated p53 responses.

Chen D, Shan J, Zhu WG, Qin J, Gu W — Wed, 10 Mar 2010 00:25:04 +0000

Transcription-independent ARF regulation in oncogenic stress-mediated p53 responses.

Nature. 2010 Mar 7;

Authors: Chen D, Shan J, Zhu WG, Qin J, Gu W

The tumour suppressor ARF is specifically required for p53 activation under oncogenic stress. Recent studies showed that p53 activation mediated by ARF, but not that induced by DNA damage, acts as a major protection against tumorigenesis in vivo under certain biological settings, suggesting that the ARF-p53 axis has more fundamental functions in tumour suppression than originally thought. Because ARF is a very stable protein in most human cell lines, it has been widely assumed that ARF induction is mediated mainly at the transcriptional level and that activation of the ARF-p53 pathway by oncogenes is a much slower and largely irreversible process by comparison with p53 activation after DNA damage. Here we report that ARF is very unstable in normal human cells but that its degradation is inhibited in cancerous cells. Through biochemical purification, we identified a specific ubiquitin ligase for ARF and named it ULF. ULF interacts with ARF both in vitro and in vivo and promotes the lysine-independent ubiquitylation and degradation of ARF. ULF knockdown stabilizes ARF in normal human cells, triggering ARF-dependent, p53-mediated growth arrest. Moreover, nucleophosmin (NPM) and c-Myc, both of which are commonly overexpressed in cancer cells, are capable of abrogating ULF-mediated ARF ubiquitylation through distinct mechanisms, and thereby promote ARF stabilization in cancer cells. These findings reveal the dynamic feature of the ARF-p53 pathway and suggest that transcription-independent mechanisms are critically involved in ARF regulation during responses to oncogenic stress.

PMID: 20208519 [PubMed - as supplied by publisher]

[...]

Release of the mitochondrial endosymbiont helps explain sterile inflammation [Letter]

Masters, S. L., Walsh, P. T. — Tue, 09 Mar 2010 17:27:20 +0000

Kupfer-type immunological synapse characteristics do not predict anti-brain tumor cytolytic T-cell function in vivo [Neuroscience]

Tue, 09 Mar 2010 17:27:19 +0000

To analyze the in vivo structure of antigen-specific immunological synapses during an effective immune response, we established brain tumors expressing…

Peptide vaccines prevent tumor growth by activating T cells that respond to native tumor antigens [Immunology]

Jordan, K. R., McMahan, R. H., Kemmler, C. B., Kappler, J. W., Slansky, J. E. — Tue, 09 Mar 2010 17:27:19 +0000

Peptide vaccines enhance the response of T cells toward tumor antigens and represent a strategy to augment antigen-independent immunotherapies of…

Perceptron learning rule derived from spike-frequency adaptation and spike-time-dependent plasticity [Neuroscience]

D'Souza, P., Liu, S.-C., Hahnloser, R. H. R. — Tue, 09 Mar 2010 17:27:19 +0000

It is widely believed that sensory and motor processing in the brain is based on simple computational primitives rooted in…

Positive allosteric modulators of the human sweet taste receptor enhance sweet taste [Neuroscience]

Tue, 09 Mar 2010 17:27:19 +0000

To identify molecules that could enhance sweetness perception, we undertook the screening of a compound library using a cell-based assay…

Molecular mechanism of the sweet taste enhancers [Neuroscience]

Tue, 09 Mar 2010 17:27:19 +0000

Positive allosteric modulators of the human sweet taste receptor have been developed as a new way of reducing dietary sugar…

Topographic organization of macaque area LIP [Neuroscience]

Tue, 09 Mar 2010 17:27:19 +0000

Despite several attempts to define retinotopic maps in the macaque lateral intraparietal area (LIP) using histological, electrophysiological, and neuroimaging methods,…

Input-specific synaptic plasticity in the amygdala is regulated by neuroligin-1 via postsynaptic NMDA receptors [Neuroscience]

