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novel genetic and molecular tools for the investigation and control of dengue virus transmission by mosquitoes.aedes aegypti is the principal vector of dengue virus (denv) throughout the tropical world. this anthropophilic mosquito species needs to be persistently infected with denv before it can transmit the virus through its saliva to a new vertebrate host. in the mosquito, denv is confronted with several innate immune pathways, among which rna interference is considered the most important. the ae. aegypti genome project opened the doors for advanced molecular studies on pathogen-vector interactions in ...024693489
beth levine receives the 2014 asci/stanley j. korsmeyer award. 201424691475
specific retrograde transduction of spinal motor neurons using lentiviral vectors targeted to presynaptic nmj receptors.to understand how receptors are involved in neuronal trafficking and to be able to utilize them for specific targeting via the peripheral route would be of great benefit. here, we describe the generation of novel lentiviral vectors with tropism to motor neurons that were made by coexpressing onto the lentiviral surface a fusogenic glycoprotein (mutated sindbis g) and an antibody against a cell-surface receptor (thy1.1, p75(ntr), or coxsackievirus and adenovirus receptor) on the presynaptic termi ...201424670531
immunologic treatments for precancerous lesions and uterine cervical cancer.development of hpv-associated cancers not only depends on efficient negative regulation of cell cycle control that supports the accumulation of genetic damage, but also relies on immune evasion that enable the virus to go undetected for long periods of time. in this way, hpv-related tumors usually present mhc class i down-regulation, impaired antigen-processing ability, avoidance of t-cell mediated killing, increased immunosuppression due to treg infiltration and secrete immunosuppressive cytoki ...201424667138
autophagy in plasma cell pathophysiology.plasma cells (pcs) are the effectors responsible for antibody (ab)-mediated immunity. they differentiate from b lymphocytes through a complete remodeling of their original structure and function. stress is a constitutive element of pc differentiation. macroautophagy, conventionally referred to as autophagy, is a conserved lysosomal recycling strategy that integrates cellular metabolism and enables adaptation to stress. in metazoa, autophagy plays diverse roles in cell differentiation. recently, ...201424659989
the role of hur in the post-transcriptional regulation of interleukin-3 in t cells.human interleukin-3 (il-3) is a lymphokine member of a class of transiently expressed mrnas harboring adenosine/uridine-rich elements (are) in their 3' untranslated regions (3'-utrs). the regulatory effects of ares are often mediated by specific are-binding proteins (are-bps). in this report, we show that the human il-3 3'-utr plays a post-transcriptional regulation role in two human transformed cell lines. more specifically, we demonstrate that the hil-3 3'-utr represses the translation of a lu ...201424658545
massive infection of seabird ticks with bacterial species related to coxiella burnetii.seabird ticks are known reservoirs of bacterial pathogens of medical importance; however, ticks parasitizing tropical seabirds have received less attention than their counterparts from temperate and subpolar regions. recently, rickettsia africae was described to infect seabird ticks of the western indian ocean and new caledonia, constituting the only available data on bacterial pathogens associated with tropical seabird tick species. here, we combined a pyrosequencing-based approach with a class ...201424657860
protein interferon-stimulated gene 15 conjugation delays but does not overcome coronavirus proliferation in a model of fulminant hepatitis.coronaviruses express a deubiquitinating protein, the papain-like protease-2 (plp2), that removes both ubiquitin and the ubiquitin-like interferon (ifn)-stimulated gene 15 (isg15) protein from target proteins. isg15 has antiviral activity against a number of viruses; therefore, we examined the effect of isg15 conjugation (isgylation) in a model of acute viral hepatitis induced by the murine hepatitis virus strain 3 (mhv-3) coronavirus. mice deficient in the isg15 deconjugating enzyme, ubiquitin- ...201424648452
consequences of in vitro host shift for st. louis encephalitis virus.understanding the potential for host range shifts and expansions of rna viruses is critical to predicting the evolutionary and epidemiological paths of these pathogens. as arthropod-borne viruses (arboviruses) experience frequent spillover from their amplification cycles and are generalists by nature, they are likely to experience a relatively high frequency of success in a range of host environments. despite this, the potential for host expansion, the genetic correlates of adaptation to novel e ...201424643879
quantitative phosphoproteomics unravels biased phosphorylation of serotonin 2a receptor at ser280 by hallucinogenic versus nonhallucinogenic agonists.the serotonin 5-ht(2a) receptor is a primary target of psychedelic hallucinogens such as lysergic acid diethylamine, mescaline, and psilocybin, which reproduce some of the core symptoms of schizophrenia. an incompletely resolved paradox is that only some 5-ht(2a) receptor agonists exhibit hallucinogenic activity, whereas structurally related agonists with comparable affinity and activity lack such a psychoactive activity. using a strategy combining stable isotope labeling by amino acids in cell ...201424637012
hepatitis c virus, mitochondria and auto/mitophagy: exploiting a host defense mechanism.hepatitis c virus (hcv) is the major reason for liver transplantation and the main cause of liver-related morbidity and mortality in a great number of countries. as for the other viruses, this pathogen interferes in more than one process and in more than one way with host cell biology. a mounting body of evidence points, in particular, toward the drastic alterations of mitochondrial physiology and functions that virus is able to induce, albeit the mechanisms have partly remained elusive. role of ...024627598
vectorology and factor delivery in induced pluripotent stem cell reprogramming.induced pluripotent stem cell (ipsc) reprogramming requires sustained expression of multiple reprogramming factors for a limited period of time (10-30 days). conventional ipsc reprogramming was achieved using lentiviral or simple retroviral vectors. retroviral reprogramming has flaws of insertional mutagenesis, uncontrolled silencing, residual expression and re-activation of transgenes, and immunogenicity. to overcome these issues, various technologies were explored, including adenoviral vectors ...201424625220
the c-terminal repeat domains of nsp3 from the old world alphaviruses bind directly to g3bp.the old world alphaviruses block stress granule assembly by sequestration of rasgap sh3-domain binding protein (g3bp). here, we show that the proline-rich sequences in the hypervariable domain of nonstructural protein 3 (nsp3) of both semliki forest virus and chikungunya virus were dispensable for binding to g3bp. nsp3 variants with or without this domain colocalized with g3bp. furthermore, we show that the c-terminal repeat motifs of nsp3 were sufficient for g3bp binding.201424623412
