Comparison of Methods for Quantification of Global DNA Methylation in Human Cells and Tissues

DNA methylation is a key epigenetic modification which, in mammals, occurs mainly at CpG dinucleotides. Most of the CpG methylation in the genome is found in repetitive regions, rich in dormant transposons and endogenous retroviruses. Global DNA hypomethylation, which is a common feature of several conditions such as ageing and cancer, can cause the undesirable activation of dormant repeat elements and lead to altered expression of associated genes. DNA hypomethylation can cause genomic instability and may contribute to mutations and chromosomal recombinations. Various approaches for quantification of global DNA methylation are widely used. Several of these approaches measure a surrogate for total genomic methyl cytosine and there is uncertainty about the comparability of these methods. Here we have applied 3 different approaches (luminometric methylation assay, pyrosequencing of the methylation status of the Alu repeat element and of the LINE1 repeat element) for estimating global DNA methylation in the same human cell and tissue samples and have compared these estimates with the “gold standard” of methyl cytosine quantification by HPLC. Next to HPLC, the LINE1 approach shows the smallest variation between samples, followed by Alu. Pearson correlations and Bland-Altman analyses confirmed that global DNA methylation estimates obtained via the LINE1 approach corresponded best with HPLC-based measurements. Although, we did not find compelling evidence that the gold standard measurement by HPLC could be substituted with confidence by any of the surrogate assays for detecting global DNA methylation investigated here, the LINE1 assay seems likely to be an acceptable surrogate in many cases.     

Movements of white-tailed deer in riparian habitat: Implications for infectious diseases

Movements of male white-tailed deer (Odocoileus virginianus) are of great concern with respect to spread of chronic wasting disease (CWD) across landscapes because most yearlings males disperse and adult males have higher prevalence of CWD than do females and younger deer. We radiocollared and monitored 85 male white-tailed deer in the middle Missouri River Valley of eastern Nebraska and western Iowa, USA from 2004 to 2008. Average size (±SE) of fixed-kernel annual home ranges (95%) and core areas (50%) for resident deer were 449 (±32) ha and 99 (±7) ha, respectively. Resident deer exhibited a high-degree of fidelity to their home ranges. Mean overlap between consecutive annual home ranges and core areas was 81% and 74%, respectively. Average dispersal distance was 17.7 ± 4.5 km (range = 3–121 km) for 22 radio-marked and 6 ear-tagged yearlings. Mean spring dispersal distance (25 km) was 150% greater than fall (10 km). Dispersal direction from Desoto National Wildlife Refuge (DNWR) was bimodal on a northwest to southeast axis that followed the Missouri River corridor. Of 22 yearlings that dispersed, 18 (82%) established adult home ranges within the river valley. Dispersal movements of yearling males represent the greatest risk for rapid spread of diseases from infected source populations. Disease management efforts in riparian habitats should target male fawns and yearling males for removal in areas within or immediately adjacent to river corridors. © 2011 The Wildlife Society.     

From transporter to transceptor: signaling from transporters provokes re-evaluation of complex trafficking and regulatory controls: endocytic internalization and intracellular trafficking of nutrient transceptors may, at least in part, be governed by their signaling function

When cells are starved of their substrate, many nutrient transporters are induced. These undergo rapid endocytosis and redirection of their intracellular trafficking when their substrate becomes available again. The discovery that some of these transporters also act as receptors, or transceptors, suggests that at least part of the sophisticated controls governing the trafficking of these proteins has to do with their signaling function rather than with control of transport. In yeast, the general amino acid permease Gap1 mediates signaling to the protein kinase A pathway. Its endocytic internalization and intracellular trafficking are subject to amino acid control. Other nutrient transceptors controlling this signal transduction pathway appear to be subject to similar trafficking regulation. Transporters with complex regulatory control have also been suggested to function as transceptors in other organisms. Hence, precise regulation of intracellular trafficking in nutrient transporters may be related to the need for tight control of nutrient-induced signaling.     

Estimated glomerular filtration rate is a poor predictor of the concentration of middle molecular weight uremic solutes in chronic kidney disease

Background

Uremic solute concentration increases as Glomerular Filtration Rate (GFR) declines. Weak associations were demonstrated between estimated GFR (eGFR) and the concentrations of several small water-soluble and protein-bound uremic solutes (MW<500Da). Since also middle molecular weight proteins have been associated with mortality and cardiovascular damage in Chronic Kidney Disease (CKD), we investigated the association between several eGFR formulae and the concentration of Low Molecular Weight Proteins (LMWP) (MW>500Da).