Tue, 09 Mar 2010 17:27:19 +0000

Despite considerable evidence for a critical role of neuroligin-1 in the specification of excitatory synapses, the cellular mechanisms and physiological…

Rituximab specifically depletes short-lived autoreactive plasma cells in a mouse model of inflammatory arthritis [Immunology]

Huang, H., Benoist, C., Mathis, D. — Tue, 09 Mar 2010 17:27:19 +0000

There is increasing appreciation of the important role of B cells in many autoimmune diseases and consequently, increasing interest in…

High-throughput identification of protein localization dependency networks [Microbiology]

Tue, 09 Mar 2010 17:27:19 +0000

Bacterial cells are highly organized with many protein complexes and DNA loci dynamically positioned to distinct subcellular sites over the…

Insulin enhances glucose-stimulated insulin secretion in healthy humans [Physiology]

Tue, 09 Mar 2010 17:27:19 +0000

Islet β-cells express both insulin receptors and insulin-signaling proteins. Recent evidence from rodents in vivo and from islets isolated from…

Structural mechanism of host Rab1 activation by the bifunctional Legionella type IV effector SidM/DrrA [Microbiology]

Zhu, Y., Hu, L., Zhou, Y., Yao, Q., Liu, L., Shao, F. — Tue, 09 Mar 2010 17:27:19 +0000

Bacterial pathogens deliver effector proteins with diverse biochemical activities into host cells, thereby modulating various host functions. Legionella pneumophila hijacks…

The PPH1 phosphatase is specifically involved in LHCII dephosphorylation and state transitions in Arabidopsis [Plant_Biology]

Tue, 09 Mar 2010 17:27:19 +0000

The ability of plants to adapt to changing light conditions depends on a protein kinase network in the chloroplast that…

Reassortment between avian H5N1 and human H3N2 influenza viruses creates hybrid viruses with substantial virulence [Microbiology]

Tue, 09 Mar 2010 17:27:19 +0000

The spread of avian H5N1 influenza viruses around the globe has become a worldwide public health concern. To evaluate the…

Toward discovery science of human brain function [Neuroscience]

Tue, 09 Mar 2010 17:27:19 +0000

Although it is being successfully implemented for exploration of the genome, discovery science has eluded the functional neuroimaging community. The…

Critical role of IRF-5 in regulation of B-cell differentiation [Immunology]

Lien, C., Fang, C.-M., Huso, D., Livak, F., Lu, R., Pitha, P. M. — Tue, 09 Mar 2010 17:27:19 +0000

IFN-regulatory factor 5 (IRF-5), a member of the IRF family, is a transcription factor that has a key role in…

Cholesterol trafficking is required for mTOR activation in endothelial cells [Pharmacology]

Xu, J., Dang, Y., Ren, Y. R., Liu, J. O. — Tue, 09 Mar 2010 17:27:19 +0000

Mammalian target of rapamycin (mTOR) constitutes a nodal point of a signaling network that regulates cell growth and proliferation in…

Arabidopsis thaliana life without phytochromes [Plant_Biology]

Strasser, B., Sanchez-Lamas, M., Yanovsky, M. J., Casal, J. J., Cerdan, P. D. — Tue, 09 Mar 2010 17:27:19 +0000

Plants use light as a source of energy for photosynthesis and as a source of environmental information perceived by photoreceptors….

Distributed neural system for general intelligence revealed by lesion mapping [Psychological_And_Cognitive_Sciences-SS]

Tue, 09 Mar 2010 17:27:19 +0000

General intelligence (g) captures the performance variance shared across cognitive tasks and correlates with real-world success. Yet it remains debated…

Functional specializations for music processing in the human newborn brain [Neuroscience]

Tue, 09 Mar 2010 17:27:19 +0000

In adults, specific neural systems with right-hemispheric weighting are necessary to process pitch, melody, and harmony as well as structure…

Negative feedback control of HIF-1 through REDD1-regulated ROS suppresses tumorigenesis [Medical_Sciences]

Tue, 09 Mar 2010 17:27:19 +0000

The HIF family of hypoxia-inducible transcription factors are key mediators of the physiologic response to hypoxia, whose dysregulation promotes tumorigenesis….

