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1.  How do environmental factors influence life cycles and development? An experimental framework for early-diverging metazoans 
Ecological developmental biology (eco-devo) explores the mechanistic relationships between the processes of individual development and environmental factors. Recent studies imply that some of these relationships have deep evolutionary origins, and may even predate the divergences of the simplest extant animals, including cnidarians and sponges. Development of these early diverging metazoans is often sensitive to environmental factors, and these interactions occur in the context of conserved signaling pathways and mechanisms of tissue homeostasis whose detailed molecular logic remain elusive. Efficient methods for transgenesis in cnidarians together with the ease of experimental manipulation in cnidarians and sponges make them ideal models for understanding causal relationships between environmental factors and developmental mechanisms. Here, we identify major questions at the interface between animal evolution and development and outline a road map for research aimed at identifying the mechanisms that link environmental factors to developmental mechanisms in early diverging metazoans.
doi:10.1002/bies.201400065
PMCID: PMC4251966  PMID: 25205353
Cnidaria; corals; environmental genomics; holobiont; hydra; sponges; symbiosis
2.  Optimization of Novel Indole-2-carboxamide Inhibitors of Neurotropic Alphavirus Replication 
Journal of medicinal chemistry  2013;56(22):9222-9241.
Neurotropic alphaviruses, which include western equine encephalitis virus (WEEV) and Fort Morgan virus, are mosquito-borne pathogens that infect the central nervous system causing acute and potentially fatal encephalitis. We previously reported a novel series of indole-2-carboxamides as alphavirus replication inhibitors, one of which conferred protection against neuroadapted Sindbis virus infection in mice. We describe here further development of this series resulting in 10-fold improvement in potency in a WEEV replicon assay and up to 40-fold increases in half-lives in mouse liver microsomes. Using a rhodamine123 uptake assay in MDR1-MDCKII cells we were able to identify structural modifications that markedly reduce recognition by P-glycoprotein, the key efflux transporter at the blood brain barrier. In a preliminary mouse PK study we were able to demonstrate that two new analogs could achieve higher and/or longer plasma drug exposures than our previous lead, and that one compound achieved measurable drug levels in the brain.
doi:10.1021/jm401330r
PMCID: PMC3895407  PMID: 24151954
antiviral; alphavirus; indole; central nervous system; encephalitis; RNA replication inhibitor; metabolic stability; P-glycoprotein
3.  T2 Texture Index of Cartilage Can Predict Early Symptomatic OA Progression: Data from the Osteoarthritis Initiative 
Objective
There is an interest in using Magnetic Resonance Imaging (MRI) to identify pre-radiographic changes in osteoarthritis (OA) and features that indicate risk for disease progression. The purpose of this study is to identify image features derived from MRI T2 maps that can accurately predict onset of OA symptoms in subjects at risk for incident knee OA.
Methods
Patients were selected from the Osteoarthritis Initiative (OAI) control cohort and incidence cohort and stratified based on the change in total WOMAC score from baseline to three year follow-up (80 non-OA progression and 88 symptomatic OA progression patients). For each patient, a series of image texture features were measured from the baseline cartilage T2 map. A linear discriminant function and feature reduction method was then trained to quantify a texture metric, the T2 texture index of cartilage (TIC), based on 22 image features, to identify a composite marker of T2 heterogeneity.
Results
Statistically significant differences were seen in the baseline T2 TIC between the non-progression and symptomatic OA progression populations. The baseline T2 TIC differentiates subjects that develop worsening of their WOMAC score OA with an accuracy between 71% and 76%. The T2 TIC differences were predominantly localized to a dominant knee compartment that correlated with the mechanical axis of the knee.
Conclusion
Baseline heterogeneity in cartilage T2 as measured with the T2 TIC index is able to differentiate and predict individuals that will develop worsening of their WOMAC score at 3-year follow-up.
doi:10.1016/j.joca.2013.06.007
PMCID: PMC3779506  PMID: 23774471
Cartilage T2; T2 heterogeneity; osteoarthritis; MRI; image biomarkers; signal texture
4.  Basigin null mutant male mice are sterile and exhibit impaired interactions between germ cells and Sertoli cells 
Developmental biology  2013;380(2):145-156.
