Sparse Rational Univariate Representation

International audienceWe present explicit worst case degree and height bounds for the rational univariate representation of the isolated roots of polynomial systems based on mixed volume. We base our estimations on height bounds of resultants and we consider the case of 0-dimensional, positive dimensional, and parametric polynomial systems

Effects of riparian plant diversity loss on aquatic microbial decomposers become more pronounced at longer times

We examined the potential long-term impacts of riparian plant diversity loss on diversity and activity of aquatic microbial decomposers. Microbial assemblages were obtained in a mixed-forest stream by immersion of mesh bags contain-ing three leaf species (alder, oak and eucalyptus), commonly found in riparian corridors of Iberian streams. Simulation of species loss was done in microcosms by including a set of all leaf species, retrieved from the stream, and non-colonized leaves of three, two or one leaf species. Leaves were renewed every month throughout six months, and microbial inoculum was ensured by a set of colonized leaves from the previous month. Microbial diversity, leaf mass loss and fungal biomass were assessed at the second and sixth months after plant species loss. Molecular diversity of fungi and bacteria, as the total number of operational taxonomic units per leaf diversity treatment, decreased with leaf diversity loss. Fungal biomass tended to decrease linearly with leaf species loss on oak and eucalyptus, suggesting more pronounced effects of leaf diver-sity on lower quality leaves. Decomposition of alder and eucalyptus leaves was affected by leaf species identity, mainly after longer times following diversity loss. Leaf decomposi-tion of alder decreased when mixed with eucalyptus, while decomposition of eucalyptus decreased in mixtures with oak. Results suggest that the effects of leaf diversity on microbial decomposers depended on leaf species number and also on which species were lost from the system, especially after longer times. This may have implications for the management of riparian forests to maintain stream ecosystem functioning.FEDER-POFC-COMPETE and the Portuguese Foundation for Science and Technology supported this study (PEst-C/ BIA/UI4050/2011, PTDC/AAC-AMB/113746/2009 and PTDC/AAC-AMB/117068/2010), S. Duarte (SFRH/BPD/47574/2008) and I. Fernandes (SFRH/BD/42215/2007)

A survey among dermatologists: diagnostics of superficial fungal infections - what is used and what is needed to initiate therapy and assess efficacy?

BACKGROUND: Superficial fungal infections are common. It is important to confirm the clinical diagnosis by mycological laboratory methods before initiating systemic antifungal treatment, especially as antifungal sensitivity and in vitro susceptibility may differ between different genera and species. For many years, the gold standard for diagnosis of superficial fungal infections has been direct fungal detection in the clinical specimen (microscopy) supplemented by culturing. Lately, newer molecular based methods for fungal identification have been developed. OBJECTIVE: This study was initiated to focus on the current usage of mycological diagnostics for superficial fungal infections by dermatologists. It was designed to investigate whether it was necessary to differentiate between initial diagnostic tests and those used at treatment follow-up in specific superficial fungal infections. METHODS: An online questionnaire was distributed among members of the EADV mycology Task Force and other dermatologists with a special interest in mycology and nail disease. RESULTS: The survey was distributed to 62 dermatologists of whom 38 (61%) completed the whole survey, 7 (11%) partially completed and 17 (27%) did not respond. Nearly, all respondents (82-100%) said that ideally they would use the result of direct microscopy (or histology) combined with a genus/species directed treatment of onychomycosis, dermatophytosis, Candida- and Malassezia-related infections. The majority of the dermatologists used a combination of clinical assessment and direct microscopy for treatment assessment and the viability of the fungus was considered more important at this visit than when initiating the treatment. Molecular based methods were not available for all responders. CONCLUSION: The available diagnostic methods are heterogeneous and their usage differs between different practices as well as between countries. The survey confirmed that dermatologists find it important to make a mycological diagnosis, particularly prior to starting oral antifungal treatment in order to confirm the diagnose and target the therapy according to genus and species

Relevance of large litter bag burial for the study of leaf breakdown in the hyporheic zone