Light-induced translocation of Drosophila visual Arrestin2 depends on Rac2 [Neuroscience]

Elsaesser, R., Kalra, D., Li, R., Montell, C. — Tue, 09 Mar 2010 17:27:19 +0000

Photoreceptor cells are remarkable in their ability to adjust their sensitivity to light over a wide range of intensities. Rapid…

Interactome analysis of longitudinal pharyngeal infection of cynomolgus macaques by group A Streptococcus [Microbiology]

Tue, 09 Mar 2010 17:27:19 +0000

Relatively little is understood about the dynamics of global host–pathogen transcriptome changes that occur during bacterial infection of mucosal surfaces….

Tissue-engineered vascular grafts transform into mature blood vessels via an inflammation-mediated process of vascular remodeling [Medical_Sciences]

Tue, 09 Mar 2010 17:27:19 +0000

Biodegradable scaffolds seeded with bone marrow mononuclear cells (BMCs) are the earliest tissue-engineered vascular grafts (TEVGs) to be used clinically….

Correction for Jordan et al., Peptide vaccines prevent tumor growth by activating T cells that respond to native tumor antigens [errata]

Tue, 09 Mar 2010 17:27:19 +0000

Amoeboid organism solves complex nutritional challenges [Ecology]

Dussutour, A., Latty, T., Beekman, M., Simpson, S. J. — Tue, 09 Mar 2010 17:27:18 +0000

A fundamental question in nutritional biology is how distributed systems maintain an optimal supply of multiple nutrients essential for life…

Climatic context and ecological implications of summer fog decline in the coast redwood region [Ecology]

Johnstone, J. A., Dawson, T. E. — Tue, 09 Mar 2010 17:27:18 +0000

Biogeographical, physiological, and paleoecological evidence suggests that the coast redwood [Sequoia sempervirens (D. Don) Endl.] is closely associated with the…

Atomic details of near-transition state conformers for enzyme phosphoryl transfer revealed by Formula rather than by phosphoranes [Biochemistry]

Tue, 09 Mar 2010 17:27:18 +0000

Prior evidence supporting the direct observation of phosphorane intermediates in enzymatic phosphoryl transfer reactions was based on the interpretation of…

Targeting mycobacterium protein tyrosine phosphatase B for antituberculosis agents [Biochemistry]

Tue, 09 Mar 2010 17:27:18 +0000

Protein tyrosine phosphatases are often exploited and subverted by pathogenic bacteria to cause human diseases. The tyrosine phosphatase mPTPB from…

Sampling the N-terminal proteome of human blood [Biochemistry]

Wildes, D., Wells, J. A. — Tue, 09 Mar 2010 17:27:18 +0000

The proteomes of blood plasma and serum represent a potential gold mine of biological and diagnostic information, but challenges such…

RFWD3-Mdm2 ubiquitin ligase complex positively regulates p53 stability in response to DNA damage [Biochemistry]

Tue, 09 Mar 2010 17:27:18 +0000

In unstressed cells, the tumor suppressor p53 is maintained at low levels by ubiquitin-mediated proteolysis mainly through Mdm2. In response…

Female-specific flightless phenotype for mosquito control [Applied_Biological_Sciences]

Tue, 09 Mar 2010 17:27:18 +0000

Dengue and dengue hemorrhagic fever are increasing public health problems with an estimated 50–100 million new infections each year. Aedes aegypti…

Rotational dynamics of DNA on the nucleosome surface markedly impact accessibility to a DNA repair enzyme [Genetics]

Hinz, J. M., Rodriguez, Y., Smerdon, M. J. — Tue, 09 Mar 2010 17:27:18 +0000

Histones play a crucial role in the organization of DNA in the nucleus, but their presence can prevent interactions with…

Vitrification and levitation of a liquid droplet on liquid nitrogen [Physics]