enhancement of antitumor immunity using a dna-based replicon vaccine derived from semliki forest virus.a dna-based replicon vaccine derived from semliki forest virus, psvk-shfcg-gm/b7.1 (fig. 1a) was designed for tumor immunotherapy as previously constructed. the expression of the fusion tumor antigen (survivin and hcgβ-ctp37) and adjuvant molecular protein (granulocyte-macrophage colony-stimulating factor/ gm-csf/b7.1) genes was confirmed by immunofluorescence assay in vitro, and immunohistochemistry assay in vivo. in this paper, the immunological effect of this vaccine was determined using immu ...201424608380
inhibition of dengue virus replication by a class of small-molecule compounds that antagonize dopamine receptor d4 and downstream mitogen-activated protein kinase signaling.dengue viruses (denv) are endemic pathogens of tropical and subtropical regions that cause significant morbidity and mortality worldwide. to date, no vaccines or antiviral therapeutics have been approved for combating denv-associated disease. in this paper, we describe a class of tricyclic small-molecule compounds-dihydrodibenzothiepines (dhbts), identified through high-throughput screening-with potent inhibitory activity against denv serotype 2. ski-417616, a highly active representative of thi ...201424599995
two alternative ways of start site selection in human norovirus reinitiation of translation.the calicivirus minor capsid protein vp2 is expressed via termination/reinitiation. this process depends on an upstream sequence element denoted termination upstream ribosomal binding site (turbs). we have shown for feline calicivirus and rabbit hemorrhagic disease virus that the turbs contains three sequence motifs essential for reinitiation. motif 1 is conserved among caliciviruses and is complementary to a sequence in the 18 s rrna leading to the model that hybridization between motif 1 and 1 ...201424599949
identification of the catalytic triad of family s46 exopeptidases, closely related to clan pa endopeptidases.the exopeptidases of family s46 are exceptional, as the closest homologs of these enzymes are the endopeptidases of clan pa. the three-dimensional structure of s46 enzymes is unknown and only one of the catalytic residues, the serine, has been identified. the catalytic histidine and aspartate residues are not experimentally identified. here we present phylogenetic and experimental data that identify all residues of the catalytic triad of s46 peptidase, dipeptidyl aminopeptidase bii (dap bii) fro ...201424598890
critical role of interferon-α constitutively produced in human hepatocytes in response to rna virus infection.several viruses are known to infect human liver and cause the hepatitis, but the interferon (ifn) response, a first-line defense against viral infection, of virus-infected hepatocytes is not clearly defined yet. we investigated innate immune system against rna viral infection in immortalized human hepatocytes (hus-e/2 cells), as the cells showed similar early innate immune responses to primary human hepatocytes (phh). the low-level constitutive expression of ifn-α1 gene, but not ifn-β and ifn-λ, ...201424587086
the role of ikkβ in venezuelan equine encephalitis virus infection.venezuelan equine encephalitis virus (veev) belongs to the genus alphavirus, family togaviridae. veev infection is characterized by extensive inflammation and studies from other laboratories implicated an involvement of the nf-κb cascade in the in vivo pathology. initial studies indicated that at early time points of veev infection, the nf-κb complex was activated in cells infected with the tc-83 strain of veev. one upstream kinase that contributes to the phosphorylation of p65 is the ikkβ compo ...201424586253
more novel hantaviruses and diversifying reservoir hosts--time for development of reservoir-derived cell culture models?due to novel, improved and high-throughput detection methods, there is a plethora of newly identified viruses within the genus hantavirus. furthermore, reservoir host species are increasingly recognized besides representatives of the order rodentia, now including members of the mammalian orders soricomorpha/eulipotyphla and chiroptera. despite the great interest created by emerging zoonotic viruses, there is still a gross lack of in vitro models, which reflect the exclusive host adaptation of mo ...201424576845
isolation of sindbis virus from a hooded crow in germany.sindbis virus (sinv) is an arbovirus that causes clinical symptoms, including arthritis, rash, and fever during acute human infections. in europe, sinv outbreaks are largely restricted to northern europe. intrigued by the isolation of sinv from mosquitoes in southwestern germany in 2009, we initiated a passive arbovirus-monitoring program in birds and analyzed a total of 685 samples. by this approach, we were able to detect a sinv in a hooded crow in germany for the first time. it was possible t ...201424575742
dynamic survey of mitochondria by ubiquitin.ubiquitin is a post-translational modifier with proteolytic and non-proteolytic roles in many biological processes. at mitochondria, it performs regulatory homeostatic functions and contributes to mitochondrial quality control. ubiquitin is essential for mitochondrial fusion, regulates mitochondria-er contacts, and participates in maternal mtdna inheritance. under stress, mitochondrial dysfunction induces ubiquitin-dependent responses that involve mitochondrial proteome remodeling and culminate ...201424569520
tissue-specific attenuation of oncolytic sindbis virus without compromised genetic stability.wild-type sindbis virus (sv) shows promise as an oncolytic agent, although potential off-target replication is a safety concern. to remove possible pathology reflecting virus replication in liver, muscle, and/or hematopoietic cells, microrna (mir)-response elements (mres) to liver-specific mir122, muscle-specific mir133a and mir206, or hematopoietic-specific mir142-3p were inserted into the sindbis viral genome. we compared the effectiveness of mres in two distinct genomic locations and found be ...201424568203
chikungunya fever in canada: fever and polyarthritis in a returned traveller. 201424566646
virus adsorption of water-stable quaternized chitosan nanofibers.the burden of unsafe drinking water is responsible for millions of deaths each year. to relieve this burden, we are in search of an inexpensive material that can adsorb pathogens from drinking water. in this pursuit, we have studied the natural carbohydrate, chitosan. to impart virus removal features, chitosan has been functionalized with a quaternary amine to form quaternized chitosan n-[(2-hydroxyl-3-trimethylammonium) propyl] chitosan (htcc). htcc can be electrospun into nanofibers with the n ...201424561959
interferon-stimulated genes: a complex web of host defenses.interferon-stimulated gene (isg) products take on a number of diverse roles. collectively, they are highly effective at resisting and controlling pathogens. in this review, we begin by introducing interferon (ifn) and the jak-stat signaling pathway to highlight features that impact isg production. next, we describe ways in which isgs both enhance innate pathogen-sensing capabilities and negatively regulate signaling through the jak-stat pathway. several isgs that directly inhibit virus infection ...201424555472
