DNA Methylation of the ABO Promoter Underlies Loss of ABO Allelic Expression in a Significant Proportion of Leukemic Patients

Background: Loss of A, B and H antigens from the red blood cells of patients with myeloid malignancies is a frequent occurrence. Previously, we have reported alterations in ABH antigens on the red blood cells of 55% of patients with myeloid malignancies. Methodology/Principal Findings: To determine the underlying molecular mechanisms of this loss, we assessed ABO allelic expression in 21 patients with ABH antigen loss previously identified by flow cytometric analysis as well as an additional 7 patients detected with ABH antigen changes by serology. When assessing ABO mRNA allelic expression, 6/12 (50%) patients with ABH antigen loss detected by flow cytometry and 5/7 (71%) of the patients with ABH antigen loss detected by serology had a corresponding ABO mRNA allelic loss of expression. We examined the ABO locus for copy number and DNA methylation alterations in 21 patients, 11 with loss of expression of one or both ABO alleles, and 10 patients with no detectable allelic loss of ABO mRNA expression. No loss of heterozygosity (LOH) at the ABO locus was observed in these patients. However in 8/11 (73%) patients with loss of ABO allelic expression, the ABO promoter was methylated compared with 2/10 (20%) of patients with no ABO allelic expression loss (P = 0.03). Conclusions/Significance: We have found that loss of ABH antigens in patients with hematological malignancies is associated with a corresponding loss of ABO allelic expression in a significant proportion of patients. Loss of ABO allelic expression was strongly associated with DNA methylation of the ABO promoter.Tina Bianco-Miotto, Damian J. Hussey, Tanya K. Day, Denise S. O'Keefe and Alexander Dobrovi

Methyl jasmonate-elicited herbivore resistance: does MeJA function as a signal without being hydrolyzed to JA?

Treatment with methyl jasmonate (MeJA) elicits herbivore resistance in many plant species and over-expression of JA carboxyl methyltransferase (JMT) constitutively increases JA-induced responses in Arabidopsis. When wild-type (WT) Nicotiana attenuata plants are treated with MeJA, a rapid transient endogenous JA burst is elicited, which in turn increases levels of nicotine and trypsin proteinase inhibitors (TPIs) and resistance to larvae of the specialist herbivore, Manduca sexta. All of these responses are impaired in plants silenced in lipoxygenase 3 expression (asLOX3) but are restored to WT levels by MeJA treatment. Whether these MeJA-induced responses are directly elicited by MeJA or by its cleavage product, JA, is unknown. Using virus-induced gene silencing (VIGS), we silenced MeJA-esterase (NaMJE) expression and found this gene responsible for most of the MeJA-cleaving activity in N. attenuata protein extracts. Silencing NaMJE in asLOX3, but not in WT plants, significantly reduced MeJA-induced nicotine levels and resistance to M. sexta, but not TPI levels. MeJA-induced transcript levels of threonine deaminase (NaTD) and phenylalanine ammonia lyase (NaPAL1) were also decreased in VIGS MJE (asLOX3) plants. Finally the performance of M. sexta larvae that fed on plants treated with JA or MeJA demonstrated that silencing NaMJE inhibited MeJA-induced but not JA-induced resistance in asLOX3 plants. From these results, we conclude that the resistance elicited by MeJA treatment is directly elicited not by MeJA but by its de-methylated product, JA

KRAS-G12C Mutation in One Real-Life and Three Population-Based Nordic Cohorts of Metastatic Colorectal Cancer

