Algorithms for matching partially labelled sequence graphs

Background

In order to find correlated pairs of positions between proteins, which are useful in predicting interactions, it is necessary to concatenate two large multiple sequence alignments such that the sequences that are joined together belong to those that interact in their species of origin. When each protein is unique then the species name is sufficient to guide this match, however, when there are multiple related sequences (paralogs) in each species then the pairing is more difficult. In bacteria a good guide can be gained from genome co-location as interacting proteins tend to be in a common operon but in eukaryotes this simple principle is not sufficient.

Results

The methods developed in this paper take sets of paralogs for different proteins found in the same species and make a pairing based on their evolutionary distance relative to a set of other proteins that are unique and so have a known relationship (singletons). The former constitute a set of unlabelled nodes in a graph while the latter are labelled. Two variants were tested, one based on a phylogenetic tree of the sequences (the topology-based method) and a simpler, faster variant based only on the inter-sequence distances (the distance-based method). Over a set of test proteins, both gave good results, with the topology method performing slightly better.

Conclusions

The methods develop here still need refinement and augmentation from constraints other than the sequence data alone, such as known interactions from annotation and databases, or non-trivial relationships in genome location. With the ever growing numbers of eukaryotic genomes, it is hoped that the methods described here will open a route to the use of these data equal to the current success attained with bacterial sequences.

     

Inducing Ni sensitivity in the Ni hyperaccumulator plant <Emphasis Type="Italic">Alyssum inflatum</Emphasis> Nyárády (Brassicaceae) by transforming with <Emphasis Type="Italic">CAX1</Emphasis>, a vacuolar membrane calcium transporter

The importance of calcium in nickel tolerance was studied in the nickel hyperaccumulator plant Alyssum inflatum by gene transformation of CAX1, a vacuolar membrane transporter that reduces cytosolic calcium. CAX1 from Arabidopsis thaliana with a CaMV35S promoter accompanying a kanamycin resistance gene was transferred into A. inflatum using Agrobacterium tumefaciens. Transformed calli were sub-cultured three times on kanamycin-rich media and transformation was confirmed by PCR using a specific primer for CAX1. At least 10 callus lines were used as a pool of transformed material. Both transformed and untransformed calli were treated with varying concentrations of either calcium (1–15 mM) or nickel (0–500 µM) to compare their responses to those ions. Increased external calcium generally led to increased callus biomass, however, the increase was greater for untransformed callus. Further, increased external calcium led to increased callus calcium concentrations. Transformed callus was less nickel tolerant than untransformed callus: under increasing nickel concentrations callus relative growth rate was significantly less for transformed callus. Transformed callus also contained significantly less nickel than untransformed callus when exposed to the highest external nickel concentration (200 µM). We suggest that transformation with CAX1 decreased cytosolic calcium and resulted in decreased nickel tolerance. This in turn suggests that, at low cytosolic calcium concentrations, other nickel tolerance mechanisms (e.g., complexation and vacuolar sequestration) are insufficient for nickel tolerance. We propose that high cytosolic calcium is an important mechanism that results in nickel tolerance by nickel hyperaccumulator plants.     

Functional expression of human pyruvate carboxylase for reduced lactic acid formation of Chinese hamster ovary cells (DG44)

To investigate the effect of human pyruvate carboxylase (hPC) on lactate formation in Chinese hamster ovary (CHO) cell lines, FLAG-tagged hPC was introduced into a dihydrofolate-deficient CHO cell line (DG44). Three clones expressing high levels of hPC, determined by Western blotting using an anti-FLAG monoclonal antibody, and a control cell line were established. Immunocytochemistry revealed that a substantial amount of expressed hPC protein was localized in the mitochondria of the cells. hPC expression did not impair cell proliferation. Rather, it improved cell viability at the end of adherent batch cultures with the serum-containing medium probably because of reduced lactate formation. Compared with control cells, specific lactate production rate of the three clones was decreased by 21–39%, which was because of a decreased specific glucose uptake rate and yield of lactate from glucose. Reduced lactate formation by hPC expression was also observed in suspension fed-batch cultures using a serum-free medium. Taken together, these results demonstrate that through the expression of the hPC enzyme, lactate formation in CHO cell culture can be efficiently reduced.     