autonomous parvoviruses neither stimulate nor are inhibited by the type i interferon response in human normal or cancer cells.members of the genus parvovirus are small, nonenveloped single-stranded dna viruses that are nonpathogenic in humans but have potential utility as cancer therapeutics. because the innate immune response to parvoviruses has received relatively little attention, we compared the response to parvoviruses to that of several other types of viruses in human cells. in normal human glia, fibroblasts, or melanocytes, vesicular stomatitis virus evoked robust beta interferon (ifn-β) responses. cytomegalovir ...201424554651
the yellow fever 17d virus as a platform for new live attenuated vaccines.the live-attenuated yellow fever 17d virus is one of the most outstanding human vaccines ever developed. it induces efficacious immune responses at a low production cost with a well-established manufacture process. these advantages make the yf17d virus attractive as a vector for the development of new vaccines. at the beginning of vector development studies, yf17d was genetically manipulated to express other flavivirus prm and e proteins, components of the viral envelope. while these 17d recombi ...201424553128
distinctive effects of the cellular inhibitor of apoptosis protein c-iap2 through stabilization by xiap in glioblastoma multiforme cells.inhibitor of apoptosis proteins (iaps) are extensively involved in nfκb signaling pathways. regulation of c-iap2 turnover by other proteins was investigated in glioblastoma multiforme (gbm) cells in the present study. when overexpressed, x-linked iap (xiap) enhanced expression of ectopic c-iap2, but not c-iap1, and endogenous c-iap2 levels were reduced once xiap expression was silenced. tnfα stimulation substantially increased c-iap2 expression, and this upregulation was impaired by suppression ...201424552816
genome-wide rnai screen identifies broadly-acting host factors that inhibit arbovirus infection.vector-borne viruses are an important class of emerging and re-emerging pathogens; thus, an improved understanding of the cellular factors that modulate infection in their respective vertebrate and insect hosts may aid control efforts. in particular, cell-intrinsic antiviral pathways restrict vector-borne viruses including the type i interferon response in vertebrates and the rna interference (rnai) pathway in insects. however, it is likely that additional cell-intrinsic mechanisms exist to limi ...201424550726
microrna-like viral small rna from dengue virus 2 autoregulates its replication in mosquito cells.micrornas (mirnas) are small regulatory rnas that play significant roles in most cellular processes. in the seemingly endless arms race between hosts and pathogens, viruses also encode mirnas that facilitate successful infection. in search of functional mirnas or viral small rnas (vsrnas) encoded by dengue virus (denv), deep sequencing data of virus-infected aedes aegypti mosquitoes were used. from six vsrnas, with candidate stem-loop structures in the 5' and 3' untranslated regions of the viral ...201424550303
efficient transfection of dissociated mouse chromaffin cells using small-volume electroporation.we have developed an improved procedure for isolating and transfecting a chromaffin cell-enriched population of primary cells from adult mouse adrenal glands. significantly, the parameters of a novel electroporation transfection technique were optimized to achieve an average transfection efficiency of 45 % on the small number of cells derived from the mouse glands. such transfection efficiency was previously unachievable with the electroporation protocols conventionally used with bovine chromaff ...201424549789
prevention of contamination by xenotropic murine leukemia virus-related virus: susceptibility to alcohol-based disinfectants and environmental stability.xenotropic murine leukemia virus-related virus (xmrv) represents a novel γ-retrovirus that is capable of infecting human cells and has been classified as a biosafety level 2 (bsl-2) organism. hence, xmrv represents a potential risk for personnel in laboratories worldwide. here, we measured the stability of xmrv and its susceptibility to alcohol-based disinfectants. to this end, we exposed an infectious xmrv reporter virus encoding a secretable luciferase to different temperatures, ph values, and ...201424532072
relating structure to evolution in class ii viral membrane fusion proteins.enveloped viruses must fuse their lipid membrane to a cellular membrane to deliver the viral genome into the cytoplasm for replication. viral envelope proteins catalyze this critical membrane fusion event. they fall into at least three distinct structural classes. class ii fusion proteins have a conserved three-domain architecture and are found in many important viral pathogens. until 2013, class ii proteins had only been found in flaviviruses and alphaviruses. however, in 2013 a class ii fusion ...201424525225
hsp90 chaperone inhibitor 17-aag attenuates aβ-induced synaptic toxicity and memory impairment.the excessive accumulation of soluble amyloid peptides (aβ) plays a crucial role in the pathogenesis of alzheimer's disease (ad), particularly in synaptic dysfunction. the role of the two major chaperone proteins, hsp70 and hsp90, in clearing misfolded protein aggregates has been established. despite their abundant presence in synapses, the role of these chaperones in synapses remains elusive. here, we report that hsp90 inhibition by 17-aag elicited not only a heat shock-like response but also u ...024523537
human-like antibodies neutralizing western equine encephalitis virus.this study describes the development of the first neutralizing antibodies against western equine encephalitis virus (weev), a member of the genus alphavirus. weev is transmitted by mosquitoes and can spread to the human central nervous system, causing symptoms ranging from mild febrile reactions to life-threatening encephalitis. weev has been classified as a biological warfare agent by the us centers for disease control and prevention. no anti-weev drugs are currently commercially available. neu ...201424518197
a gammaherpesvirus bcl-2 ortholog blocks b cell receptor-mediated apoptosis and promotes the survival of developing b cells in vivo.gammaherpesviruses such as epstein-barr virus (ebv) and kaposi's sarcoma-associated herpesvirus (kshv, hhv-8) establish lifelong latency in their hosts and are associated with the development of several types of malignancies, including a subset of b cell lymphomas. these viruses are thought to co-opt the process of b cell differentiation to latently infect a fraction of circulating memory b cells, resulting in the establishment of a stable latency setpoint. however, little is known about how thi ...201424516386
the major cellular sterol regulatory pathway is required for andes virus infection.the bunyaviridae comprise a large family of rna viruses with worldwide distribution and includes the pathogenic new world hantavirus, andes virus (andv). host factors needed for hantavirus entry remain largely enigmatic and therapeutics are unavailable. to identify cellular requirements for andv infection, we performed two parallel genetic screens. analysis of a large library of insertionally mutagenized human haploid cells and a sirna genomic screen converged on components (srebp-2, scap, s1p a ...201424516383
