Sequence Similarity Network Reveals Common Ancestry of Multidomain Proteins

We address the problem of homology identification in complex multidomain families with varied domain architectures. The challenge is to distinguish sequence pairs that share common ancestry from pairs that share an inserted domain but are otherwise unrelated. This distinction is essential for accuracy in gene annotation, function prediction, and comparative genomics. There are two major obstacles to multidomain homology identification: lack of a formal definition and lack of curated benchmarks for evaluating the performance of new methods. We offer preliminary solutions to both problems: 1) an extension of the traditional model of homology to include domain insertions; and 2) a manually curated benchmark of well-studied families in mouse and human. We further present Neighborhood Correlation, a novel method that exploits the local structure of the sequence similarity network to identify homologs with great accuracy based on the observation that gene duplication and domain shuffling leave distinct patterns in the sequence similarity network. In a rigorous, empirical comparison using our curated data, Neighborhood Correlation outperforms sequence similarity, alignment length, and domain architecture comparison. Neighborhood Correlation is well suited for automated, genome-scale analyses. It is easy to compute, does not require explicit knowledge of domain architecture, and classifies both single and multidomain homologs with high accuracy. Homolog predictions obtained with our method, as well as our manually curated benchmark and a web-based visualization tool for exploratory analysis of the network neighborhood structure, are available at http://www.neighborhoodcorrelation.org. Our work represents a departure from the prevailing view that the concept of homology cannot be applied to genes that have undergone domain shuffling. In contrast to current approaches that either focus on the homology of individual domains or consider only families with identical domain architectures, we show that homology can be rationally defined for multidomain families with diverse architectures by considering the genomic context of the genes that encode them. Our study demonstrates the utility of mining network structure for evolutionary information, suggesting this is a fertile approach for investigating evolutionary processes in the post-genomic era.     

A comparison of Anopheles gambiae and Plasmodium falciparum genetic structure over space and time

Population genetic structure and subdivision are key factors affecting the evolution of organisms. In this study, we analysed and compared the population genetic structure of the malaria parasite Plasmodium falciparum and its mosquito vector Anopheles gambiae over space and time in the Nianza Province, near Victoria Lake in Kenya. The parasites were collected from mosquitoes caught in six villages separated by up to 68 km in 2002 and 2003. A total of 545 oocysts were dissected from 122 infected mosquitoes and genotyped at seven microsatellite markers. Five hundred and forty-seven mosquitoes, both infected and uninfected, were genotyped at eight microsatellites. For the parasite and the vector, the analysis revealed no (or very little) genetic differentiation among villages. This may be explained by high local population sizes for the parasite and the mosquito. The small level of genetic differentiation observed between populations may explain the speed at which antimalarial drug resistance and insecticide resistance spread into the African continent.     

Correction of glycogenosis type 2 by muscle-specific lentiviral vector

Glycogen storage disease type II (GSDII) or Pompe disease is an inherited disease of glycogen metabolism caused by a lack of functional lysosomal acid α-glucosidase (GAA). Affected individuals store glycogen in lysosomes resulting in fatal hypertrophic cardiomyopathy and respiratory failure in the most severe form. We investigated for the first time the use of lentiviral vectors to correct the GSDII phenotype in human and murine GAA-deficient cells. Fibroblasts from infantile and adult GSDII patients were efficiently transduced by a GAA-expressing lentiviral vector placed under the control of the strong MND promoter, leading to a complete restoration of enzymatic activity. We also developed a muscle-specific lentiviral vector based on the synthetic C5–12 promoter and tested it on deficient myogenic satellite cells derived from a GSDII mouse model. GAA was expressed as a correctly processed protein allowing a complete enzymatic and metabolic correction in myoblasts and differentiated myotubes, as well as a significant mannose-6-phosphate (M6P)-dependent secretion reuptake by naive cells. Transduced cells showed lysosomal glycogen clearance, as demonstrated by electron microscopy. These results form the basis for a therapeutic approach of GSDII using lentiviral vector-mediated gene transfer into muscle stem cells.     

Is the hepatic metabolism of glucose and linoleic acid influenced by species in overfed ducks?

