An Excess of Deleterious Variants in VEGF-A Pathway Genes in Down-Syndrome-Associated Atrioventricular Septal Defects

About half of people with trisomy 21 have a congenital heart defect (CHD), whereas the remainder have a structurally normal heart, demonstrating that trisomy 21 is a significant risk factor but is not causal for abnormal heart development. Atrioventricular septal defects (AVSD) are the most commonly occurring heart defects in Down syndrome (DS), and ∼65% of all AVSD is associated with DS. We used a candidate-gene approach among individuals with DS and complete AVSD (cases = 141) and DS with no CHD (controls = 141) to determine whether rare genetic variants in genes involved in atrioventricular valvuloseptal morphogenesis contribute to AVSD in this sensitized population. We found a significant excess (p < 0.0001) of variants predicted to be deleterious in cases compared to controls. At the most stringent level of filtering, we found potentially damaging variants in nearly 20% of cases but fewer than 3% of controls. The variants with the highest probability of being damaging in cases only were found in six genes: COL6A1, COL6A2, CRELD1, FBLN2, FRZB, and GATA5. Several of the case-specific variants were recurrent in unrelated individuals, occurring in 10% of cases studied. No variants with an equal probability of being damaging were found in controls, demonstrating a highly specific association with AVSD. Of note, all of these genes are in the VEGF-A pathway, even though the candidate genes analyzed in this study represented numerous biochemical and developmental pathways, suggesting that rare variants in the VEGF-A pathway might contribute to the genetic underpinnings of AVSD in humans.     

Roles of mouse sperm‐associated alpha‐L‐fucosidases in fertilization

Sperm‐associated α‐L ‐fucosidases have been implicated in fertilization in many species. Previously, we documented the existence of α‐L ‐fucosidase in mouse cauda epididymal contents, and showed that sperm‐associated α‐L ‐fucosidase is cryptically stored within the acrosome and reappears within the sperm equatorial segment after the acrosome reaction. The enrichment of sperm membrane‐associated α‐L ‐fucosidase within the equatorial segment of acrosome‐reacted cells implicates its roles during fertilization. Here, we document the absence of α‐L ‐fucosidase in mouse oocytes and early embryos, and define roles of sperm associated α‐L ‐fucosidase in fertilization using specific inhibitors and competitors. Mouse sperm were pretreated with deoxyfuconojirimycin (DFJ, an inhibitor of α‐L ‐fucosidase) or with anti‐fucosidase antibody; alternatively, mouse oocytes were pretreated with purified human liver α‐L ‐fucosidase. Five‐millimolar DFJ did not inhibit sperm–zona pellucida (ZP) binding, membrane binding, or fusion and penetration, but anti‐fucosidase antibody and purified human liver α‐L ‐fucosidase significantly decreased the frequency of these events. To evaluate sperm‐associated α‐L ‐fucosidase enzyme activity in post‐fusion events, DFJ‐pretreated sperm were microinjected into oocytes, and 2‐pronuclear (2‐PN) embryos were treated with 5 mM DFJ with no significant effects, suggesting that α‐L ‐fucosidase enzyme activity does not play a role in post‐fusion events and/or early embryo development in mice. The recognition and binding of mouse sperm to the ZP and oolemma involves the glycoprotein structure of α‐L ‐fucosidase, but not its catalytic action. These observations suggest that deficits in fucosidase protein and/or the presence of anti‐fucosidase antibody may be responsible for some types of infertility. Mol. Reprod. Dev. 80: 273–285, 2013. © 2013 Wiley Periodicals, Inc.     

The Active and the Inactive Form of the hAT1 Receptor Have an Identical Ligand-Binding Environment: An MPA Study on a Constitutively Active Angiotensin II Receptor Mutant

Several models of activation mechanisms were proposed for G protein-coupled receptors (GPCRs), yet no direct methods exist for their elucidation. The availability of constitutively active mutants has given an opportunity to study active receptor conformations within acceptable limits using models such as the angiotensin II type 1 (AT₁)¹ receptor mutant N111G-hAT₁ which displays an important constitutive activity. Recently, by using methionine proximity assay, we showed for the hAT₁ receptor that TMD III, VI, and VII form the ligand-binding pocket of the C-terminal amino acid of an antagonistic AngII analogue. In the present contribution, we investigated whether the same residues would also constitute the ligand-binding contacts in constitutively activated mutant (CAM) receptors. For this purpose, the same Met mutagenesis strategy was carried out on the N111G double mutants. Analysis of 43 receptors mutants in the N111G-hAT₁ series, photolabeled and CNBr digested, showed that there were only subtle structural changes between the wt-receptor and its constitutively active form.     

Cryopreservation and storage of mussel (Mytilus spp.) haemocytes for latent analysis by the Comet assay

Estuarine and coastal habitats are known to be polluted by a range of chemical contaminants from both industrial and domestic sources. Blue mussels (Mytilus spp.), which inhabit these areas, are widely used as bio-indicators in eco-toxicological studies, because of their sedentary nature and their ability to bio-accumulate contaminants. The analysis of DNA damage in mussel haemocytes is a valuable tool for biomonitoring but sampling issues related to storage, handling and transportation have often limited its application in large-scale monitoring programmes. This study uses a trial and error method to evaluate and validate a suitable protocol for cryopreservation of mussel haemocytes, thereby allowing material collected in the field to be analysed later under controlled laboratory conditions. Three different cell-culture media, i.e. Leibovitz-15, Hank's balanced salt solution and mussel physiological saline, along with four different cryoprotectants, i.e. dimethyl sulphoxide (10% and 20%), 1,2-propanediol (10%), ethylene glycol (10%) and glycerol (10%) were tested to assess their suitability for cryopreservation of mussel haemocytes for analysis in the comet assay. Experimental studies where mussel haemocytes were also exposed to UV radiation or benzo(a)pyrene were conducted in order to mimic environmental stresses and to verify the effectiveness of newly defined cryopreservation protocols. The comet assay was used to demonstrate that mussel haemocytes could be preserved at cryogenic temperatures for a month without altering levels of DNA damage, which could possibly be used for lab or field studies where time constraints or facilities do not allow instant analysis.     