mers-cov papain-like protease has deisgylating and deubiquitinating activities.coronaviruses encode papain-like proteases (plpro) that are often multifunctional enzymes with protease activity to process the viral replicase polyprotein and deubiquitinating (dub)/deisgylating activity, which is hypothesized to modify the innate immune response to infection. here, we investigate the predicted dub activity of the plpro domain of the recently described middle east respiratory syndrome coronavirus (mers-cov). we found that expression of mers-cov plpro reduces the levels of ubiqu ...201324503068
generation of infectious virus particles from inducible transgenic genomes.arboviruses like dengue virus, yellow fever virus, and west nile virus are enveloped particles spread by mosquitoes, infecting millions of humans per year, with neither effective vaccines, nor specific antiviral therapies [1,2]. previous studies of infection and virus replication utilize either purified virus particles or deficient genomes that do not complete the viral life cycle [1,2]. here we describe transgenic drosophila strains expressing trans-complementing genomes (referred to as 'replic ...024502780
dynamics of the emergence and establishment of a newly dominant genotype of japanese encephalitis virus throughout asia.in recent years, genotype i (gi) of japanese encephalitis virus (jev) has displaced genotype iii (giii) as the dominant virus genotype throughout asia. in this study, the largest collection of giii and gi envelope gene-derived viral sequences assembled to date was used to reconstruct the spatiotemporal chronology of genotype displacement throughout asia and to determine the evolutionary and epidemiological dynamics underlying this significant event. gi consists of two clades, gi-a and gi-b, with ...201424501419
the xxiiird phage/virus assembly meeting.the xxiiird phage/virus assembly (pva) meeting returned to its birthplace in lake arrowhead, ca on september 8-13, 2013 (fig. 1). the original meeting occurred in 1968, organized by bob edgar (caltech, pasadena, ca usa), fred eiserling (university of california, los angeles, los angeles, ca usa) and bill wood (caltech, pasadena, ca usa). the organizers of the 2013 meeting were bill gelbart (university of california, los angeles, los angeles, ca usa) and jack johnson (scripps research institute, ...024498537
estimating the burden of japanese encephalitis virus and other encephalitides in countries of the mekong region.diverse aetiologies of viral and bacterial encephalitis are widely recognized as significant yet neglected public health issues in the mekong region. a robust analysis of the corresponding health burden is lacking. we retrieved 75 articles on encephalitis in the region published in english or in french from 1965 through 2011. review of available data demonstrated that they are sparse and often derived from hospital-based studies with significant recruitment bias. almost half (35 of 75) of articl ...201424498443
pharmacological rescue of ras signaling, glua1-dependent synaptic plasticity, and learning deficits in a fragile x model.fragile x syndrome, caused by the loss of fmr1 gene function, is the most common form of inherited mental retardation, with no effective treatment. using a tractable animal model, we investigated mechanisms of action of a few fda-approved psychoactive drugs that modestly benefit the cognitive performance in fragile x patients. here we report that compounds activating serotonin (5ht) subtype 2b receptors (5ht2b-rs) or dopamine (da) subtype 1-like receptors (d1-rs) and/or those inhibiting 5ht2a-rs ...024493647
surveillance for arboviral zoonoses in new zealand birds.given the significant burden that emerging infectious diseases place on global economies and public health, the monitoring and mitigation of, and early response to, potential infectious diseases are of the highest priority. the objective of this study was to survey for known and other potential arboviral zoonoses in multiple bird species at four locations in new zealand.201324478919
role of phosphatidylserine receptors in enveloped virus infection.we recently demonstrated that a soluble protein, gas6, can facilitate viral entry by bridging viral envelope phosphatidylserine to axl, a receptor tyrosine kinase expressed on target cells. the interaction between phosphatidylserine, gas6, and axl was originally shown to be a molecular mechanism through which phagocytes recognize phosphatidylserine exposed on dead cells. since our initial report, several groups have confirmed that axl/gas6, as well as other phosphatidylserine receptors, facilita ...201424478428
roles of the putative integrin-binding motif of the human metapneumovirus fusion (f) protein in cell-cell fusion, viral infectivity, and pathogenesis.human metapneumovirus (hmpv) is a relatively recently identified paramyxovirus that causes acute upper and lower respiratory tract infection. entry of hmpv is unusual among the paramyxoviruses, in that fusion is accomplished by the fusion (f) protein without the attachment glycoprotein (g protein). it has been suggested that hmpv f protein utilizes integrin αvβ1 as a cellular receptor. consistent with this, the f proteins of all known hmpv strains possess an integrin-binding motif ((329)rgd(331) ...201424478423
chemistry and biological activities of flavonoids: an overview.there has been increasing interest in the research on flavonoids from plant sources because of their versatile health benefits reported in various epidemiological studies. since flavonoids are directly associated with human dietary ingredients and health, there is need to evaluate structure and function relationship. the bioavailability, metabolism, and biological activity of flavonoids depend upon the configuration, total number of hydroxyl groups, and substitution of functional groups about th ...201324470791
structural plasticity of the semliki forest virus glycome upon interspecies transmission.cross-species viral transmission subjects parent and progeny alphaviruses to differential post-translational processing of viral envelope glycoproteins. alphavirus biogenesis has been extensively studied, and the semliki forest virus e1 and e2 glycoproteins have been shown to exhibit differing degrees of processing of n-linked glycans. however the composition of these glycans, including that arising from different host cells, has not been determined. here we determined the chemical composition o ...201424467287
effect of mixed infections of sindbis and la crosse viruses on replication of each virus in vitro.mixed viral infections are ubiquitous in natural populations and may have significant but unpredictable biological and epidemiological consequences. these infections may be acquired simultaneously (coinfection) or result from two single infections at different time intervals (superinfection). this study investigated the effect of mixed infections of two mosquito-borne viruses, sindbis (sinv) and la crosse (lacv), on the replication of each virus in vertebrate baby hamster kidney (bhk) and invert ...201424463260