Materials and Methods

In 95 CKD-patients (CKD-stage 2–5 not on dialysis), associations between different eGFR-formulae (creatinine, CystatinC-based or both) and the natural logarithm of the concentration of several LMWP’s were analyzed: i.e. parathyroid hormone (PTH), Cystatin C (CystC), interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-α), leptin, retinol binding protein (RbP), immunoglobin light chains kappa and lambda (Ig-κ and Ig-λ), beta-2-microglobulin (β₂M), myoglobin and fibroblast growth factor-23 (FGF-23)).

Results

The regression coefficients (R²) between eGFR, based on the CKD-EPI-Crea-CystC-formula as reference, and the examined LMWP’s could be divided into three groups. Most of the LMWP’s associated weakly (R² <0.2) (FGF-23, leptin, IL-6, TNF-α, Ig-κ, Ig-λ) or intermediately (R² 0.2–0.7) (RbP, myoglobin, PTH). Only β₂M and CystC showed a strong association (R² >0.7). Almost identical R²-values were found per LMWP for all eGFR-formulae, with exception of CystC and β₂M which showed weaker associations with creatinine-based than with CystC-based eGFR.

Conclusion

The association between eGFR and the concentration of several LMWP’s is inconsistent, with in general low R²-values. Thus, the use of eGFR to evaluate kidney function does not reflect the concentration of several LMWP’s with proven toxic impact in CKD.     

A comparative study of seed yield parameters in Arabidopsis thaliana mutants and transgenics

Because seed yield is the major factor determining the commercial success of grain crop cultivars, there is a large interest to obtain more understanding of the genetic factors underlying this trait. Despite many studies, mainly in the model plant Arabidopsis thaliana, have reported transgenes and mutants with effects on seed number and/or seed size, knowledge about seed yield parameters remains fragmented. This study investigated the effect of 46 genes, either in gain- and/or loss-of-function situations, with a total of 64 Arabidopsis lines being examined for seed phenotypes such as seed size, seed number per silique, number of inflorescences, number of branches on the main inflorescence and number of siliques. Sixteen of the 46 genes, examined in 14 Arabidopsis lines, were reported earlier to directly affect in seed size and/or seed number or to indirectly affect seed yield by their involvement in biomass production. Other genes involved in vegetative growth, flower or inflorescence development or cell division were hypothesized to potentially affect the final seed size and seed number. Analysis of this comprehensive data set shows that of the 14 lines previously described to be affected in seed size or seed number, only nine showed a comparable effect. Overall, this study provides the community with a useful resource for identifying genes with effects on seed yield and candidate genes underlying seed QTL. In addition, this study highlights the need for more thorough analysis of genes affecting seed yield.     

Treatment-associated polymorphisms in protease are significantly associated with higher viral load and lower CD4 count in newly diagnosed drug-naive HIV-1 infected patients

Background

Bone marrow stromal cell antigen 2 (BST-2) is a cellular factor that restricts the egress of viruses such as human immunodeficiency virus (HIV-1) from the surface of infected cells, preventing infection of new cells. BST-2 is variably expressed in most cell types, and its expression is enhanced by cytokines such as type I interferon alpha (IFN-??). In this present study, we used the beta-retrovirus, mouse mammary tumor virus (MMTV) as a model to examine the role of mouse BST-2 in host infection in vivo.

Results

By using RNA interference, we show that loss of BST-2 enhances MMTV replication in cultured mammary tumor cells and in vivo. In cultured cells, BST-2 inhibits virus accumulation in the culture medium, and co-localizes at the cell surface with virus structural proteins. Furthermore, both scanning electron micrograph (SEM) and transmission electron micrograph (TEM) show that MMTV accumulates on the surface of IFN??-stimulated cells.

Conclusions

Our data provide evidence that BST-2 restricts MMTV release from naturally infected cells and that BST-2 is an antiviral factor in vivo.     

Evaluation of fences for containing feral swine under simulated depopulation conditions

Populations of feral swine (Sus scrofa) are estimated to include >2 million animals in the state of Texas, USA, alone. Feral swine damage to property, crops, and livestock exceeds $50 million annually. These figures do not include the increased risks and costs associated with the potential for feral swine to spread disease to domestic livestock. Thus, effective bio-security measures will be needed to quickly isolate affected feral swine populations during disease outbreaks. We evaluated enclosures built of 0.86-m-tall traditional hog panels for containing feral swine during 35 trials, each involving 6 recently caught animals exposed to increasing levels of motivation. During trials, fences were 97% successful when enclosures were entered by humans for maintenance purposes; 83% effective when pursued by walking humans discharging paintball projectors; and in limited testing, 100% successful when pursued and removed by gunners in a helicopter. In addition to being effective in containing feral swine, enclosures constructed of hog panels required simple hand tools, took <5 min/m to erect, and were inexpensive ($5.73/m excluding labor) relative to other fencing options. As such, hog-panel fences are suitable for use by state and federal agencies for rapid deployment in disease response situations, but also exhibit utility for general control of other types of damage associated with feral swine. © 2011 The Wildlife Society.