Basigin (BSG) is a multifunctional glycoprotein that plays an important role in male reproduction since male knockout (KO) mice are sterile. The Bsg KO testis lacks elongated spermatids and mature spermatozoa, a phenotype similar to that of alpha-mannosidase IIx (MX) KO mice. MX regulates formation of N-acetylglucosamine (GlcNAc) terminated N-glycans that participate in germ cell-Sertoli cell adhesion. Results showed that Bsg KO spermatocytes displayed normal homologous chromosome synapsis and progression through meiosis. However, only punctate expression of the round spermatid marker SP-10 in the acrosomal granule of germ cells of Bsg KO mice was detected indicating that spermatogenesis in Bsg KO mice was arrested at the early round spermatid stages. We observed a large increase in the number of germ cells undergoing apoptosis in Bsg KO testes. Using lectin blotting, we determined that GlcNAc terminated N-glycans are linked to BSG. GlcNAc terminated N-glycans were significantly reduced in Bsg KO testes. These observations indicate that BSG may act as a germ cell-Sertoli cell attachment molecule. Loss of BSG significantly reduced adhesion between GC-2 and SF7 cells. Moreover, wild type testes showed strong expression of N-cadherin (CDH2) while expression was greatly reduced in the testes of Bsg KO mice. In addition, the integrity of the blood-testis barrier (BTB) was compromised in Bsg KO testes. In conclusion, although some Bsg KO spermatogonia can undergo normal progression to the spermatocyte stage, BSG-mediated germ cell-Sertoli cell interactions appear to be necessary for integrity of the BTB and spermatocyte progression to mature spermatozoa.
doi:10.1016/j.ydbio.2013.05.023
PMCID: PMC3778672  PMID: 23727514
Basigin; BSG; N-glycans; α-Mannosidase II; Spermatogenesis; Blood-testis barrier
5.  Nonlinear System Modeling with Random Matrices: Echo State Networks Revisited 
Echo state networks (ESNs) are a novel form of recurrent neural networks (RNNs) that provide an efficient and powerful computational model approximating nonlinear dynamical systems. A unique feature of an ESN is that a large number of neurons (the “reservoir”) are used, whose synaptic connections are generated randomly, with only the connections from the reservoir to the output modified by learning. Why a large randomly generated fixed RNN gives such excellent performance in approximating nonlinear systems is still not well understood. In this brief, we apply random matrix theory to examine the properties of random reservoirs in ESNs under different topologies (sparse or fully connected) and connection weights (Bernoulli or Gaussian). We quantify the asymptotic gap between the scaling factor bounds for the necessary and sufficient conditions previously proposed for the echo state property. We then show that the state transition mapping is contractive with high probability when only the necessary condition is satisfied, which corroborates and thus analytically explains the observation that in practice one obtains echo states when the spectral radius of the reservoir weight matrix is smaller than 1.
doi:10.1109/TNNLS.2011.2178562
PMCID: PMC4107715  PMID: 24808467
Circular law; concentration of measure; echo state networks; echo state property; random matrix theory; recurrent neural networks
6.  The “Naked Coral” Hypothesis Revisited – Evidence for and Against Scleractinian Monophyly 
PLoS ONE  2014;9(4):e94774.
The relationship between Scleractinia and Corallimorpharia, Orders within Anthozoa distinguished by the presence of an aragonite skeleton in the former, is controversial. Although classically considered distinct groups, some phylogenetic analyses have placed the Corallimorpharia within a larger Scleractinia/Corallimorpharia clade, leading to the suggestion that the Corallimorpharia are “naked corals” that arose via skeleton loss during the Cretaceous from a Scleractinian ancestor. Scleractinian paraphyly is, however, contradicted by a number of recent phylogenetic studies based on mt nucleotide (nt) sequence data. Whereas the “naked coral” hypothesis was based on analysis of the sequences of proteins encoded by a relatively small number of mt genomes, here a much-expanded dataset was used to reinvestigate hexacorallian phylogeny. The initial observation was that, whereas analyses based on nt data support scleractinian monophyly, those based on amino acid (aa) data support the “naked coral” hypothesis, irrespective of the method and with very strong support. To better understand the bases of these contrasting results, the effects of systematic errors were examined. Compared to other hexacorallians, the mt genomes of “Robust” corals have a higher (A+T) content, codon usage is far more constrained, and the proteins that they encode have a markedly higher phenylalanine content, leading us to suggest that mt DNA repair may be impaired in this lineage. Thus the “naked coral” topology could be caused by high levels of saturation in these mitochondrial sequences, long-branch effects or model violations. The equivocal results of these extensive analyses highlight the fundamental problems of basing coral phylogeny on mitochondrial sequence data.
doi:10.1371/journal.pone.0094774
PMCID: PMC3989238  PMID: 24740380
7.  Neurotropic Arboviruses Induce Interferon Regulatory Factor 3-Mediated Neuronal Responses That Are Cytoprotective, Interferon Independent, and Inhibited by Western Equine Encephalitis Virus Capsid 
Journal of Virology  2013;87(3):1821-1833.