Tue, 09 Mar 2010 17:27:18 +0000

The vitrification of a liquid occurs when ice crystal formation is prevented in the cryogenic environment through ultrarapid cooling. In…

Specificity landscapes of DNA binding molecules elucidate biological function [Applied_Biological_Sciences]

Tue, 09 Mar 2010 17:27:18 +0000

Evaluating the specificity spectra of DNA binding molecules is a nontrivial challenge that hinders the ability to decipher gene regulatory…

ParA2, a Vibrio cholerae chromosome partitioning protein, forms left-handed helical filaments on DNA [Biochemistry]

Tue, 09 Mar 2010 17:27:18 +0000

Most bacterial chromosomes contain homologs of plasmid partitioning (par) loci. These loci encode ATPases called ParA that are thought to…

Multiple translational products from a five-nucleotide ribozyme [Biochemistry]

Turk, R. M., Chumachenko, N. V., Yarus, M. — Tue, 09 Mar 2010 17:27:18 +0000

An indispensable step in protein biosynthesis is the 2^‘(3^‘) aminoacylation of tRNA by aminoacyl-tRNA synthetases. Here we show that a…

Genetic and dietary regulation of lipid droplet expansion in Caenorhabditis elegans [Genetics]

Tue, 09 Mar 2010 17:27:18 +0000

Dietary fat accumulates in lipid droplets or endolysosomal compartments that undergo selective expansion under normal or pathophysiological conditions. We find…

Appropriate maturation and folding of 16S rRNA during 30S subunit biogenesis are critical for translational fidelity [Biochemistry]

Roy-Chaudhuri, B., Kirthi, N., Culver, G. M. — Tue, 09 Mar 2010 17:27:18 +0000

Ribosomal protein S5 is critical for small ribosomal subunit (SSU) assembly and is indispensable for SSU function. Previously, we identified…

Arboreality has allowed for the evolution of increased longevity in mammals [Evolution]

Shattuck, M. R., Williams, S. A. — Tue, 09 Mar 2010 17:27:18 +0000

The evolutionary theory of aging predicts that species will experience delayed senescence and increased longevity when rates of extrinsic mortality…

Mutation-selection models of coding sequence evolution with site-heterogeneous amino acid fitness profiles [Evolution]

Rodrigue, N., Philippe, H., Lartillot, N. — Tue, 09 Mar 2010 17:27:18 +0000

Modeling the interplay between mutation and selection at the molecular level is key to evolutionary studies. To this end, codon-based…

Follistatin-like-1, a diffusible mesenchymal factor determines the fate of epithelium [Developmental_Biology]

Umezu, T., Yamanouchi, H., Iida, Y., Miura, M., Tomooka, Y. — Tue, 09 Mar 2010 17:27:18 +0000

Mesenchyme is generally believed to play critical roles in “secondary induction” during organogenesis. Because of the complexity of tissue interactions…

Dual role of lignin in plant litter decomposition in terrestrial ecosystems [Environmental_Sciences-PS]

Austin, A. T., Ballare, C. L. — Tue, 09 Mar 2010 17:27:18 +0000

Plant litter decomposition is a critical step in the formation of soil organic matter, the mineralization of organic nutrients, and…

Fluctuation enhanced electrochemical reaction rates at the nanoscale [Chemistry]

Garcia-Morales, V., Krischer, K. — Tue, 09 Mar 2010 17:27:18 +0000

The electrode potential constitutes a dynamical variable whenever an electrode is resistively coupled to the electric circuit. We show that…

Phylogenetic analysis of 83 plastid genes further resolves the early diversification of eudicots [Evolution]

Moore, M. J., Soltis, P. S., Bell, C. D., Burleigh, J. G., Soltis, D. E. — Tue, 09 Mar 2010 17:27:18 +0000

Although Pentapetalae (comprising all core eudicots except Gunnerales) include 70% of all angiosperms, the origin of and relationships among the…

Lattice thermal conductivity of MgO at conditions of Earth’s interior [Geophysics]