Background: KRAS mutations, present in over 40% of metastatic colorectal cancer (mCRC), are negative predictive factors for anti-EGFR therapy. Mutations in KRAS-G12C have a cysteine residue for which drugs have been developed. Published data on this specific mutation are conflicting; thus, we studied the frequency and clinical characteristics in a real-world and population-based setting.Methods: Patients from three Nordic population-based cohorts and the real-life RAXO-study were combined. RAS and BRAF tests were performed in routine healthcare, except for one cohort. The dataset consisted of 2,559 patients, of which 1,871 could be accurately classified as KRAS, NRAS, and BRAF-V600E. Demographics, treatments, and outcomes were compared using logistic regression. Overall survival (OS) was estimated with Kaplan-Meier, and differences were compared using Cox regression, adjusted for baseline factors.Results: The KRAS-G12C frequency was 2%-4% of all tested in the seven cohorts (mean 3%) and 4%-8% of KRAS mutated tumors in the cohorts (mean 7%). Metastasectomies and ablations were performed more often (38% vs. 28%, p = 0.040), and bevacizumab was added more often (any line 74% vs. 59%, p = 0.007) for patients with KRAS-G12C- vs. other KRAS-mutated tumors, whereas chemotherapy was given to similar proportions. OS did not differ according to KRAS mutation, neither overall (adjusted hazard ratio (HR) 1.03; 95% CI 0.74-1.42, reference KRAS-G12C) nor within treatment groups defined as "systemic chemotherapy, alone or with biologics", "metastasectomy and/or ablations", or "best supportive care", RAS and BRAF wild-type tumors (n = 548) differed similarly to KRAS-G12C, as to other KRAS- or NRAS-mutated (n = 66) tumors.Conclusions: In these real-life and population-based cohorts, there were no significant differences in patient characteristics and outcomes between patients with KRAS-G12C tumors and those with other KRAS mutations. This contrasts with the results of most previous studies claiming differences in many aspects, often with worse outcomes for those with a KRAS-G12C mutation, although not consistent. When specific drugs are developed, as for this mutation, differences in outcome will hopefully emerge.ill hopefully emerge.</p

Healthy diet in primary care: views of general practitioners and nurses from Europe

Background: Most of the national colleges of general practitioners (GPs) do not have their own dietary/nutritional tools, and GPs and nurses do not have the time, knowledge, or skills to advise their patients about desirable dietary practices. Objective: To assess the usefulness of a simple and practical guide on healthy diet to be used by European GPs and nurses. Design: A postal survey was mailed to 171 GPs and nurses from 12 European countries to obtain information about the usefulness of a guide on healthy diet developed by EUROPREV. Results: The perception of health professionals is that the main source of information on healthy diet for the population was the media. In all, 95% of GPs and nurses reported that the guide was useful; 93, 95, and 82% reported that the concepts were concise, easy to understand, and realistic, respectively. Also, 77% reported that the type of counselling recommended was feasible and could be applied, 94% reported that the implementation measures proposed could be effective and 88% reported that the Traditional Mediterranean Diet Pyramid is useful, but some concerns about the content were mentioned. Conclusions: GPs and nurses from Europe think that a practical guide on healthy diet developed by EUROPREV could be used to advise patients in primary care, although the Traditional Mediterranean Diet Pyramid should be modified

Reverse Genetics in Ecological Research

By precisely manipulating the expression of individual genetic elements thought to be important for ecological performance, reverse genetics has the potential to revolutionize plant ecology. However, untested concerns about possible side-effects of the transformation technique, caused by Agrobacterium infection and tissue culture, on plant performance have stymied research by requiring onerous sample sizes. We compare 5 independently transformed Nicotiana attenuata lines harboring empty vector control (EVC) T-DNA lacking silencing information with isogenic wild types (WT), and measured a battery of ecologically relevant traits, known to be important in plant-herbivore interactions: phytohormones, secondary metabolites, growth and fitness parameters under stringent competitive conditions, and transcriptional regulation with microarrays. As a positive control, we included a line silenced in trypsin proteinase inhibitor gene (TPI) expression, a potent anti-herbivore defense known to exact fitness costs in its expression, in the analysis. The experiment was conducted twice, with 10 and 20 biological replicates per genotype. For all parameters, we detected no difference between any EVC and WT lines, but could readily detect a fitness benefit of silencing TPI production. A statistical power analyses revealed that the minimum sample sizes required for detecting significant fitness differences between EVC and WT was 2–3 orders of magnitude larger than the 10 replicates required to detect a fitness effect of TPI silencing. We conclude that possible side-effects of transformation are far too low to obfuscate the study of ecologically relevant phenotypes

Accuracy of 1-Hour Plasma Glucose During the Oral Glucose Tolerance Test in Diagnosis of Type 2 Diabetes in Adults : A Meta-analysis