Biogenic and Risk Elements in Wines from the Slovak Market with the Estimation of Consumer Exposure

Wine consumption delivers macroelements and microelements necessary for the proper metabolism. On the other hand, wine can be an important source of toxic metals. The aim of this study was to estimate the concentrations of Ca, Cd, Cu, Fe, Hg, Mg, Ni, Pb, and Zn in the Slovak and non-Slovak wines. The concentration of metals was evaluated with respect to the type, the alcohol content, and the age of Slovak wine. The general scheme of concentrations found was as follows Ca > Mg > Fe > Zn > Pb > Cd > Ni > Cu > Hg. The type of wine and the alcohol content do not have a significant impact on metal concentrations. Also, the age of wine has no influence on the mean concentration of metals, except for Zn. Metal concentrations in Slovak and non-Slovak wines indicate similar contents of metals, except for Ni. The contribution to both dietary reference values (DRVs) and provisional tolerable weekly intake (PTWI) evaluations in the Slovak wine suggested low dietary exposure to Ca, Cu, Fe, Mg, Ni, Zn, Cd, Hg, and Pb, respectively. However, we do not suggest that the consumption of all Slovak wines is healthy. The maximum Pb concentrations in Slovak wines exceed the maximum permitted level proposed by the European Commission. This might be proved by the results of the margin of the exposure (MOE) value evaluation in the samples containing the maximum Pb concentrations, showing a high risk of CKD and SBP in high and extreme consumption groups.     

Characteristics of Environmental <Emphasis Type="Italic">Paracoccidioides brasiliensis</Emphasis> Isolates

The ecological niche or exact habitat of the fungus Paracoccidioides brasiliensis is not known, and few isolates have been obtained from the environment. In this study, ten isolates were analyzed with respect to antigenic composition, serology, pathogenicity, and molecular aspects. Gp43 is considered to be the molecular basis for the serodiagnosis of paracoccidioidomycosis; however, in this study only six of the environmental isolates secreted this molecule (four in great amounts and two in small amounts). Other molecules were also produced. When exoantigens from these isolates were tested using immunodiffusion, only four preparations were positive by ID tests. However, when these exoantigens were tested by ELISA, all of them except one were able to detect anti-P. brasiliensis antibodies. In Western blot assays, these exoantigens showed different reactivities. Isolates that secreted gp43 presented positive reactions for this molecule, and isolates that did not secrete gp43 gave positive reactions for other minor molecules. RAPD analysis revealed that there is great genetic variation between these environmental isolates. These isolates were non-pathogenic: no mortality was observed among the inoculated mice during an 18-month follow-up period.     

DNMT3b 39179G‎T Polymorphism and the Risk of Adenocarcinoma of the Colon in Koreans

DNA-methyltransferase-3B (DNMT3b) plays an important role in the generation of aberrant methylation in carcinogenesis. DNMT3b SNP has been associated with susceptibility to lung, head, neck, and breast cancer, but its association with the development of colon cancer has not been reported. We investigated the relationship between the 39179G‎T polymorphism in the DNMT3b gene, which is involved in de novo methylation and is associated with the risk of adenocarcinoma of the colon in Koreans. The DNMT3b 39179G‎T genotypes were determined by a PCR-RFLP method in 248 adenocarcinomas of colon cancer patients and in 248 healthy controls matched as to age and sex. When stratified by sex and age, a significantly reduced risk of the combined GT and GG genotypes was observed in younger patients (<59, adjusted OR = 0.255, 95% CI = 0.133–0.489) and in male patients (adjusted OR = 0.383, 95% CI = 0.225–0.652). The DNMT3b 39179G‎T polymorphism may be a genetic determinant of adenocarcinoma of the colon, especially in younger Korean men.     