Climate‐induced changes in the distribution of freshwater fish: observed and predicted trends

1. Climate change could be one of the main threats faced by aquatic ecosystems and freshwater biodiversity. Improved understanding, monitoring and forecasting of its effects are thus crucial for researchers, policy makers and biodiversity managers. 2. Here, we provide a review and some meta‐analyses of the literature reporting both observed and predicted climate‐induced effects on the distribution of freshwater fish. After reviewing three decades of research, we summarise how methods in assessing the effects of climate change have evolved, and whether current knowledge is geographically or taxonomically biased. We conducted multispecies qualitative and quantitative analyses to find out whether the observed responses of freshwater fish to recent changes in climate are consistent with those predicted under future climate scenarios. 3. We highlight the fact that, in recent years, freshwater fish distributions have already been affected by contemporary climate change in ways consistent with anticipated responses under future climate change scenarios: the range of most cold‐water species could be reduced or shift to higher altitude or latitude, whereas that of cool‐ and warm‐water species could expand or contract. 4. Most evidence about the effects of climate change is underpinned by the large number of studies devoted to cold‐water fish species (mainly salmonids). Our knowledge is still incomplete, however, particularly due to taxonomic and geographic biases. 5. Observed and expected responses are well correlated among families, suggesting that model predictions are supported by empirical evidence. The observed effects are of greater magnitude and show higher variability than the predicted effects, however, indicating that other drivers of changes may be interacting with climate and seriously affecting freshwater fish. 6. Finally, we suggest avenues of research required to address current gaps in what we know about the climate‐induced effects on freshwater fish distribution, including (i) the need for more long‐term data analyses, (ii) the assessment of climate‐induced effects at higher levels of organisation (e.g. assemblages), (iii) methodological improvements (e.g. accounting for uncertainty among projections and species’ dispersal abilities, combining both distributional and empirical approaches and including multiple non‐climatic stressors) and (iv) systematic confrontation of observed versus predicted effects across multi‐species assemblages and at several levels of biological organisation (i.e. populations and assemblages).     

No relation between ACEml/D polymorphism and high altitude pulmonary edema in the Han Chinese

Objectives To explore whether the angiotensin T -converting enzyme （ACE） I/D （insertion/ deletion） polymorphism is associated with the susceptibility to high altitude pulmonary edema （HAPE） in the Han Chinese. Methods One hundred and forty-seven HAPE-p （HAPE patients） and 193 HAPE-r （HAPE resistants） were enrolled from the Yushu earthquake reconstruction workers in Qinghai province where the altitude is over 3 500 m above sea level. Blood samples were collected from each of the HAPE-p and HAPE-r groups. Information about physiological phenotypes was obtained via fieldwork investigation. The ACE-I/D polymorphism in HAPE-p and HAPE-r was detected by polymerase chain reaction （PCR）. Results The SaO2 was significantly lower while HR was significantly higher in HAPE-p group than those in HAPE-r group. The genotype frequencies of ACE-I/D for II, ID, DD in HAPE-r and HAPE-p groups were 0.430, 0.446, 0.124 and 0.435, 0.469, 0.095, respectively, the allelic frequencies of I and D were 0.650, 0.350 and 0.670, 0.330, respectively. The OR of ID, DD and D alleles relative to II for HAPE was 0.961 （0.610-1.514）, 1.322 （0.634-2.758） and 1.080 （0.783-1.489）. There was no significant difference of the genotypic and the allelic frequencies in ACE-I/D polymorphism between HAPE-p and HAPE-r groups. Conclusions There is no relation between ACE-I/D polymorphism and HAPE in the Han Chinese.     

Interactive effects of pests increase seed yield

Loss in seed yield and therefore decrease in plant fitness due to simultaneous attacks by multiple herbivores is not necessarily additive, as demonstrated in evolutionary studies on wild plants. However, it is not clear how this transfers to crop plants that grow in very different conditions compared to wild plants. Nevertheless, loss in crop seed yield caused by any single pest is most often studied in isolation although crop plants are attacked by many pests that can cause substantial yield losses. This is especially important for crops able to compensate and even overcompensate for the damage. We investigated the interactive impacts on crop yield of four insect pests attacking different plant parts at different times during the cropping season. In 15 oilseed rape fields in Sweden, we estimated the damage caused by seed and stem weevils, pollen beetles, and pod midges. Pest pressure varied drastically among fields with very low correlation among pests, allowing us to explore interactive impacts on yield from attacks by multiple species. The plant damage caused by each pest species individually had, as expected, either no, or a negative impact on seed yield and the strongest negative effect was caused by pollen beetles. However, seed yield increased when plant damage caused by both seed and stem weevils was high, presumably due to the joint plant compensatory reaction to insect attack leading to overcompensation. Hence, attacks by several pests can change the impact on yield of individual pest species. Economic thresholds based on single species, on which pest management decisions currently rely, may therefore result in economically suboptimal choices being made and unnecessary excessive use of insecticides.