OBJECTIVE One-hour plasma glucose (1-h PG) during the oral glucose tolerance test (OGTT) is an accurate predictor of type 2 diabetes. We performed a meta-analysis to determine the optimum cutoff of 1-h PG for detection of type 2 diabetes using 2-h PG as the gold standard. RESEARCH DESIGN AND METHODS We included 15 studies with 35,551 participants from multiple ethnic groups (53.8% Caucasian) and 2,705 newly detected cases of diabetes based on 2-h PG during OGTT. We excluded cases identified only by elevated fasting plasma glucose and/or HbA(1c). We determined the optimal 1-h PG threshold and its accuracy at this cutoff for detection of diabetes (2-h PG >= 11.1 mmol/L) using a mixed linear effects regression model with different weights to sensitivity/specificity (2/3, 1/2, and 1/3). RESULTS Three cutoffs of 1-h PG, at 10.6 mmol/L, 11.6 mmol/L, and 12.5 mmol/L, had sensitivities of 0.95, 0.92, and 0.87 and specificities of 0.86, 0.91, and 0.94 at weights 2/3, 1/2, and 1/3, respectively. The cutoff of 11.6 mmol/L (95% CI 10.6, 12.6) had a sensitivity of 0.92 (0.87, 0.95), specificity of 0.91 (0.88, 0.93), area under the curve 0.939 (95% confidence region for sensitivity at a given specificity: 0.904, 0.946), and a positive predictive value of 45%. CONCLUSIONS The 1-h PG of >= 11.6 mmol/L during OGTT has a good sensitivity and specificity for detecting type 2 diabetes. Prescreening with a diabetes-specific risk calculator to identify high-risk individuals is suggested to decrease the proportion of false-positive cases. Studies including other ethnic groups and assessing complication risk are warranted.Peer reviewe

Coordinated Multiwavelength Observations of BL Lacertae in 2000

BL Lacertae was the target of an extensive multiwavelength monitoring campaign in the second half of 2000. Simultaneous or quasi-simultaneous observations were taken at radio (UMRAO and Metsaehovi) and optical(WEBT collaboration) frequencies, in X-rays (BeppoSAX and RXTE), and at VHE gamma-rays (HEGRA). The WEBT optical campaign achieved an unprecedented time coverage, virtually continuous over several 10 - 20 hour segments. It revealed intraday variability on time scales of ~ 1.5 hours and evidence for spectral hardening associated with increasing optical flux. During the campaign, BL Lacertae underwent a major transition from a rather quiescent state prior to September 2000, to a flaring state for the rest of the year. This was also evident in the X-ray activity of the source. BeppoSAX observations on July 26/27 revealed a rather low X-ray flux and a hard spectrum, while a BeppoSAX pointing on Oct. 31 - Nov. 2, 2000, indicated significant variability on time scales of < a few hours, and provided evidence for the synchrotron spectrum extending out to ~ 10 keV during that time. During the July 26/27 observation, there is a tantalizing, though not statistically significant, indication of a time delay of ~ 4 - 5 hr between the BeppoSAX and the R-band light curve. Also, a low-significance detection of a time delay of 15 d between the 14.5 GHz and the 22 GHz radio light curves is reported. Several independent methods to estimate the co-moving magnetic field in the source are presented, suggesting a value of ~ 2 e_B^{2/7} G, where e_B is the magnetic-field equipartition factor w.r.t. the electron energy density in the jet.Comment: Accepted for publication in Ap

The Biochemistry, Ultrastructure, and Subunit Assembly Mechanism of AMPA Receptors

The AMPA-type ionotropic glutamate receptors (AMPA-Rs) are tetrameric ligand-gated ion channels that play crucial roles in synaptic transmission and plasticity. Our knowledge about the ultrastructure and subunit assembly mechanisms of intact AMPA-Rs was very limited. However, the new studies using single particle EM and X-ray crystallography are revealing important insights. For example, the tetrameric crystal structure of the GluA2cryst construct provided the atomic view of the intact receptor. In addition, the single particle EM structures of the subunit assembly intermediates revealed the conformational requirement for the dimer-to-tetramer transition during the maturation of AMPA-Rs. These new data in the field provide new models and interpretations. In the brain, the native AMPA-R complexes contain auxiliary subunits that influence subunit assembly, gating, and trafficking of the AMPA-Rs. Understanding the mechanisms of the auxiliary subunits will become increasingly important to precisely describe the function of AMPA-Rs in the brain. The AMPA-R proteomics studies continuously reveal a previously unexpected degree of molecular heterogeneity of the complex. Because the AMPA-Rs are important drug targets for treating various neurological and psychiatric diseases, it is likely that these new native complexes will require detailed mechanistic analysis in the future. The current ultrastructural data on the receptors and the receptor-expressing stable cell lines that were developed during the course of these studies are useful resources for high throughput drug screening and further drug designing. Moreover, we are getting closer to understanding the precise mechanisms of AMPA-R-mediated synaptic plasticity