Cell-intrinsic innate immune responses mediated by the transcription factor interferon regulatory factor 3 (IRF-3) are often vital for early pathogen control, and effective responses in neurons may be crucial to prevent the irreversible loss of these critical central nervous system cells after infection with neurotropic pathogens. To investigate this hypothesis, we used targeted molecular and genetic approaches with cultured neurons to study cell-intrinsic host defense pathways primarily using the neurotropic alphavirus western equine encephalitis virus (WEEV). We found that WEEV activated IRF-3-mediated neuronal innate immune pathways in a replication-dependent manner, and abrogation of IRF-3 function enhanced virus-mediated injury by WEEV and the unrelated flavivirus St. Louis encephalitis virus. Furthermore, IRF-3-dependent neuronal protection from virus-mediated cytopathology occurred independently of autocrine or paracrine type I interferon activity. Despite being partially controlled by IRF-3-dependent signals, WEEV also disrupted antiviral responses by inhibiting pattern recognition receptor pathways. This antagonist activity was mapped to the WEEV capsid gene, which disrupted signal transduction downstream of IRF-3 activation and was independent of capsid-mediated inhibition of host macromolecular synthesis. Overall, these results indicate that innate immune pathways have important cytoprotective activity in neurons and contribute to limiting injury associated with infection by neurotropic arboviruses.
doi:10.1128/JVI.02858-12
PMCID: PMC3554193  PMID: 23192868
8.  Unsupervised, low latency anomaly detection of algorithmically generated domain names by generative probabilistic modeling 
Journal of Advanced Research  2014;5(4):423-433.
We propose a method for detecting anomalous domain names, with focus on algorithmically generated domain names which are frequently associated with malicious activities such as fast flux service networks, particularly for bot networks (or botnets), malware, and phishing. Our method is based on learning a (null hypothesis) probability model based on a large set of domain names that have been white listed by some reliable authority. Since these names are mostly assigned by humans, they are pronounceable, and tend to have a distribution of characters, words, word lengths, and number of words that are typical of some language (mostly English), and often consist of words drawn from a known lexicon. On the other hand, in the present day scenario, algorithmically generated domain names typically have distributions that are quite different from that of human-created domain names. We propose a fully generative model for the probability distribution of benign (white listed) domain names which can be used in an anomaly detection setting for identifying putative algorithmically generated domain names. Unlike other methods, our approach can make detections without considering any additional (latency producing) information sources, often used to detect fast flux activity. Experiments on a publicly available, large data set of domain names associated with fast flux service networks show encouraging results, relative to several baseline methods, with higher detection rates and low false positive rates.
doi:10.1016/j.jare.2014.01.001
PMCID: PMC4294760
Anomaly detection; Algorithmically generated domain names; Malicious domain names; Domain name modeling; Fast flux
9.  Discovery of Potent Broad Spectrum Antivirals Derived from Marine Actinobacteria 
PLoS ONE  2013;8(12):e82318.
Natural products provide a vast array of chemical structures to explore in the discovery of new medicines. Although secondary metabolites produced by microbes have been developed to treat a variety of diseases, including bacterial and fungal infections, to date there has been limited investigation of natural products with antiviral activity. In this report, we used a phenotypic cell-based replicon assay coupled with an iterative biochemical fractionation process to identify, purify, and characterize antiviral compounds produced by marine microbes. We isolated a compound from Streptomyces kaviengensis, a novel actinomycetes isolated from marine sediments obtained off the coast of New Ireland, Papua New Guinea, which we identified as antimycin A1a. This compound displays potent activity against western equine encephalitis virus in cultured cells with half-maximal inhibitory concentrations of less than 4 nM and a selectivity index of greater than 550. Our efforts also revealed that several antimycin A analogues display antiviral activity, and mechanism of action studies confirmed that these Streptomyces-derived secondary metabolites function by inhibiting the cellular mitochondrial electron transport chain, thereby suppressing de novo pyrimidine synthesis. Furthermore, we found that antimycin A functions as a broad spectrum agent with activity against a wide range of RNA viruses in cultured cells, including members of the Togaviridae, Flaviviridae, Bunyaviridae, Picornaviridae, and Paramyxoviridae families. Finally, we demonstrate that antimycin A reduces central nervous system viral titers, improves clinical disease severity, and enhances survival in mice given a lethal challenge with western equine encephalitis virus. Our results provide conclusive validation for using natural product resources derived from marine microbes as source material for antiviral drug discovery, and they indicate that host mitochondrial electron transport is a viable target for the continued development of broadly active antiviral compounds.
doi:10.1371/journal.pone.0082318
PMCID: PMC3857800  PMID: 24349254
10.  Correction: Coral Thermal Tolerance: Tuning Gene Expression to Resist Thermal Stress 
PLoS ONE  2013;8(10):10.1371/annotation/2b623dcd-9413-47b0-990d-c46c8e662ad4.
doi:10.1371/annotation/2b623dcd-9413-47b0-990d-c46c8e662ad4
PMCID: PMC3805605
11.  The Schistosoma japonicum genome reveals features of host-parasite interplay 
Nature  2009;460(7253):345-351.