Particulate organic matter is the major source of energy for most low-order streams, but a large part of this litter is buried within bed sediment during floods and thus become poorly available for benthic food webs. The fate of this buried litter is little studied. In most cases, measures of breakdown rates consist of burying a known mass of litter within the stream sediment and following its breakdown over time. We tested this method using large litter bags (15 x 15 cm) and two field experiments. First, we used litter large bags filled with Alnus glutinosa leaves (buried at 20 cm depth with a shovel) in six stations within different land-use contexts and with different sediment grain sizes. Breakdown rates were surprisingly high (0.0011–0.0188 day-1) and neither correlate with most of the physico-chemical characteristics measured in the interstitial habitats nor with the land-use around the stream. In contrast, the rates were negatively correlated with a decrease in oxygen concentrations between surface and buried bags and positively correlated with both the percentage of coarse particles (20–40 mm) in the sediment and benthic macro-invertebrate richness. These results suggest that the vertical exchanges with surface water in the hyporheic zone play a crucial role in litter breakdown. Second, an experimental modification of local sediment (removing fine particles with a shovel to increase vertical exchanges) highlighted the influence of grain size on water and oxygen exchanges, but had no effect on hyporheic breakdown rates. Burying large litter bags within sediments may thus not be a relevant method, especially in clogged conditions, due to changes induced through the burial process in the vertical connectivity between surface and interstitial habitats that modify organic matter processing

Bifunctional Anti-Huntingtin Proteasome-Directed Intrabodies Mediate Efficient Degradation of Mutant Huntingtin Exon 1 Protein Fragments

Huntington's disease (HD) is a fatal autosomal dominant neurodegenerative disorder caused by a trinucleotide (CAG)n repeat expansion in the coding sequence of the huntingtin gene, and an expanded polyglutamine (>37Q) tract in the protein. This results in misfolding and accumulation of huntingtin protein (htt), formation of neuronal intranuclear and cytoplasmic inclusions, and neuronal dysfunction/degeneration. Single-chain Fv antibodies (scFvs), expressed as intrabodies that bind htt and prevent aggregation, show promise as immunotherapeutics for HD. Intrastriatal delivery of anti-N-terminal htt scFv-C4 using an adeno-associated virus vector (AAV2/1) significantly reduces the size and number of aggregates in HDR6/1 transgenic mice; however, this protective effect diminishes with age and time after injection. We therefore explored enhancing intrabody efficacy via fusions to heterologous functional domains. Proteins containing a PEST motif are often targeted for proteasomal degradation and generally have a short half life. In ST14A cells, fusion of the C-terminal PEST region of mouse ornithine decarboxylase (mODC) to scFv-C4 reduces htt exon 1 protein fragments with 72 glutamine repeats (httex1-72Q) by ∼80–90% when compared to scFv-C4 alone. Proteasomal targeting was verified by either scrambling the mODC-PEST motif, or via proteasomal inhibition with epoxomicin. For these constructs, the proteasomal degradation of the scFv intrabody proteins themselves was reduced<25% by the addition of the mODC-PEST motif, with or without antigens. The remaining intrabody levels were amply sufficient to target N-terminal httex1-72Q protein fragment turnover. Critically, scFv-C4-PEST prevents aggregation and toxicity of httex1-72Q fragments at significantly lower doses than scFv-C4. Fusion of the mODC-PEST motif to intrabodies is a valuable general approach to specifically target toxic antigens to the proteasome for degradation

Selection and characterisation of a phage-displayed human antibody (Fab) reactive to the lung resistance-related major vault protein

The major vault protein is the main component on multimeric vault particles, that are likely to play an essential role in normal cell physiology and to be associated with multidrug resistance of tumour cells. In order to unravel the function of vaults and their putative contribution to multidrug resistance, specific antibodies are invaluable tools. Until now, only conventional major vault protein-reactive murine monoclonal antibodies have been generated, that are most suitable for immunohistochemical analyses. The phage display method allows for selection of human antibody fragments with potential use in clinical applications. Furthermore, cDNA sequences encoding selected antibody fragments are readily identified, facilitating various molecular targeting approaches. In order to obtain such human Fab fragments recognising major vault protein we used a large non-immunized human Fab fragment phage library. Phages displaying major vault protein-reactive Fabs were obtained through several rounds of selection on major vault protein-coated immunotubes and subsequent amplification in TG1 E coli bacteria. Eventually, one major vault protein-reactive clone was selected and further examined. The anti-major vault protein Fab was found suitable for immunohistochemical and Western blot analysis of tumour cell lines and human tissues. BIAcore analysis showed that the binding affinity of the major vault protein-reactive clone almost equalled that of the murine anti-major vault protein Mabs. The cDNA sequence of this human Fab may be exploited to generate an intrabody for major vault protein-knock out studies. Thus, this human Fab fragment should provide a valuable tool in elucidating the contribution(s) of major vault protein/vaults to normal physiology and cellular drug resistance mechanisms

