Can transposable element copy number distribution parameters be estimated from natural populations of Drosophila melanogaster?

The copy frequency distribution of a transposable element family in a Drosophila melanogaster natural population is generally characterised by the values of the Charlesworths' model parameters α and β (Charlesworth & Charlesworth, 1983). The estimation of these parameters is made using the observed distribution of the occupied sites in a population sample. Several results have been interpreted as due either to the influence of stochastic factors or to deterministic factors (transposition, excision, selection…). The accuracy of this method was tested by estimations performed on samples from simulated populations. The results show that with the sample size usually used for natural population studies, the confidence intervals are too large to reasonably deduce either the element copy number distribution or the values of transposition and excision rate and selective coefficients.     

Specificity of the interaction of furfural with DNA

Furfural or 2-furaldehyde is a dietary mutagen and is present in various frequently consumed food products. The alkaline unwinding assay and protection of cleavage sites from the action of various restriction enzymes was used to study the interaction of furfural with DNA. Alkaline unwinding experiments showed the formation of an increasing number of strand breaks in duplex DNA both with increasing furfural concentration and with time of reaction. Treatment of lambda phage DNA with furfural protected cleavage with restriction endonucleases DraI and SspI but not with ApaI, BssHII and SacII. These results indicate that under the conditions used furfural reacts exclusively with AT base pairs. A minimum of 3-4 consecutive AT base pairs are required for this reaction. This was determined by the use of several restriction enzymes whose hexanucleotide recognition sequences contain subsets of AT base pairs.     

Somaclonal variation as a tool to develop pest resistant plants of Torenia fournieri ‘Compacta Blue’

Callus cultures of Torenia fournieri Compacta Blue were initiated on a modified Murashige and Skoog salt medium (MS) with 2.26 M 2,4-dichlorophenoxyacetic acid. Shoots were regenerated from these cultures using MS medium amended with 2.46 M indolebutyric acid and 8.88 M benzyladenine. These shoot cultures were subjected to two-spotted spidermite (Tetranychus urticae Koch.) and the greenhouse whitefly [Trialeurodes vaporariorum (Westwood)]. Pests were allowed to feed until such time that their populations started to decrease due to lack of food. The remaining live tissue of the Torenia was placed on MS medium amended with 2.28 M zeatin to induce new adventitious shoots and plantlets. Newly regenerated plantlets were acclimated to greenhouse conditions and evaluated for resistance to the pest to which they were subjected in vitro. Highly significant differences in pest numbers were found in somaclones for both the two-spotted spidermite and greenhouse whitefly when compared to control plants. A wide range of variability was observed among the somaclonal population. There were significantly fewer mite eggs laid on plants regenerated from in vitro cultures screened with two-spotted spidermites than on seed sown controls. Regenerants from cultures screened with whiteflies in vitro had fewer eggs, immatures and live adults than controls.Abbreviations BA benzyladenine - IBA indolebutyric acid - 2,4-d 2,4-dichlorophenoxyacetic acid - MS Murashige and Skoog salt medium Storrs Agricultural Research Station Scientific Publication 1641.     

The plant glycosyltransferase clone collection for functional genomics

The glycosyltransferases (GTs) are an important and functionally diverse family of enzymes involved in glycan and glycoside biosynthesis. Plants have evolved large families of GTs which undertake the array of glycosylation reactions that occur during plant development and growth. Based on the Carbohydrate‐Active enZymes (CAZy) database, the genome of the reference plant Arabidopsis thaliana codes for over 450 GTs, while the rice genome (Oryza sativa) contains over 600 members. Collectively, GTs from these reference plants can be classified into over 40 distinct GT families. Although these enzymes are involved in many important plant specific processes such as cell‐wall and secondary metabolite biosynthesis, few have been functionally characterized. We have sought to develop a plant GTs clone resource that will enable functional genomic approaches to be undertaken by the plant research community. In total, 403 (88%) of CAZy defined Arabidopsis GTs have been cloned, while 96 (15%) of the GTs coded by rice have been cloned. The collection resulted in the update of a number of Arabidopsis GT gene models. The clones represent full‐length coding sequences without termination codons and are Gateway^® compatible. To demonstrate the utility of this JBEI GT Collection, a set of efficient particle bombardment plasmids (pBullet) was also constructed with markers for the endomembrane. The utility of the pBullet collection was demonstrated by localizing all members of the Arabidopsis GT14 family to the Golgi apparatus or the endoplasmic reticulum (ER). Updates to these resources are available at the JBEI GT Collection website http://www.addgene.org/ .     