Schistosoma japonicum is a parasitic flatworm that causes human schistosomiasis, a significant cause of morbidity in China and the Philippines. Here we present a draft genomic sequence for the worm, which is the first reported for any flatworm, indeed for the superphylum Lophotrochozoa. The genome provides a global insight into the molecular architecture and host interaction of this complex metazoan pathogen, revealing that it can exploit host nutrients, neuroendocrine hormones and signaling pathways for growth, development and maturation. Having a complex nervous system and a well developed sensory system, S. japonicum can accept stimulation of the corresponding ligands as a physiological response to different environments, such as fresh water or the tissues of its intermediate and mammalian hosts. Numerous proteinases, including cercarial elastase, are implicated in mammalian skin penetration and haemoglobin degradation. The genomic information will serve as a valuable platform to facilitate development of new interventions for schistosomiasis control.
doi:10.1038/nature08140
PMCID: PMC3747554  PMID: 19606140
12.  Male Mice That Do Not Express Group VIA Phospholipase A2 Produce Spermatozoa with Impaired Motility and Have Greatly Reduced Fertility* 
The Journal of biological chemistry  2004;279(37):38194-38200.
The Group VIA Phospholipase A2 (iPLA2β) is the first recognized cytosolic Ca2+-independent PLA2 and has been proposed to participate in arachidonic acid (20:4) incorporation into glycerophosphocholine lipids, cell proliferation, exocytosis, apoptosis, and other processes. To study iPLA2β functions, we disrupted its gene by homologous recombination to generate mice that do not express iPLA2β. Heterozygous iPLA2β+/− breeding pairs yield a Mendelian 1:2:1 ratio of iPLA2β+/+, iPLA2β+/−, and iPLA2β−/− pups and a 1:1 male:female gender distribution of iPLA2β−/− pups. Several tissues of wild-type mice express iPLA2β mRNA, immunoreactive protein, and activity, and testes express the highest levels. Testes or other tissues of iPLA2β−/− mice express no iPLA2β mRNA or protein, but iPLA2β−/− testes are not deficient in 20:4-containing glycerophosphocholine lipids, indicating that iPLA2β does not play an obligatory role in formation of such lipids in that tissue. Spermatozoa from iPLA2β−/− mice have reduced motility and impaired ability to fertilize mouse oocytes in vitro and in vivo, and inhibiting iPLA2β with a bromoenol lactone suicide substrate reduces motility of wild-type spermatozoa in a time- and concentration-dependent manner.Mating iPLA2β−/− male mice with iPLA2β+/+, iPLA2β+/−, or iPLA2β−/− female mice yields only about 7% of the number of pups produced by mating pairs with an iPLA2β+/+ or iPLA2β+/− male, but iPLA2β−/− female mice have nearly normal fertility. These findings indicate that iPLA2β plays an important functional role in spermatozoa, suggest a target for developing male contraceptive drugs, and complement reports that disruption of the Group IVA PLA2 (cPLA2α) gene impairs female reproductive ability.
doi:10.1074/jbc.M406489200
PMCID: PMC3733543  PMID: 15252026
13.  The acute transcriptional response of the coral Acropora millepora to immune challenge: expression of GiMAP/IAN genes links the innate immune responses of corals with those of mammals and plants 
BMC Genomics  2013;14:400.
Background
As a step towards understanding coral immunity we present the first whole transcriptome analysis of the acute responses of Acropora millepora to challenge with the bacterial cell wall derivative MDP and the viral mimic poly I:C, defined immunogens provoking distinct but well characterised responses in higher animals.
Results
These experiments reveal similarities with the responses both of arthropods and mammals, as well as coral-specific effects. The most surprising finding was that MDP specifically induced three members of the GiMAP gene family, which has been implicated in immunity in mammals but is absent from Drosophila and Caenorhabditis. Like their mammalian homologs, GiMAP genes are arranged in a tandem cluster in the coral genome.