Tang, X., Dong, J. — Tue, 09 Mar 2010 17:27:18 +0000

Thermal conductivity of the Earth’s lower mantle greatly impacts the mantle convection style and affects the heat conduction from the…

Atrazine induces complete feminization and chemical castration in male African clawed frogs (Xenopus laevis) [Ecology]

Tue, 09 Mar 2010 17:27:18 +0000

The herbicide atrazine is one of the most commonly applied pesticides in the world. As a result, atrazine is the…

Inaugural Article: The Lucretian swerve: The biological basis of human behavior and the criminal justice system [Neuroscience]

Cashmore, A. R. — Tue, 09 Mar 2010 17:27:17 +0000

It is widely believed, at least in scientific circles, that living systems, including mankind, obey the natural physical laws. However,…

Inaugural Article: Some features of the spread of epidemics and information on a random graph [Applied_Mathematics]

Durrett, R. — Tue, 09 Mar 2010 17:27:17 +0000

Random graphs are useful models of social and technological networks. To date, most of the research in this area has…

Rontgen’s electrode-free elastomer actuators without electromechanical pull-in instability [Applied_Physical_Sciences]

Keplinger, C., Kaltenbrunner, M., Arnold, N., Bauer, S. — Tue, 09 Mar 2010 17:27:17 +0000

Electrical actuators made from films of dielectric elastomers coated on both sides with stretchable electrodes may potentially be applied in…

Bioorganometallic mechanism of action, and inhibition, of IspH [Chemistry]

Wang, W., Wang, K., Liu, Y.-L., No, J.-H., Li, J., Nilges, M. J., Oldfield, E. — Tue, 09 Mar 2010 17:27:17 +0000

We have investigated the mechanism of action of Aquifex aeolicus IspH [E-4-hydroxy-3-methyl-but-2-enyl diphosphate (HMBPP) reductase], together with its inhibition, using…

Solving the apparent diversity-accuracy dilemma of recommender systems [Applied_Physical_Sciences]

Zhou, T., Kuscsik, Z., Liu, J.-G., Medo, M., Wakeling, J. R., Zhang, Y.-C. — Tue, 09 Mar 2010 17:27:17 +0000

Recommender systems use data on past user preferences to predict possible future likes and interests. A key challenge is that…

Nanowire platform for mapping neural circuits [Commentary]

Xie, C., Cui, Y. — Tue, 09 Mar 2010 17:27:17 +0000

Fermentable sugars by chemical hydrolysis of biomass [Applied_Biological_Sciences]

Binder, J. B., Raines, R. T. — Tue, 09 Mar 2010 17:27:17 +0000

Abundant plant biomass has the potential to become a sustainable source of fuels and chemicals. Realizing this potential requires the…

In This Issue [This Week in PNAS]

Tue, 09 Mar 2010 17:27:17 +0000

Molecular basis of cyclooxygenase enzymes (COXs) selective inhibition [Biophysics_And_Computational_Biology]

Tue, 09 Mar 2010 16:03:35 +0000

The widely used nonsteroidal anti-inflammatory drugs block the cyclooxygenase enzymes (COXs) and are clinically used for the treatment of inflammation,…

Predicting organic hydrogen atom transfer rate constants using the Marcus cross relation [Biochemistry]

Warren, J. J., Mayer, J. M. — Tue, 09 Mar 2010 16:03:35 +0000

Chemical reactions that involve net hydrogen atom transfer (HAT) are ubiquitous in chemistry and biology, from the action of antioxidants…

Superheating of ice crystals in antifreeze protein solutions [Biophysics_And_Computational_Biology]

Celik, Y., Graham, L. A., Mok, Y.-F., Bar, M., Davies, P. L., Braslavsky, I. — Tue, 09 Mar 2010 16:03:34 +0000

It has been argued that for antifreeze proteins (AFPs) to stop ice crystal growth, they must irreversibly bind to the…

Reverse Engineering of Bacterial Chemotaxis Pathway via Frequency Domain Analysis