the alternate triad motif of the poly(adp-ribose) polymerase-like domain of the human zinc finger antiviral protein is essential for its antiviral activity.the human zinc finger antiviral protein (hzap) gene is spliced to yield a short (hzap-s) and a long (hzap-l) isoform. the long isoform possesses a poly(adp-ribose) polymerase (parp)-like domain in its c-terminus predicted to be inactive due to alterations in its triad motif compared with bona fide parps. using sindbis virus as prototype member of alphaviruses we confirmed that hzap-l is a more potent inhibitor of alphaviruses than hzap-s. specific small interfering rna knockdown of hzap-l but no ...201424457973
alphavirus mutator variants present host-specific defects and attenuation in mammalian and insect models.arboviruses cycle through both vertebrates and invertebrates, which requires them to adapt to disparate hosts while maintaining genetic integrity during genome replication. to study the genetic mechanisms and determinants of these processes, we use chikungunya virus (chikv), a re-emerging human pathogen transmitted by the aedes mosquito. we previously isolated a high fidelity (or antimutator) polymerase variant, c483y, which had decreased fitness in both mammalian and mosquito hosts, suggesting ...201424453971
from scourge to cure: tumour-selective viral pathogenesis as a new strategy against cancer.tumour mutations corrupt cellular pathways, and accumulate to disrupt, dysregulate, and ultimately avoid mechanisms of cellular control. yet the very changes that tumour cells undergo to secure their own growth success also render them susceptible to viral infection. enhanced availability of surface receptors, disruption of antiviral sensing, elevated metabolic activity, disengagement of cell cycle controls, hyperactivation of mitogenic pathways, and apoptotic avoidance all render the malignant ...201424453963
inefficient type i interferon-mediated antiviral protection of primary mouse neurons is associated with the lack of apolipoprotein l9 expression.we examined the antiviral response promoted by type i interferons (ifn) in primary mouse neurons. ifn treatment of neuron cultures strongly upregulated the transcription of ifn-stimulated genes but conferred a surprisingly low resistance to infection by neurotropic viruses such as theiler's murine encephalomyelitis virus (tmev) or vesicular stomatitis virus (vsv). response of primary mouse neurons to ifn treatment was heterogeneous, as many neurons failed to express the typical ifn response mark ...201424453359
targeting γ-herpesvirus 68 bcl-2-mediated down-regulation of autophagy.γ-herpesviruses (γhvs) are common human pathogens that encode homologs of the anti-apoptotic cellular bcl-2 proteins, which are critical to viral reactivation and oncogenic transformation. the murine γhv68 provides a tractable in vivo model for understanding general features of these important human pathogens. bcl-xl, a cellular bcl-2 homolog, and the murine γhv68 bcl-2 homolog, m11, both bind to a bh3 domain within the key autophagy effector beclin 1 with comparable affinities, resulting in the ...201424443581
gene delivery in malignant b cells using the combination of lentiviruses conjugated to anti-transferrin receptor antibodies and an immunoglobulin promoter.we previously developed an antibody-avidin fusion protein (ch128.1av) specific for the human transferrin receptor 1 (tfr1; cd71) to be used as a delivery vector for cancer therapy and showed that ch128.1av delivers the biotinylated plant toxin saporin-6 into malignant b cells. however, as a result of widespread expression of tfr1, delivery of the toxin to normal cells is a concern. therefore, we explored the potential of a dual targeted lentiviral-mediated gene therapy strategy to restrict gene ...024436117
approaches to the treatment of disease induced by chikungunya virus.chikungunya virus, a re-emerging mosquito-borne alphavirus, causes fever, rash and persistent arthralgia/arthritis in humans. severe outbreaks have occurred resulting in infections of millions of people in southeast asia and africa. currently there are no antiviral drugs or vaccines for prevention and treatment of chikungunya infections. herein we report the current status of research on antiviral drugs and vaccines for chikungunya virus infections.024434329
attenuating mutations in nsp1 reveal tissue-specific mechanisms for control of ross river virus infection.ross river virus (rrv) is one of a group of mosquito-transmitted alphaviruses that cause debilitating, and often chronic, musculoskeletal disease in humans. previously, we reported that replacement of the nonstructural protein 1 (nsp1) gene of the mouse-virulent rrv strain t48 with that from the mouse-avirulent strain dc5692 generated a virus that was attenuated in a mouse model of disease. here we find that the six nsp1 nonsynonymous nucleotide differences between strains t48 and dc5692 are det ...201424429363
zoonotic encephalitides caused by arboviruses: transmission and epidemiology of alphaviruses and flaviviruses.in this review, we mainly focus on zoonotic encephalitides caused by arthropod-borne viruses (arboviruses) of the families flaviviridae (genus flavivirus) and togaviridae (genus alphavirus) that are important in both humans and domestic animals. specifically, we will focus on alphaviruses (eastern equine encephalitis virus, western equine encephalitis virus, venezuelan equine encephalitis virus) and flaviviruses (japanese encephalitis virus and west nile virus). most of these viruses were origin ...201324427764
differential regulation of gabab receptor trafficking by different modes of n-methyl-d-aspartate (nmda) receptor signaling.inhibitory gabab receptors (gababrs) can down-regulate most excitatory synapses in the cns by reducing postsynaptic excitability. functional gababrs are heterodimers of gabab1 and gabab2 subunits and here we show that the trafficking and surface expression of gababrs is differentially regulated by synaptic or pathophysiological activation of nmda receptors (nmdars). activation of synaptic nmdars using a chemltp protocol increases gababr recycling and surface expression. in contrast, excitotoxic ...201424425870
design of a novel integration-deficient lentivector technology that incorporates genetic and posttranslational elements to target human dendritic cells.as sentinels of the immune system, dendritic cells (dcs) play an essential role in regulating cellular immune responses. one of the main challenges of developing dc-targeted therapies includes the delivery of antigen to dcs in order to promote the activation of antigen-specific effector cd8 t cells. with the goal of creating antigen-directed immunotherapeutics that can be safely administered directly to patients, immune design has developed a platform of novel integration-deficient lentiviral ve ...201324419083
salmonid alphavirus replication in mosquito cells: towards a novel vaccine production system.salmonid alphavirus (sav) causes pancreas disease and sleeping disease in atlantic salmon (salmo salar) and rainbow trout (oncorhynchus mykiss) and confers a major burden to the aquaculture industry. a commercial inactivated whole virus vaccine propagated in a salmon cell line at low temperature provides effective protection against sav infections. alphaviruses (family togaviridae) are generally transmitted between vertebrate hosts via blood-sucking arthropod vectors, typically mosquitoes. sav i ...201424418177