Conclusions
A phylogenomic survey of this gene family implies ancient origins, multiple independent losses and lineage-specific expansions during animal evolution. Whilst functional convergence cannot be ruled out, GiMAP expression in corals may reflect an ancestral role in immunity, perhaps in phagolysosomal processing.
doi:10.1186/1471-2164-14-400
PMCID: PMC3723955  PMID: 23768317
Innate immunity; Evolution; GTPase; Coral disease; Cnidaria; Transcriptomics
14.  The Skeletal Proteome of the Coral Acropora millepora: The Evolution of Calcification by Co-Option and Domain Shuffling 
Molecular Biology and Evolution  2013;30(9):2099-2112.
In corals, biocalcification is a major function that may be drastically affected by ocean acidification (OA). Scleractinian corals grow by building up aragonitic exoskeletons that provide support and protection for soft tissues. Although this process has been extensively studied, the molecular basis of biocalcification is poorly understood. Notably lacking is a comprehensive catalog of the skeleton-occluded proteins—the skeletal organic matrix proteins (SOMPs) that are thought to regulate the mineral deposition. Using a combination of proteomics and transcriptomics, we report the first survey of such proteins in the staghorn coral Acropora millepora. The organic matrix (OM) extracted from the coral skeleton was analyzed by mass spectrometry and bioinformatics, enabling the identification of 36 SOMPs. These results provide novel insights into the molecular basis of coral calcification and the macroevolution of metazoan calcifying systems, whereas establishing a platform for studying the impact of OA at molecular level. Besides secreted proteins, extracellular regions of transmembrane proteins are also present, suggesting a close control of aragonite deposition by the calicoblastic epithelium. In addition to the expected SOMPs (Asp/Glu-rich, galaxins), the skeletal repertoire included several proteins containing known extracellular matrix domains. From an evolutionary perspective, the number of coral-specific proteins is low, many SOMPs having counterparts in the noncalcifying cnidarians. Extending the comparison with the skeletal OM proteomes of other metazoans allowed the identification of a pool of functional domains shared between phyla. These data suggest that co-option and domain shuffling may be general mechanisms by which the trait of calcification has evolved.
doi:10.1093/molbev/mst109
PMCID: PMC3748352  PMID: 23765379
biomineralization; calcium carbonate skeleton; scleractinian; proteomics; evolution
15.  Novel Inhibitors of Neurotropic Alphavirus Replication That Improve Host Survival in a Mouse Model of Acute Viral Encephalitis 
Journal of Medicinal Chemistry  2012;55(7):3535-3545.
Arboviral encephalitis is a potentially devastating human disease with no approved therapies that target virus replication. We previously discovered a novel class of thieno[3,2-b]pyrrole-based inhibitors active against neurotropic alphaviruses such as western equine encephalitis virus (WEEV) in cultured cells. In this report we describe initial development of these novel antiviral compounds, including bioisosteric replacement of the 4H-thieno[3,2-b]pyrrole core with indole to improve metabolic stability and the introduction of chirality to assess target enantioselectivity. Selected modifications enhanced antiviral activity while maintaining low cytotoxicity, increased stability to microsomal metabolism, and also revealed striking enantiospecific activity in cultured cells. Furthermore, we demonstrate improved outcomes (both symptoms and survival) following treatment with indole analog 9h (CCG-203926) in an in vivo mouse model of alphaviral encephalitis that closely correlate with the enantiospecific in vitro antiviral activity. These results represent a substantial advancement in the early preclinical development of a promising class of novel antiviral drugs against virulent neurotropic alphaviruses.
doi:10.1021/jm300214e
PMCID: PMC3329717  PMID: 22428985
antiviral; alphavirus; indole; central nervous system; encephalitis; RNA replication inhibitor
16.  Activation of the Type I Interferon Pathway Is Enhanced in Response to Human Neuronal Differentiation 
PLoS ONE  2013;8(3):e58813.
Despite the crucial role of innate immunity in preventing or controlling pathogen-induced damage in most, if not all, cell types, very little is known about the activity of this essential defense system in central nervous system neurons, especially in humans. In this report we use both an established neuronal cell line model and an embryonic stem cell-based system to examine human neuronal innate immunity and responses to neurotropic alphavirus infection in cultured cells. We demonstrate that neuronal differentiation is associated with increased expression of crucial type I interferon signaling pathway components, including interferon regulatory factor-9 and an interferon receptor heterodimer subunit, which results in enhanced interferon stimulation and subsequent heightened antiviral activity and cytoprotective responses against neurotropic alphaviruses such as western equine encephalitis virus. These results identify important differentiation-dependent changes in innate immune system function that control cell-autonomous neuronal responses. Furthermore, this work demonstrates the utility of human embryonic stem cell-derived cultures as a platform to study the interactions between innate immunity, virus infection, and pathogenesis in central nervous system neurons.