Effect of High-Intensity Interval Training on Cardiac Apoptosis Markers in Methamphetamine-Dependent Rats

Chronic methamphetamine use increases apoptosis, leading to heart failure and sudden cardiac death. Previous studies have shown the importance of high-intensity interval training (HIIT) in reducing indices of cardiac tissue apoptosis in different patients, but in the field of sports science, the molecular mechanisms of apoptosis in methamphetamine-dependent rats are still unclear. The present article aimed to investigate the changes in cardiac apoptosis markers in methamphetamine-dependent rats in response to HIIT. Left ventricular tissue was used to evaluate caspase-3, melusin, FAK, and IQGAP1 gene expression. Rats were divided into four groups: sham, methamphetamine (METH), METH-control, and METH-HIIT. METH was injected for 21 days and then the METH-HIIT group performed HIIT for 8 weeks at 5 sessions per week. The METH groups showed increased caspase-3 gene expression and decreased melusin, FAK, and IQGAP1 when compared to the sham group. METH-HIIT showed decreased caspase-3 and increased melusin and FAK gene expression compared with the METH and METH-control groups. The IQGAP1 gene was higher in METH-HIIT when compared with METH, while no difference was observed between METH-HIIT and METH-control. Twenty-one days of METH exposure increased apoptosis markers in rat cardiac tissue; however, HIIT might have a protective effect, as shown by the apoptosis markers.     

Human Papillomavirus Type 16 Integration Analysis by Real-time PCR Assay in Associated Cancers

Human papillomavirus (HPV) is a common viral infection worldwide associated with a variety of cancers. The integration of the HPV genome in these patients causes chromosomal instability and triggers carcinogenesis. The aim of this study was to investigate the HPV-16 genome physical status in four major cancers related to HPV infection. Formalin-fixed paraffin-embedded blocks from our previous projects on head and neck, colorectal, penile, and cervical cancers were collected, and HPV-16–positive specimens were used for further analysis. The DNA extraction copy number of E2 and E7 genes was calculated by qualitative real-time PCR method. Serially diluted standards that were cloned in PUC57 plasmid were used. Standard curve and melting curve analysis was used for quantification. Of the 672 specimens studied, 76 (11.3%) were HPV-16 positive. We found that 35.6% (16/45) were integrated. Statistical analysis showed that there were significant correlations between integration of HPV-16 and cervical cancer end-stage carcinogenesis (P < .0001), episomal form, and ASCUS lesions (P = .045). Significant correlation in penile cancer patients was seen between the episomal form and high-grade cancer stage (P = .037). Integration is a major factor in the carcinogenesis mechanism of HPV and has different prevalence in various cancers with a higher rate in progression except in penile cancer.     

Functional characterization of Ape1 variants identified in the human population

Apurinic/apyrimidinic (AP) sites are common mutagenic and cytotoxic DNA lesions. Ape1 is the major human repair enzyme for abasic sites and incises the phosphodiester backbone 5′ to the lesion to initiate a cascade of events aimed at removing the AP moiety and maintaining genetic integrity. Through resequencing of genomic DNA from 128 unrelated individuals, and searching published reports and sequence databases, seven amino acid substitution variants were identified in the repair domain of human Ape1. Functional characterization revealed that three of the variants, L104R, E126D and R237A, exhibited ~40–60% reductions in specific incision activity. A fourth variant, D283G, is similar to the previously characterized mutant D283A found to exhibit ~10% repair capacity. The most common substitution (D148E; observed at an allele frequency of 0.38) had no impact on endonuclease and DNA binding activities, nor did a G306A substitution. A G241R variant showed slightly enhanced endonuclease activity relative to wild-type. In total, four of seven substitutions in the repair domain of Ape1 imparted reduced function. These reduced function variants may represent low penetrance human polymorphisms that associate with increased disease susceptibility.     