the role of viral genomic rna and nucleocapsid protein in the autophagic clearance of hantavirus glycoprotein gn.hantaviruses have tri-segmented negative sense rna genome. the viral m-segment rna encodes a glycoprotein precursor (gpc), which is cleaved into two glycoprotein molecules gn and gc that form spikes on the viral envelope. we previously reported that gn is degraded shortly after synthesis by the host autophagy machinery. however, gn being an important integral component of the virion, must escape degradation during the packaging and assembly stage of virus replication cycle. the mechanism regulat ...201424412713
the role of hts in drug discovery at the university of michigan.high throughput screening (hts) is an integral part of a highly collaborative approach to drug discovery at the university of michigan. the hts lab is one of four core centers that provide services to identify, produce, screen and follow-up on biomedical targets for faculty. key features of this system are: protein cloning and purification, protein crystallography, small molecule and sirna hts, medicinal chemistry and pharmacokinetics. therapeutic areas that have been targeted include anti-bacte ...024409957
functional cross-talk between distant domains of chikungunya virus non-structural protein 2 is decisive for its rna-modulating activity.chikungunya virus (chikv) non-structural protein 2 (nsp2) is a multifunctional protein that is considered a master regulator of the viral life cycle and a main viral factor responsible for cytopathic effects and subversion of antiviral defense. the c-terminal part of nsp2 possesses protease activity, whereas the n-terminal part exhibits ntpase and rna triphosphatase activity and is proposed to have helicase activity. bioinformatics analysis classified chikv nsp2 into helicase superfamily 1. howe ...201424407286
human pathogenic bacteria, fungi, and viruses in drosophila: disease modeling, lessons, and shortcomings.drosophila has been the invertebrate model organism of choice for the study of innate immune responses during the past few decades. many drosophila-microbe interaction studies have helped to define innate immunity pathways, and significant effort has been made lately to decipher mechanisms of microbial pathogenesis. here we catalog 68 bacterial, fungal, and viral species studied in flies, 43 of which are relevant to human health. we discuss studies of human pathogens in flies revealing not only ...201424398387
phosphorylation of initiation factor eif2 in response to stress conditions is mediated by acidic ribosomal p1/p2 proteins in saccharomyces cerevisiae.eukaryotic cells contain an unusually large cytoplasmic pool of p1/p2 phosphoproteins, which form the highly flexible 60s subunit stalk that is required to interact with and activate soluble translation factors. in cells, cytoplasmic p1/p2 proteins are exchanged for ribosome-bound proteins in a process that can modulate ribosome function and translation. here, we analysed different s. cerevisiae stalk mutants grown under stress conditions that result in eif2α phosphorylation. these mutants eithe ...201324391917
small rna analysis in sindbis virus infected human hek293 cells.in contrast to the defence mechanism of rna interference (rnai) in plants and invertebrates, its role in the innate response to virus infection of mammals is a matter of debate. since rnai has a well-established role in controlling infection of the alphavirus sindbis virus (sinv) in insects, we have used this virus to investigate the role of rnai in sinv infection of human cells.201324391886
sorting through the roles of beclin 1 in microglia and neurodegeneration.beclin 1 has a well-established role in regulating autophagy, a cellular degradation pathway. although the yeast ortholog of beclin 1 (atg6/vps30) was discovered to also regulate vacuolar protein sorting nearly 30 years ago, the varied functions of beclin 1 in mammalian cells are only beginning to be sorted out. we recently described a role for beclin 1 in regulating recycling of phagocytic receptors in microglia, a function analogous to that of its yeast ortholog. microglia lacking beclin 1 hav ...201424385262
antimicrobial and cytotoxic activities of turbinariaconoides (j.agardh) kuetz.brown alga, turbinariaconoideswas successively extracted with n-hexane, cyclohexane, methanol and ethanol:water (1:1). the extracts were evaluated for antibacterial and antifungal activities by disc diffusion method. minimal inhibitory concentration was determined for active extracts by broth dilution method. the antiviral activity and cytotoxicity of the extracts were tested in human embryonic lung (hel) cells (herpes simplex virus-1, herpes simplex virus-2, vaccinia virus, vesicular stomatitis ...201024381606
identical strength of the t cell responses against e2, nsp1 and capsid chikv proteins in recovered and chronic patients after the epidemics of 2005-2006 in la reunion island.to characterize the immunity developed by patients infected by chikungunya virus (chikv), we studied the intensity and specificity of chikv-specific t cells mediated responses in chronic and recovered patients at 12 to 24 months post-infection. t cells were challenged in vitro against chikv synthetic peptides covering the length of three viral proteins, capsid, e2 and nsp1 proteins as well as all inactivated virus particles. cytokine production was assessed by elispot and intracellular labeling. ...201324376836
antiviral autophagy restrictsrift valley fever virus infection and is conserved from flies to mammals.autophagy has been implicated as a component of host defense, but the significance of antimicrobial autophagy in vivo and the mechanism by which it is regulated during infection are poorly defined. here we found that antiviral autophagy was conserved in flies and mammals during infection with rift valley fever virus (rvfv), a mosquito-borne virus that causes disease in humans and livestock. in drosophila, toll-7 limited rvfv replication and mortality through activation of autophagy. rvfv infecti ...201324374193
effective virus inactivation and removal by steps of biotest pharmaceuticals igiv production process.the virus validation of three steps of biotest pharmaceuticals igiv production process is described here. the steps validated are precipitation and removal of fraction iii of the cold ethanol fractionation process, solvent/detergent treatment and 35 nm virus filtration. virus validation was performed considering combined worst case conditions. by these validated steps sufficient virus inactivation/removal is achieved, resulting in a virus safe product.201224371563
inhibition of translation by ifit family members is determined by their ability to interact selectively with the 5'-terminal regions of cap0-, cap1- and 5'ppp- mrnas.ribosomal recruitment of cellular mrnas depends on binding of eif4f to the mrna's 5'-terminal 'cap'. the minimal 'cap0' consists of n7-methylguanosine linked to the first nucleotide via a 5'-5' triphosphate (ppp) bridge. cap0 is further modified by 2'-o-methylation of the next two riboses, yielding 'cap1' (m7gpppnmn) and 'cap2' (m7gpppnmnm). however, some viral rnas lack 2'-o-methylation, whereas others contain only ppp- at their 5'-end. interferon-induced proteins with tetratricopeptide repeats ...201324371270