doi:10.1371/journal.pone.0058813
PMCID: PMC3591356  PMID: 23505563
17.  Human Neuronal Cells Possess Functional Cytoplasmic and Toll-like Receptor-Mediated Innate Immune Pathways Influenced by Phosphatidylinositol-3 Kinase Signaling1 
Innate immune pathways are early defense responses important for the immediate control and eventual clearance of many pathogens, where signaling is initiated via pattern recognition receptor-mediated events that occur in a ligand- and cell-type specific manner. Within CNS neurons, innate immune pathways are likely crucial to control pathogens that target these essential yet virtually irreplaceable cells. However, relatively little is known about the induction and regulation of neuronal pattern recognition receptor signaling. In this report, we used human neuronal cell lines and primary rat neuronal cultures to examine pattern recognition receptor expression and function. We found that several innate immune receptor ligands, including Sendai virus, the dsRNA mimetic polyinosinic-polycytidylic acid, and LPS all activated differentiation-dependent neuronal innate immune pathways. Functional genetic analyses revealed that interferon regulatory factor 3-mediated pathways that resulted in IFNβ transcriptional upregulation were activated in cultured human neuronal cells by the pattern recognition receptors TLR3, melanoma differentiated-associated gene 5, or retinoic acid inducible gene I in a ligand-specific manner. Furthermore, genome-wide transcriptional array and targeted genetic and pharmacologic analyses identified PI3K signaling as crucial for the induction of innate immune pathways in neurons. These results indicate that human neuronal cells possess specific and functional pattern recognition receptor pathways essential for the effective induction of innate immune responses, and suggest that neurons can play an active role in defense against neurotropic pathogens.
This is an author-produced version of a manuscript accepted for publication in The Journal of Immunology (The JI). The American Association of Immunologists, Inc. (AAI), publisher of The JI, holds the copyright to this manuscript. This manuscript has not yet been copyedited or subjected to editorial proofreading by The JI; hence it may differ from the final version published in The JI (online and in print). AAI (The JI) is not liable for errors or omissions in the author-produced version on the manuscript or in any version derived from it by the United States National Institutes of Health or any other third party. The final, citable version of record can be found at www.jimmunol.org.
doi:10.4049/jimmunol.0904133
PMCID: PMC2887731  PMID: 20483728
18.  Margin-Maximizing Feature Elimination Methods for Linear and Nonlinear Kernel-Based Discriminant Functions 
Feature selection for classification in high-dimensional spaces can improve generalization, reduce classifier complexity, and identify important, discriminating feature “markers.” For support vector machine (SVM) classification, a widely used technique is recursive feature elimination (RFE). We demonstrate that RFE is not consistent with margin maximization, central to the SVM learning approach. We thus propose explicit margin-based feature elimination (MFE) for SVMs and demonstrate both improved margin and improved generalization, compared with RFE. Moreover, for the case of a nonlinear kernel, we show that RFE assumes that the squared weight vector 2-norm is strictly decreasing as features are eliminated. We demonstrate this is not true for the Gaussian kernel and, consequently, RFE may give poor results in this case. MFE for nonlinear kernels gives better margin and generalization. We also present an extension which achieves further margin gains, by optimizing only two degrees of freedom—the hyperplane’s intercept and its squared 2-norm—with the weight vector orientation fixed. We finally introduce an extension that allows margin slackness. We compare against several alternatives, including RFE and a linear programming method that embeds feature selection within the classifier design. On high-dimensional gene microarray data sets, University of California at Irvine (UCI) repository data sets, and Alzheimer’s disease brain image data, MFE methods give promising results.
doi:10.1109/TNN.2010.2041069
PMCID: PMC3580203  PMID: 20194055
Alzheimer’s; classifier margin; discriminant function; feature elimination; Gaussian kernel; margin maximization; medical imaging; microarray; magnetic resonance imaging (MRI); neurodegenerative; polynomial kernel; recursive feature elimination (RFE); support vector machine (SVM)
19.  An Overview of Population Genetic Data Simulation 
Abstract
Simulation studies in population genetics play an important role in helping to better understand the impact of various evolutionary and demographic scenarios on sequence variation and sequence patterns, and they also permit investigators to better assess and design analytical methods in the study of disease-associated genetic factors. To facilitate these studies, it is imperative to develop simulators with the capability to accurately generate complex genomic data under various genetic models. Currently, a number of efficient simulation software packages for large-scale genomic data are available, and new simulation programs with more sophisticated capabilities and features continue to emerge. In this article, we review the three basic simulation frameworks—coalescent, forward, and resampling—and some of the existing simulators that fall under these frameworks, comparing them with respect to their evolutionary and demographic scenarios, their computational complexity, and their specific applications. Additionally, we address some limitations in current simulation algorithms and discuss future challenges in the development of more powerful simulation tools.