There are genetic differences in the hepatic glucose and linoleic acid metabolisms between Muscovy and Pekin ducks ad libitum-fed. To understand the effect of overfeeding on the hepatic metabolisms in these two species of ducks, we compared the different pathways of glucose and linoleic acid reaching the liver of Muscovy (Cairina moschata) (n = 6) and Pekin (Anas platyrhynchos) (n = 6) ducks overfed for 1 week and sacrificed 2–4 h after their last meal by using the ex vivo method of liver slices incubated for 16 h with [U-¹⁴C]-glucose, [1-¹⁴C]-linoleic acid and [³⁵S]-methionine added to the survival medium. The glucose was the main precursor of triacylglycerol synthesis in the liver of these two species and its hepatic metabolism was similar between species. The hepatic uptake of linoleic acid was 1.7-fold higher (P = 0.020) in the Muscovy duck than in the Pekin duck leading to a 1.9-fold higher (P = 0.017) esterification of this fatty acid in the liver of the Muscovy duck than in that of the Pekin duck. Finally, both species after 1 week of overfeeding exhibited the same capacity to secrete VLDL remaining insufficient to avoid hepatic steatosis.     

Assessment of the genetic diversity of Xanthomonas axonopodis pv. phaseoli and Xanthomonas fuscans subsp. fuscans as a basis to identify putative pathogenicity genes and a type III secretion system of the SPI-1 family by multiple suppression subtractive hybridizations

Fluorescent amplified fragment length polymorphism revealed that strains of Xanthomonas axonopodis pv. phaseoli and Xanthomonas fuscans subsp. fuscans are genetically distinct and can be grouped into four genetic lineages. Four suppression subtractive hybridizations were then performed to isolate DNA fragments present in these bean pathogens and absent from closely related xanthomonads. Virulence gene candidates were identified such as homologs of hemagglutinins, TonB-dependent receptors, zinc-dependent metalloproteases, type III effectors, and type IV secretion system components. Unexpectedly, homologs of the type III secretion apparatus components (SPI-1 family), usually reported in animal pathogens and insect symbionts, were also detected.     

The transport of sugars to developing embryos is not via the bulk endosperm in oilseed rape seeds

The fate of sucrose (Suc) supplied via the phloem to developing oilseed rape (Brassica napus) seeds has been investigated by supplying [(14)C]Suc to pedicels of detached, developing siliques. The method gives high, sustained rates of lipid synthesis in developing embryos within the silique comparable with those on the intact plant. At very early developmental stages (3 d after anthesis), the liquid fraction that occupies most of the interior of the seed has a very high hexose-to-Suc ratio and [(14)C]Suc entering the seeds is rapidly converted to hexoses. Between 3 and 12 d after anthesis, the hexose-to-Suc ratio of the liquid fraction of the seed remains high, but the fraction of [(14)C]Suc converted to hexose falls dramatically. Instead, most of the [(14)C]Suc entering the seed is rapidly converted to products in the growing embryo. These data, together with light and nuclear magnetic resonance microscopy, reveal complex compartmentation of sugar metabolism and transport within the seed during development. The bulk of the sugar in the liquid fraction of the seed is probably contained within the central vacuole of the endosperm. This sugar is not in contact with the embryo and is not on the path taken by carbon from the phloem to the embryo. These findings have important implications for the sugar switch model of embryo development and for understanding the relationship between the embryo and the surrounding endosperm.     

9 cis,11 trans conjugated linoleic acid (CLA) is synthesised and desaturated into conjugated 18:3 in bovine adipose tissues