a single-amino-acid polymorphism in chikungunya virus e2 glycoprotein influences glycosaminoglycan utilization.chikungunya virus (chikv) is a reemerging arbovirus responsible for outbreaks of infection throughout asia and africa, causing an acute illness characterized by fever, rash, and polyarthralgia. although chikv infects a broad range of host cells, little is known about how chikv binds and gains access to the target cell interior. in this study, we tested whether glycosaminoglycan (gag) binding is required for efficient chikv replication using chikv vaccine strain 181/25 and clinical isolate sl1564 ...201324371059
structural basis for the regulatory function of a complex zinc-binding domain in a replicative arterivirus helicase resembling a nonsense-mediated mrna decay helicase.all positive-stranded rna viruses with genomes>∼7 kb encode helicases, which generally are poorly characterized. the core of the nidovirus superfamily 1 helicase (hel1) is associated with a unique n-terminal zinc-binding domain (zbd) that was previously implicated in helicase regulation, genome replication and subgenomic mrna synthesis. the high-resolution structure of the arterivirus helicase (nsp10), alone and in complex with a polynucleotide substrate, now provides first insights into the str ...201324369429
subgenomic reporter rna system for detection of alphavirus infection in mosquitoes.current methods for detecting real-time alphavirus (family togaviridae) infection in mosquitoes require the use of recombinant viruses engineered to express a visibly detectable reporter protein. these altered viruses expressing fluorescent proteins, usually from a duplicated viral subgenomic reporter, are effective at marking infection but tend to be attenuated due to the modification of the genome. additionally, field strains of viruses cannot be visualized using this approach unless infectiou ...201324367703
genome-wide rnai screen identifies novel host proteins required for alphavirus entry.the enveloped alphaviruses include important and emerging human pathogens such as chikungunya virus and eastern equine encephalitis virus. alphaviruses enter cells by clathrin-mediated endocytosis, and exit by budding from the plasma membrane. while there has been considerable progress in defining the structure and function of the viral proteins, relatively little is known about the host factors involved in alphavirus infection. we used a genome-wide sirna screen to identify host factors that pr ...201324367265
ghrelin triggers the synaptic incorporation of ampa receptors in the hippocampus.ghrelin is a peptide mainly produced by the stomach and released into circulation, affecting energy balance and growth hormone release. these effects are guided largely by the expression of the ghrelin receptor growth hormone secretagogue type 1a (ghs-r1a) in the hypothalamus and pituitary. however, ghs-r1a is expressed in other brain regions, including the hippocampus, where its activation enhances memory retention. herein we explore the molecular mechanism underlying the action of ghrelin on h ...201324367106
type 1 metabotropic glutamate receptors (mglu1) trigger the gating of glud2 delta glutamate receptors.the orphan glud2 receptor belongs to the ionotropic glutamate receptor family but does not bind glutamate. ligand-gated glud2 currents have never been evidenced, and whether glud2 operates as an ion channel has been a long-standing question. here, we show that glud2 gating is triggered by type 1 metabotropic glutamate receptors, both in a heterologous expression system and in purkinje cells. thus, glud2 is not only an adhesion molecule at synapses but also works as a channel. this gating mechani ...201324357660
the role of innate immunity in conditioning mosquito susceptibility to west nile virus.arthropod-borne viruses (arboviruses) represent an emerging threat to human and livestock health globally. in particular, those transmitted by mosquitoes present the greatest challenges to disease control efforts. an understanding of the molecular basis for mosquito innate immunity to arbovirus infection is therefore critical to investigations regarding arbovirus evolution, virus-vector ecology, and mosquito vector competence. in this review, we discuss the current state of understanding regardi ...201324351797
gene therapy strategies for hiv/aids: preclinical modeling in humanized mice.in the absence of an effective vaccine and lack of a complete cure, gene therapy approaches to control hiv infection offer feasible alternatives. due to the chronic nature of infection, a wide window of opportunity exists to gene modify the hiv susceptible cells that continuously arise from the bone marrow source. to evaluate promising gene therapy approaches that employ various anti-hiv therapeutic molecules, an ideal animal model is necessary to generate important efficacy and preclinical data ...201324351796
lentiviral vector-based therapy in head and neck cancer (review).head and neck squamous cell carcinoma (hnscc) is the sixth most common neoplasm worldwide. despite advances in multimodality treatments involving surgery, radiation and chemotherapy, the five-year survival rate has remained at ~50% for the past 35 years. therefore, the early detection of recurrent or persistent disease is extremely important. replication-incompetent hiv-1-based lentiviral vectors have emerged as powerful and safe tools for gene delivery. commonly, hnscc is a locoregional disease ...201324348811
exploring early stages of the chemical unfolding of proteins at the proteome scale.after decades of using urea as denaturant, the kinetic role of this molecule in the unfolding process is still undefined: does urea actively induce protein unfolding or passively stabilize the unfolded state? by analyzing a set of 30 proteins (representative of all native folds) through extensive molecular dynamics simulations in denaturant (using a range of force-fields), we derived robust rules for urea unfolding that are valid at the proteome level. irrespective of the protein fold, presence ...201324348236
cytoplasmic viruses: rage against the (cellular rna decay) machine. 201324339774
interferon-stimulated poly(adp-ribose) polymerases are potent inhibitors of cellular translation and virus replication.the innate immune response is the first line of defense against most viral infections. its activation promotes cell signaling, which reduces virus replication in infected cells and leads to induction of the antiviral state in yet-uninfected cells. this inhibition of virus replication is a result of the activation of a very broad spectrum of specific cellular genes, with each of their products usually making a small but detectable contribution to the overall antiviral state. the lack of a strong, ...201324335297
infection of myofibers contributes to increased pathogenicity during infection with an epidemic strain of chikungunya virus.chikungunya virus (chikv) is an alphavirus transmitted by mosquitoes that is known to cause severe arthritis and myositis in affected patients. the ongoing epidemic began in eastern africa in 2004 and then spread to islands of the indian ocean, india, and southeast asia, ultimately afflicting millions. during this outbreak, more severe disease manifestations, including fatalities, have been documented. the reasons for this change in pathogenesis are multifactorial but likely include mutations th ...201324335291