doi:10.1089/cmb.2010.0188
PMCID: PMC3244809  PMID: 22149682
backward simulators; disease association study; forward simulators; genome simulation; resampling
20.  The Neuronal Calcium Sensor Protein Acrocalcin: A Potential Target of Calmodulin Regulation during Development in the Coral Acropora millepora 
PLoS ONE  2012;7(12):e51689.
To understand the calcium-mediated signalling pathways underlying settlement and metamorphosis in the Scleractinian coral Acropora millepora, a predicted protein set derived from larval cDNAs was scanned for the presence of EF-hand domains (Pfam Id: PF00036). This approach led to the identification of a canonical calmodulin (AmCaM) protein and an uncharacterised member of the Neuronal Calcium Sensor (NCS) family of proteins known here as Acrocalcin (AmAC). While AmCaM transcripts were present throughout development, AmAC transcripts were not detected prior to gastrulation, after which relatively constant mRNA levels were detected until metamorphosis and settlement. The AmAC protein contains an internal CaM-binding site and was shown to interact in vitro with AmCaM. These results are consistent with the idea that AmAC is a target of AmCaM in vivo, suggesting that this interaction may regulate calcium-dependent processes during the development of Acropora millepora.
doi:10.1371/journal.pone.0051689
PMCID: PMC3524228  PMID: 23284743
21.  Maturation-dependent responses of human neuronal cells to western equine encephalitis virus infection and type I interferons 
Virology  2007;372(1):208-220.
Innate cell-autonomous antiviral responses are essential first lines of defense against central nervous system infections but may also contribute to neuropathogenesis. We investigated the relationships between innate immunity and neuronal differentiation using an in vitro culture system with human cell lines to analyze cellular responses to the neurotropic alphavirus western equine encephalitis virus. Human neuronal cells displayed a maturation-dependent reduction in virus-induced cytopathology that was independent of autocrine interferon α or β activity. In addition, maturation was associated with enhanced responsiveness to exogenous stimuli, such that differentiated neurons required five- to ten-fold less type I interferon to suppress viral replication or virus-induced cytopathology compared to immature cells, although this enhanced responsiveness extended to only a subset of unique type I interferons. These results demonstrate that maturation-dependent changes in human neuronal cells may be key determinants in the innate immune response to infections with neurotropic alphaviruses.
doi:10.1016/j.virol.2007.10.025
PMCID: PMC3518402  PMID: 18022665
Neurons; Differentiation; Alphavirus; Interferon
22.  Coral Thermal Tolerance: Tuning Gene Expression to Resist Thermal Stress 
PLoS ONE  2012;7(11):e50685.
The acclimatization capacity of corals is a critical consideration in the persistence of coral reefs under stresses imposed by global climate change. The stress history of corals plays a role in subsequent response to heat stress, but the transcriptomic changes associated with these plastic changes have not been previously explored. In order to identify host transcriptomic changes associated with acquired thermal tolerance in the scleractinian coral Acropora millepora, corals preconditioned to a sub-lethal temperature of 3°C below bleaching threshold temperature were compared to both non-preconditioned corals and untreated controls using a cDNA microarray platform. After eight days of hyperthermal challenge, conditions under which non-preconditioned corals bleached and preconditioned corals (thermal-tolerant) maintained Symbiodinium density, a clear differentiation in the transcriptional profiles was revealed among the condition examined. Among these changes, nine differentially expressed genes separated preconditioned corals from non-preconditioned corals, with 42 genes differentially expressed between control and preconditioned treatments, and 70 genes between non-preconditioned corals and controls. Differentially expressed genes included components of an apoptotic signaling cascade, which suggest the inhibition of apoptosis in preconditioned corals. Additionally, lectins and genes involved in response to oxidative stress were also detected. One dominant pattern was the apparent tuning of gene expression observed between preconditioned and non-preconditioned treatments; that is, differences in expression magnitude were more apparent than differences in the identity of genes differentially expressed. Our work revealed a transcriptomic signature underlying the tolerance associated with coral thermal history, and suggests that understanding the molecular mechanisms behind physiological acclimatization would be critical for the modeling of reefs in impending climate change scenarios.
doi:10.1371/journal.pone.0050685
PMCID: PMC3511300  PMID: 23226355
23.  Identification of Thieno[3,2-b]Pyrrole Derivatives as Novel Small Molecule Inhibitors of Neurotropic Alphaviruses 
The Journal of infectious diseases  2009;199(7):950-957.