The prevalence of factor V Leiden,prothrombin G20210A and methylenetetrahydrofolate reductase polymorphism C677T among G6PD deficient individuals from Western Iran

It has been suggested that the allele frequency of thrombophilic mutations is affected by glucose-6-phosphate dehydrogenase (G6PD) deficiency. The prevalence of thrombophilic mutations were studied in sixty G6PD deficient individuals including 57 males and three females with the mean age of 15 ± 3.08 and 110 age and sex matched healthy individuals consisted of 95 males and 15 females with the mean age of 16.19 ± 2.17 from the Kermanshah Province of Iran. Using a combination of PCR-RFLP technique, single strand conformation polymorphism (SSCP) analysis and DNA sequencing polymorphic G6PD mutations were identified. The factor V Leiden, prothrombin G20210A and methylenetetrahydrofolate reductase (MTHFR) C677T were detected by PCR-RFLP method using MnlI, HindIII and HinfI restriction enzymes, respectively. Three mutations, G6PD Mediterranean, G6PD Chatham and G6PD Cosenza were identified in 60 G6PD deficient individuals with highest prevalence of G6PD Mediterranean (91.6%). In G6PD deficient individuals the prevalence of factor V Leiden tended to be higher (5%) compared to healthy individuals (2.7%). The prevalence of prothrombin G20210A mutation in G6PD deficient individuals was 1.7%. However, in normal subjects the prevalence of this mutation was 2.7%. The frequency of T allele in G6PD deficient individuals were insignificantly higher (29.16%) than those in healthy individuals (26.8%). Our finding indicates that the prevalence of factor V Leiden, prothrombin G20210A and MTHFR C677T in G6PD deficient individuals is not statistically different compared to normal subjects and G6PD deficiency is not associated with these thrombophilic mutations in Western Iran.     

The calcium-free form of atorvastatin inhibits amyloid-β(1–42) aggregation in vitro

Alzheimer''s disease is characterized by the presence of extraneuronal amyloid plaques composed of amyloid-beta (Aβ) fibrillar aggregates in the brains of patients. In mouse models, it has previously been shown that atorvastatin (Ator), a cholesterol-lowering drug, has some reducing effect on the production of cerebral Aβ. A meta-analysis on humans showed moderate effects in the short term but no improvement in the Alzheimer''s Disease Assessment Scale—Cognitive Subscale behavioral test. Here, we explore a potential direct effect of Ator on Aβ42 aggregation. Using NMR-based monomer consumption assays and CD spectroscopy, we observed a promoting effect of Ator in its original form (Ator-calcium) on Aβ42 aggregation, as expected because of the presence of calcium ions. The effect was reversed when applying a CaCO₃-based calcium ion scavenging method, which was validated by the aforementioned methods as well as thioflavin-T fluorescence assays and transmission electron microscopy. We found that the aggregation was inhibited significantly when the concentration of calcium-free Ator exceeded that of Aβ by at least a factor of 2. The ¹H–¹⁵N heteronuclear single quantum correlation and saturation-transfer difference NMR data suggest that calcium-free Ator exerts its effect through interaction with the ¹⁶KLVF¹⁹ binding site on the Aβ peptide via its aromatic rings as well as hydroxyl and methyl groups. On the other hand, molecular dynamics simulations confirmed that the increasing concentration of Ator is necessary for the inhibition of the conformational transition of Aβ from an α-helix-dominant to a β-sheet-dominant structure.     

Identification of an Immunoglobulin M (IgM) Antibody Against Enolase 1 Protein (ENO1) Derived from HEK-293 Cells in Patients with Kidney Failure

Kidney failure is one of the most important challenges in medicine. In this study, we used HEK-293 kidney cells to evaluate and compare changes in the expr