the influence of viral rna secondary structure on interactions with innate host cell defences.rna viruses infecting vertebrates differ fundamentally in their ability to establish persistent infections with markedly different patterns of transmission, disease mechanisms and evolutionary relationships with their hosts. although interactions with host innate and adaptive responses are complex and persistence mechanisms likely multi-factorial, we previously observed associations between bioinformatically predicted rna secondary formation in genomes of positive-stranded rna viruses with their ...201324335283
genome-wide rnai screen identifies sec61a and vcp as conserved regulators of sindbis virus entry.alphaviruses are a large class of insect-borne human pathogens and little is known about the host-factor requirements for infection. to identify such factors, we performed a genome-wide rnai screen using model drosophila cells and validated 94 genes that impacted infection of sindbis virus (sinv), the prototypical alphavirus. we identified a conserved role for sec61a and valosin-containing protein (vcp) in facilitating sinv entry in insects and mammals. sec61a and vcp selectively regulate traffi ...201324332855
[construction and characterization of egfp reporter gene labeled sindbis virus].to construct and characterize egfp reporter gene labeled sindbis virus (sinv).201324319966
enhanced arbovirus surveillance with deep sequencing: identification of novel rhabdoviruses and bunyaviruses in australian mosquitoes.viral metagenomics characterizes known and identifies unknown viruses based on sequence similarities to any previously sequenced viral genomes. a metagenomics approach was used to identify virus sequences in australian mosquitoes causing cytopathic effects in inoculated mammalian cell cultures. sequence comparisons revealed strains of liao ning virus (reovirus, seadornavirus), previously detected only in china, livestock-infecting stretch lagoon virus (reovirus, orbivirus), two novel dimarhabdov ...201324314645
carbocyclic 5'-nor "reverse" fleximers. design, synthesis, and preliminary biological activity.a series of 5'-nor carbocyclic "reverse" flexible nucleosides or "fleximers" have been designed wherein the nucleobase scaffold resembles a "split" purine as well as a substituted pyrimidine. this modification was employed to explore recognition by both purine and pyrimidine metabolizing enzymes. the synthesis of the carbocyclic fleximers and the results of their preliminary biological screening are described herein.201124312722
the no/cgmp pathway inhibits transient camp signals through the activation of pde2 in striatal neurons.the no-cgmp signaling plays an important role in the regulation of striatal function although the mechanisms of action of cgmp specifically in medium spiny neurons (msns) remain unclear. using genetically encoded fluorescent biosensors, including a novel epac-based sensor (epac-s(h150)) with increased sensitivity for camp, we analyze the cgmp response to no and whether it affected camp/pka signaling in msns. the cygnet2 sensor for cgmp reported large responses to no donors in both striatonigral ...201324302895
a genome-wide analysis of rna pseudoknots that stimulate efficient -1 ribosomal frameshifting or readthrough in animal viruses.programmed -1 ribosomal frameshifting (prf) and stop codon readthrough are two translational recoding mechanisms utilized by some rna viruses to express their structural and enzymatic proteins at a defined ratio. efficient recoding usually requires an rna pseudoknot located several nucleotides downstream from the recoding site. to assess the strategic importance of the recoding pseudoknots, we have carried out a large scale genome-wide analysis in which we used an in-house developed program to d ...201324298557
glun3a expression restricts spine maturation via inhibition of git1/rac1 signaling.nmda-type glutamate receptors (nmdars) guide the activity-dependent remodeling of excitatory synapses and associated dendritic spines during critical periods of postnatal brain development. whereas mature nmdars composed of glun1 and glun2 subunits mediate synapse plasticity and promote spine growth and stabilization, juvenile nmdars containing glun3a subunits are thought to inhibit these processes via yet unknown mechanisms. here, we report that glun3a binds g protein-coupled receptor kinase-in ...201324297929
rim1α sumoylation is required for fast synaptic vesicle exocytosis.the rapid, activity-dependent quantal presynaptic release of neurotransmitter is vital for brain function. the complex process of vesicle priming, fusion, and retrieval is very precisely controlled and requires the spatiotemporal coordination of multiple protein-protein interactions. here, we show that posttranslational modification of the active zone protein rab3-interacting molecule 1α (rim1α) by the small ubiquitin-like modifier 1 (sumo-1) functions as a molecular switch to direct these inter ...201324290762
pan-viral specificity of ifn-induced genes reveals new roles for cgas in innate immunity.the type i interferon (ifn) response protects cells from viral infection by inducing hundreds of interferon-stimulated genes (isgs), some of which encode direct antiviral effectors. recent screening studies have begun to catalogue isgs with antiviral activity against several rna and dna viruses. however, antiviral isg specificity across multiple distinct classes of viruses remains largely unexplored. here we used an ectopic expression assay to screen a library of more than 350 human isgs for eff ...201324284630
full inactivation of human influenza virus by high hydrostatic pressure preserves virus structure and membrane fusion while conferring protection to mice against infection.whole inactivated vaccines (wivs) possess greater immunogenicity than split or subunit vaccines, and recent studies have demonstrated that wivs with preserved fusogenic activity are more protective than non-fusogenic wivs. in this work, we describe the inactivation of human influenza virus x-31 by high hydrostatic pressure (hhp) and analyze the effects on the structure by spectroscopic measurements, light scattering, and electron microscopy. we also investigated the effects of hhp on the glycopr ...201324282553
rubella virus capsid protein structure and its role in virus assembly and infection.rubella virus (rv) is a leading cause of birth defects due to infectious agents. when contracted during pregnancy, rv infection leads to severe damage in fetuses. despite its medical importance, compared with the related alphaviruses, very little is known about the structure of rv. the rv capsid protein is an essential structural component of virions as well as a key factor in virus-host interactions. here we describe three crystal structures of the structural domain of the rv capsid protein. th ...201324282305
autophagy enhances the replication of classical swine fever virus in vitro.autophagy plays an important role in cellular responses to pathogens. however, the impact of the autophagy machinery on classical swine fever virus (csfv) infection is not yet confirmed. in this study, we showed that csfv infection significantly increases the number of autophagy-like vesicles in the cytoplasm of host cells at the ultrastructural level. we also found the formation of 2 ubiquitin-like conjugation systems upon virus infection, including lc3-i/lc3-ii conversion and atg12-atg5 conjug ...201324262968
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