Neurotropic alphaviruses such as western, eastern, and Venezuelan equine encephalitis viruses cause serious and potentially fatal central nervous system infections in humans and are high-priority potential bioterrorism agents. There are currently no widely available vaccines or licensed therapies for these virulent pathogens. To identify potential novel antiviral drugs, we developed a cell-based assay with a western equine encephalitis virus replicon that expresses a luciferase reporter gene and screened a small molecule diversity library of 51,028 compounds. We identified and validated a thieno[3,2-b]pyrrole compound with a half maximal inhibitory concentration of <10 µmol/L, a selectivity index >20, and potent activity against live virus in cultured neuronal cells. Furthermore, a structure-activity relationship analysis with 20 related compounds identified several with enhanced activity profiles, including 6 with submicromolar half maximal inhibitory concentrations. In conclusion, we have identified a novel class of promising inhibitors with potent activity against virulent neurotropic alphaviruses.
doi:10.1086/597275
PMCID: PMC2788236  PMID: 19239364
24.  Expression of Putative Immune Response Genes during Early Ontogeny in the Coral Acropora millepora 
PLoS ONE  2012;7(7):e39099.
Background
Corals, like many other marine invertebrates, lack a mature allorecognition system in early life history stages. Indeed, in early ontogeny, when corals acquire and establish associations with various surface microbiota and dinoflagellate endosymbionts, they do not efficiently distinguish between closely and distantly related individuals from the same population. However, very little is known about the molecular components that underpin allorecognition and immunity responses or how they change through early ontogeny in corals.
Methodology/Principal Findings
Patterns in the expression of four putative immune response genes (apextrin, complement C3, and two CELIII type lectin genes) were examined in juvenile colonies of Acropora millepora throughout a six-month post-settlement period using quantitative real-time PCR (qPCR). Expression of a CELIII type lectin gene peaked in the fourth month for most of the coral juveniles sampled and was significantly higher at this time than at any other sampling time during the six months following settlement. The timing of this increase in expression levels of putative immune response genes may be linked to allorecognition maturation which occurs around this time in A.millepora. Alternatively, the increase may represent a response to immune challenges, such as would be involved in the recognition of symbionts (such as Symbiodinium spp. or bacteria) during winnowing processes as symbioses are fine-tuned.
Conclusions/Significance
Our data, although preliminary, are consistent with the hypothesis that lectins may play an important role in the maturation of allorecognition responses in corals. The co-expression of lectins with apextrin during development of coral juveniles also raises the possibility that these proteins, which are components of innate immunity in other invertebrates, may influence the innate immune systems of corals through a common pathway or system. However, further studies investigating the expression of these genes in alloimmune-challenged corals are needed to further clarify emerging evidence of a complex innate immunity system in corals.
doi:10.1371/journal.pone.0039099
PMCID: PMC3391189  PMID: 22792163
25.  Major Cellular and Physiological Impacts of Ocean Acidification on a Reef Building Coral 
PLoS ONE  2012;7(4):e34659.
As atmospheric levels of CO2 increase, reef-building corals are under greater stress from both increased sea surface temperatures and declining sea water pH. To date, most studies have focused on either coral bleaching due to warming oceans or declining calcification due to decreasing oceanic carbonate ion concentrations. Here, through the use of physiology measurements and cDNA microarrays, we show that changes in pH and ocean chemistry consistent with two scenarios put forward by the Intergovernmental Panel on Climate Change (IPCC) drive major changes in gene expression, respiration, photosynthesis and symbiosis of the coral, Acropora millepora, before affects on biomineralisation are apparent at the phenotype level. Under high CO2 conditions corals at the phenotype level lost over half their Symbiodinium populations, and had a decrease in both photosynthesis and respiration. Changes in gene expression were consistent with metabolic suppression, an increase in oxidative stress, apoptosis and symbiont loss. Other expression patterns demonstrate upregulation of membrane transporters, as well as the regulation of genes involved in membrane cytoskeletal interactions and cytoskeletal remodeling. These widespread changes in gene expression emphasize the need to expand future studies of ocean acidification to include a wider spectrum of cellular processes, many of which may occur before impacts on calcification.
doi:10.1371/journal.pone.0034659
PMCID: PMC3324498  PMID: 22509341

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