An Engineered Viral Protease Exhibiting Substrate Specificity for a Polyglutamine Stretch Prevents Polyglutamine-Induced Neuronal Cell Death

BACKGROUND: Polyglutamine (polyQ)-induced protein aggregation is the hallmark of a group of neurodegenerative diseases, including Huntington's disease. We hypothesized that a protease that could cleave polyQ stretches would intervene in the initial events leading to pathogenesis in these diseases. To prove this concept, we aimed to generate a protease possessing substrate specificity for polyQ stretches. METHODOLOGY/PRINCIPAL FINDINGS: Hepatitis A virus (HAV) 3C protease (3CP) was subjected to engineering using a yeast-based method known as the Genetic Assay for Site-specific Proteolysis (GASP). Analysis of the substrate specificity revealed that 3CP can cleave substrates containing glutamine at positions P5, P4, P3, P1, P2', or P3', but not substrates containing glutamine at the P2 or P1' positions. To accommodate glutamine at P2 and P1', key residues comprising the active sites of the S2 or S1' pockets were separately randomized and screened. The resulting sets of variants were combined by shuffling and further subjected to two rounds of randomization and screening using a substrate containing glutamines from positions P5 through P3'. One of the selected variants (Var26) reduced the expression level and aggregation of a huntingtin exon1-GFP fusion protein containing a pathogenic polyQ stretch (HttEx1(97Q)-GFP) in the neuroblastoma cell line SH-SY5Y. Var26 also prevented cell death and caspase 3 activation induced by HttEx1(97Q)-GFP. These protective effects of Var26 were proteolytic activity-dependent. CONCLUSIONS/SIGNIFICANCE: These data provide a proof-of-concept that proteolytic cleavage of polyQ stretches could be an effective modality for the treatment of polyQ diseases

Genetic markers and phosphoprotein forms of beta-catenin pβ-Cat552 and pβ-Cat675 are prognostic biomarkers of cervical cancer

BACKGROUND: Cervical cancer (CC) remains a leading cause of gynaecological cancer-related mortality world wide and constitutes the third most common malignancy in women. The RAIDs consortium (http://www. raids-fp7.eu/) conducted a prospective European study [BioRAIDs (NCT02428842)] with the objective to stratify CC patients for innovative treatments. A “metagene” of genomic markers in the PI3K pathway and epigenetic regulators had been previously associated with poor outcome [2]. METHODS: To detect new, more specific, targets for treatment of patients who resist standard chemo-radiation, a high-dimensional Cox model was applied to define dominant molecular variants, copy number variations, and reverse phase protein arrays (RPPA). FINDINGS: Survival analysis on 89 patients with all omics data available, suggested loss-of-function (LOF) or activating molecular alterations in nine genes to be candidate biomarkers for worse prognosis in patients treated by chemo-radiation while LOF of ATRX, MED13 as well as CASP8 were associated with better prognosis. When protein expression data by RPPA were factored in, the supposedly low molecula

Intraspecific traits change biodiversity effects on ecosystem functioning under metal stress

The online version of this article (doi:10.1007/s00442-011-1930-3) contains supplementary material, which is available to authorized users.Studies investigating the impacts of biodiversity loss on ecosystem processes have often reached different conclusions, probably because insufficient attention has been paid to some aspects including (1) which biodiversity measure (e.g., species number, species identity or trait) better explains ecosystem functioning, (2) the mechanisms underpinning biodiversity effects, and (3) how can environmental context modulates biodiversity effects. Here, we investigated how species number (one to three species) and traits of aquatic fungal decomposers (by replacement of a functional type from an unpolluted site by another from a metal-polluted site) affect fungal production (biomass acumulation) and plant litter decomposition in the presence and absence of metal stress. To examine the putative mechanisms that explain biodiversity effects, we determined the contribution of each fungal species to the total biomass produced in multicultures by real-time PCR. In the absence of metal, positive diversity effects were observed for fungal production and leaf decomposition as a result of species complementarity. Metal stress decreased diversity effects on leaf decomposition in assemblages containing the functional type from the unpolluted site, probably due to competitive interactions between fungi. However, dominance effect maintained positive diversity effects under metal stress in assemblages containing the functional type from the metal-polluted site. These findings emphasize the importance of intraspecific diversity in modulating diversity effects under metal stress, providing evidence that trait-based diversity measures should be incorporated when examining biodiversity effects.The Portuguese Foundation for Science and Technology supported I. Fernandes (SFRH/BD/42215/2007