Nicolas Guex et al. — Tue, 09 Mar 2010 08:00:00 +0000

Chemotaxis is defined as a behavior involving organisms sensing attractants or repellents and leading towards or away from them. Therefore, it is possible to reengineer chemotaxis network to control the movement of bacteria to our advantage. Understanding the design principles of chemotaxis pathway is a prerequisite and an important topic in synthetic biology. Here, we provide guidelines for chemotaxis pathway design by employing control theory and reverse engineering concept on pathway dynamic design. We first analyzed the mathematical models for two most important kinds of E. coli chemotaxis pathway—adaptive and non-adaptive pathways, and concluded that the control units of the pathway de facto function as a band-pass filter and a low-pass filter, respectively, by abstracting the frequency response properties of the pathways. The advantage of the band-pass filter is established, and we demonstrate how to tune the three key parameters of it—A (max amplification), ω₁ (down cut-off frequency) and ω₂ (up cut-off frequency) to optimize the chemotactic effect. Finally, we hypothesized a similar but simpler version of the dynamic pathway model based on the principles discovered and show that it leads to similar properties with native E. coli chemotactic behaviors. Our study provides an example of simulating and designing biological dynamics in silico and indicates how to make use of the native pathway’s features in this process. Furthermore, the characteristics we discovered and tested through reverse engineering may help to understand the design principles of the pathway and promote the design of artificial chemotaxis pathways.

BAAV Transcytosis Requires an Interaction with β-1-4 Linked- Glucosamine and gp96

Nicolas Guex et al. — Tue, 09 Mar 2010 08:00:00 +0000

Cell surface carbohydrates play an important role in virus entry and intracellular trafficking. Bovine Adeno-Associated Virus (BAAV) uses plasma membrane gangliosides for transduction and infection. In addition, independent of the infectious pathway, BAAV also has the ability to pass through barrier epithelia and endothelia using a transcytosis pathway dependent upon the presence of cell surface carbohydrates. Thus, in order to better define the carbohydrate interactions that are necessary for BAAV infection or transcytosis, a glycan microarray composed of both natural and synthetic carbohydrates was probed with HA-tagged BAAV particles. This identified chitotriose, a trimer of β-1-4-linked N-acetyl glucosamine, as having an interaction with BAAV. Competition experiments showed that the BAAV interaction with this carbohydrate is not necessary for infection but is instead important in the transcytosis pathway. The β-1-4-linked N-acetyl glucosamine modification has been reported on gp96, a glycoprotein involved in the transcytosis of bacteria and toxins. Significantly, immunoprecipitation and competition experiments with an anti-gp96 antibody and a soluble form of gp96, respectively, showed this glycoprotein can also interact with BAAV to serve as a receptor for its transcytosis.

Computational Study of Evolutionary Selection Pressure on Rainbow Trout Estrogen Receptors

Nicolas Guex et al. — Tue, 09 Mar 2010 08:00:00 +0000

Molecular dynamics simulations were used to determine the binding affinities between the hormone 17-estradiol (E2) and different estrogen receptor (ER) isoforms in the rainbow trout, Oncorhynchus mykiss. Previous phylogenetic analysis indicates that a whole genome duplication prior to the divergence of ray-finned fish led to two distinct ER isoforms, ER and ER, and the recent whole genome duplication in the ancestral salmonid created two ER isoforms, ER and ER. The objective of our computational studies is to provide insight into the underlying evolutionary pressures on these isoforms. For the ER subtype our results show that E2 binds preferentially to ER over ER. Tests of lineage specific N/S ratios indicate that the ligand binding domain of the ER gene is evolving under relaxed selection relative to all other ER genes. Comparison with the highly conserved DNA binding domain suggests that ER may be undergoing neofunctionalization possibly by binding to another ligand. By contrast, both ER and ER bind similarly to E2 and the best fitting model of selection indicates that the ligand binding domain of all ER genes are evolving under the same level of purifying selection, comparable to ER.