Although endogenous synthesis of conjugated linoleic acid (CLA) in the mammary gland of lactating cows has been already well documented, no study has determined so far as to which tissue and/or organ is involved in CLA synthesis in the growing ruminant except one study showing that CLA synthesis does not occur in ruminant liver. In this context, adipose tissue appears to be a good candidate for endogenous synthesis of CLA in the growing ruminant. The aim of this study was to compare the respective metabolisms of 11trans 18:1 (vaccenic acid, VA) and 9cis,11trans 18:2 (rumenic acid) to that of stearic acid (the preferential substrate of Δ9 desaturase) in adipose tissues (subcutaneous, SC and intermuscular, IM) of six Charolais steers by using the in vitromethod of incubated tissue slices. Samples of SC and IM adipose tissues were incubated at 37°C for 16 h under an atmosphere of 95% O2/5% CO2 in a medium supplemented with 0.75 mM of fatty acid (FA) mixture (representative of circulating non-esterified FA) and 186 μM [1-14C]-18:0 or 58.6 μM [1-14C]-VA or 56 μM [1-14C]-9cis,11trans CLA. Viability of explants was verified by measuring metabolic functions (glucose uptake and glucose-6-phosphate dehydrogenase activity). After 16 h of incubation, FA uptake was similar for all FA (18:0, VA and 9cis,11trans 18:2) in both SC and IM adipose tissues (around 40%). Once in adipose tissue, all FA were preferentially esterified (>80% of cell FA) favouring neutral lipid synthesis (around 90% of esterified FA). Stearic acid was highly (27%) desaturated into oleic acid in SC adipose tissue whereas this desaturation was much lower (6.8%) in IM adipose tissue (P < 0.0001). VA was desaturated into 9cis,11trans CLA at a low extent of about 2.5% to 4.4% in both adipose tissues probably because of a limited affinity of Δ9 desaturase for VA. 9cis,11trans CLA was itself converted by desaturation into 6cis, 9cis,11trans 18:3 at the intensity of 10.8% and 14.5% of cell 9cis,11trans CLA in SC and IM adipose tissues, respectively. In conclusion, bovine adipose tissues of the growing ruminant were especially involved in the endogenous synthesis of CLA from VA and in its desaturation into conjugated derivative, mainly 6cis, 9cis,11trans 18:3, of which biological properties need to be elucidated.     

Food restriction and refeeding in lambs influence muscle antioxidant status

Compensatory growth, a frequent phenomenon observed in ruminants due to seasonal variation in food availability, affects protein metabolism including protein oxidation. These oxidation processes may have an impact on animal health as well as on meat protein degradation during post mortem aging (ie meat maturation). Sixteen male lambs were randomly divided into four groups. One group was fed ad libitum (C) and one group was food-restricted to 60% of the intake of the C group (R). The last two groups were restricted similarly to the R group and refed either ad libitum (RAL) or similarly to the C group (pair-feeding) (RPF). Muscles samples were taken immediately after slaughter. The present study showed that the restriction/refeeding pattern had no effect on protein oxidation in the muscles studied (longissimus dorsi (LD), semitendinosus (ST) and supraspinatus (SP)). However, total antioxidant capacity decreased after food restriction (-51%, -43%, P < 0.01 for ST and LD muscles, respectively) and re-increased only after ad libitum refeeding. This alteration in the total antioxidant status can partially be explained by the similar pattern of change observed in the glutathione concentration of the muscles (-25%, P < 0.05 for ST muscle and NS for the other muscles). However, none of the concentrations of other water-soluble antioxidants studied (carnosine, anserine, glutathione peroxidase and superoxide dismutase) were altered during compensatory growth. This study showed that an inappropriate feeding level following a nutritional stress induced alterations in the total antioxidant status (particularly that of glutathione), which may have consequences on animal health. Other consequences of a decrease of the animal antioxidant status in vivo could be an alteration of the protein oxidation processes during meat maturation.     

INS VNTR is not associated with childhood obesity in 1,023 families: a family-based study

Previous studies have described genetic associations of the insulin gene variable number tandem repeat (INS VNTR) variant with childhood obesity and associated phenotypes. We aimed to assess the contribution of INS VNTR genotypes to childhood obesity and variance of insulin resistance, insulin secretion, and birth weight using family-based design. Participants were either French or German whites. We used transmission disequilibrium tests (TDTs) for assessing binary traits and quantitative pedigree disequilibrium tests for assessing continuous traits. In contrast to previous findings, we did not observe any familial association with childhood obesity (T = 50%, P = 0.77) in the 1,023 families tested. In French obese children, INS VNTR did not associate with fasting insulin levels (P = 0.23) and class I allele showed only borderline association with increased insulin secretion index at 30 min (P = 0.03). INS VNTR did not associate with birth weight in obese children (P = 0.98) and TDT analyses in 350 French families with history of low birth weight (LBW) showed no association with this condition (P = 0.92). In summary, our study, the largest performed so far, does not support the previously reported associations between INS VNTR and childhood obesity, insulin resistance, or birth weight, and does not suggest any major role for this variant in modulating these traits.