A second-generation diagnostic single nucleotide polymorphism (SNP)-based assay, optimized to distinguish among eight poplar (Populus L.) species and their early hybrids

Rapid identification of Populus L. species and hybrids can be achieved with relatively little effort through the use of primer extension-based single nucleotide polymorphism (SNP) genotyping assays. We present an optimized set of 36 SNP markers from 28 gene regions that diagnose eight poplar species (Populus angustifolia James, Populus balsamifera L., Populus deltoides Bartram, Populus fremontii Watson, Populus laurifolia Ledeb., Populus maximowiczii Henry, Populus nigra L., and Populus trichocarpa Torr. & Gray). A total of 700 DNA sequences from six Populus species (1–15 individuals per species) were used to construct the array. A set of flanking and probe oligonucleotides was developed and tested. The accuracy of the SNP assay was validated by genotyping 448 putatively “pure” individuals from 14 species of Populus. Overall, the SNP assay had a high success rate (97.6 %) and will prove useful for the identification of all Aigeiros Duby and Tacamahaca Spach. species and their early-generation hybrids within natural populations and breeding programs. Null alleles and intraspecific polymorphisms were detected for a few locus/species combinations in the Aigeiros and Tacamahaca sections. When we attempted to genotype aspens of the section Populus (Populus alba L., Populus grandidentata Michx., Populus tremula L., and Populus tremuloides Michx.), the success rate of the SNP array decreased by 13 %, demonstrating moderate cross-sectional transferability.     

Patient and observer reported outcome measures to evaluate health-related quality of life in inherited metabolic diseases: a scoping review

Background

Health-related Quality of Life (HrQoL) is a multidimensional measure, which has gained clinical and social relevance. Implementation of a patient-centred approach to both clinical research and care settings, has increased the recognition of patient and/or observer reported outcome measures (PROMs or ObsROMs) as informative and reliable tools for HrQoL assessment. Inherited Metabolic Diseases (IMDs) are a group of heterogeneous conditions with phenotypes ranging from mild to severe and mostly lacking effective therapies. Consequently, HrQoL evaluation is particularly relevant.

Objectives

We aimed to: (1) identify patient and/or caregiver-reported HrQoL instruments used among IMDs; (2) identify the main results of the application of each HrQoL tool and (3) evaluate the main limitations of HrQoL instruments and study design/methodology in IMDs.

Methods

A scoping review was conducted using methods outlined by Arksey and O’Malley. Additionally, we critically analysed each article to identify the HrQoL study drawbacks.

Results

Of the 1954 studies identified, 131 addressed HrQoL of IMDs patients using PROMs and/or ObsROMs, both in observational or interventional studies. In total, we identified 32 HrQoL instruments destined to self- or proxy-completion; only 2% were disease-specific. Multiple tools (both generic and disease-specific) proved to be responsive to changes in HrQoL; the SF-36 and PedsQL questionnaires were the most frequently used in the adult and pediatric populations, respectively. Furthermore, proxy data often demonstrated to be a reliable approach complementing self-reported HrQoL scores. Nevertheless, numerous limitations were identified especially due to the rarity of these conditions.

Conclusions

HrQoL is still not frequently assessed in IMDs. However, our results show successful examples of the use of patient-reported HrQoL instruments in this field. The importance of HrQoL measurement for clinical research and therapy development, incites to further research in HrQoL PROMs’ and ObsROMs’ creation and validation in IMDs.

     

Strain-induced inflammation in pulmonary alveolar tissue due to mechanical ventilation

Inflammation is the body’s attempt at self-protection to remove harmful stimuli, including damaged cells, irritants, or pathogens and begin the healing process. In this study, strain-induced inflammation in pulmonary alveolar tissue under high tidal volume is investigated through a combination of an inflammation model and fluid structure interaction (FSI) analysis. A realistic three-dimensional organ model for alveolar sacs is built, and FSI is employed to evaluate strain distribution in alveolar tissue for different tidal volume (TV) values under the mechanical ventilation (MV) condition. The alveolar tissue is treated as a hyperelastic solid and provides the environment for the tissue constituents. The influence of different strain distributions resulting from different tidal volumes is investigated. It is observed that strain is highly distributed in the inlet area. In addition, strain versus time curves in different locations through the alveolar model reveals that middle layers in the alveolar region would undergo higher levels of strain during breathing under the MV condition. Three different types of strain distributions in the alveolar region from the FSI simulation are transferred to the CA model to study population dynamics of cell constituents under MV for different TVs; 200, 500 and 1000 mL, respectively. The CA model results suggests that strain distribution plays a significant role in population dynamics. An interplay between strain magnitude and distribution appears to influence healing capability. Results suggest that increasing TV leads to an exponential rise in tissue damage by inflammation.