Quantification of mitochondrial DNA copy number: Pre-analytical factors

Mitochondrial DNA (mtDNA) content is important for understanding many cellular processes. Several pre-analytical factors, from sample collection to DNA extraction can affect measurement of mtDNA copy number. In the present study, whole blood samples yielded a higher mtDNA copy number than buffy coat samples. mtDNA content is affected by the cell separation method used and the time between blood withdrawal and cell separation. Thus, reference values must be established with the same type of sample. As to the DNA isolation and purification method, the manual phenol method can give randomly false high values. The QIAamp DNA Mini Kit provided the most highly reproducible mtDNA/nDNA yield.     

The Evolutionary History of the Transposable Element Penelope in the Drosophila virilis Group of Species

We have used phylogenetic techniques to study the evolutionary history of the Penelope transposable element in the Drosophila virilis species group. Two divergent types of Penelope have been detected, one previously described, clade I, and a new one which we have termed clade III. The phylogeny of some copies of the Penelope clade I element was partially consistent with the species phylogeny of the D. montana subphylad, suggesting cospeciation and allowing the estimation of the evolutionary rate of Penelope. Divergence times of elements found in different species are younger than the age of the species, suggesting horizontal transfer events. Electronic Supplementary Material Electronic Supplementary material is available for this article at and accessible for authorised users. [Reviewing Editor: Dr. Dmitri Petrov]     

Genome-Wide Divergence of DNA Methylation Marks in Cerebral and Cerebellar Cortices

Background

Emerging evidence suggests that DNA methylation plays an expansive role in the central nervous system (CNS). Large-scale whole genome DNA methylation profiling of the normal human brain offers tremendous potential in understanding the role of DNA methylation in brain development and function.

Methodology/Significant Findings

Using methylation-sensitive SNP chip analysis (MSNP), we performed whole genome DNA methylation profiling of the prefrontal, occipital, and temporal regions of cerebral cortex, as well as cerebellum. These data provide an unbiased representation of CpG sites comprising 377,509 CpG dinucleotides within both the genic and intergenic euchromatic region of the genome. Our large-scale genome DNA methylation profiling reveals that the prefrontal, occipital, and temporal regions of the cerebral cortex compared to cerebellum have markedly different DNA methylation signatures, with the cerebral cortex being hypermethylated and cerebellum being hypomethylated. Such differences were observed in distinct genomic regions, including genes involved in CNS function. The MSNP data were validated for a subset of these genes, by performing bisulfite cloning and sequencing and confirming that prefrontal, occipital, and temporal cortices are significantly more methylated as compared to the cerebellum.

Conclusions

These findings are consistent with known developmental differences in nucleosome repeat lengths in cerebral and cerebellar cortices, with cerebrum exhibiting shorter repeat lengths than cerebellum. Our observed differences in DNA methylation profiles in these regions underscores the potential role of DNA methylation in chromatin structure and organization in CNS, reflecting functional specialization within cortical regions.     

A Novel Analytical Framework for Dissecting the Genetic Architecture of Behavioral Symptoms in Neuropsychiatric Disorders

Background

For diagnosis of neuropsychiatric disorders, a categorical classification system is often utilized as a simple way for conceptualizing an often complex clinical picture. This approach provides an unsatisfactory model of mental illness, since in practice patients do not conform to these prototypical diagnostic categories. Family studies show notable familial co-aggregation between schizophrenia and bipolar illness and between schizoaffective disorders and both bipolar disorder and schizophrenia, revealing that mental illness does not conform to such categorical models and is likely to follow a continuum encompassing a spectrum of behavioral symptoms.

Results and Methodology

We introduce an analytic framework to dissect the phenotypic heterogeneity present in complex psychiatric disorders based on the conceptual paradigm of a continuum of psychosis. The approach identifies subgroups of behavioral symptoms that are likely to be phenotypically and genetically homogenous. We have evaluated this approach through analysis of simulated data with simulated behavioral traits and predisposing genetic factors. We also apply this approach to a psychiatric dataset of a genome scan for schizophrenia for which extensive behavioral information was collected for each individual patient and their families. With this approach, we identified significant evidence for linkage among depressed individuals with two distinct symptom profiles, that is individuals with sleep disturbance symptoms with linkage on chromosome 2q13 and also a mutually exclusive group of individuals with symptoms of concentration problems with linkage on chromosome 2q35. In addition we identified a subset of individuals with schizophrenia defined by language disturbances with linkage to chromosome 2p25.1 and a group of patients with a phenotype intermediate between those of schizophrenia and schizoaffective disorder with linkage to chromosome 2p21.

Conclusions

The findings presented are novel and demonstrate the efficacy of this approach in detection of genes underlying such complex human disorders as schizophrenia and depression.     

Fire frequency effects on soil and pollinators: what shapes sexual plant reproduction?

Increased fire frequency can significantly erode both soil properties and plant–pollinator interactions affecting plant reproductive success but they have seldom been assessed simultaneously. Here, we evaluate soil properties, pollinator assemblage and the reproductive success of two native Fabaceae herbs, Desmodium uncinatum and Rhynchosia edulis, growing in unburned, low and high fire frequency sites of Chaco Serrano across two consecutive years. Desmodium uncinatum is outcrossing with a high dependence on pollinators, whereas R. edulis is autogamous and completely independent of pollinators. We found that soil water content, nitrates and electrical conductivity significantly decreased in low and high fire frequency sites. Pollinator richness and composition visiting each plant species was similar across all fire frequency scenarios. However, fruit set of the exogamous D. uncinatum was strongly reduced in frequently burned sites, whereas fruit set of the autogamous R. edulis showed no significant changes. In both species, the probability of setting fruits was positively related to soil quality across fire frequency scenarios, implying that decreased reproduction was mainly driven by limitation of abiotic resources shaped by increased fire frequency. Because the pollinator-dependent D. uncinatum has a higher reproductive cost, reduced soil quality induced by fire frequency had stronger effects on its reproduction. Chronic reduction of sexual reproduction in frequently burned sites with depleted soils will limit population recruitment with negative consequences on long-term plant population persistence.     

New bis(azolylcarbonyl)pyridine chromium(III) complexes as initiators for ethylene polymerization

Reaction of 2,6-pyridinedicarbonyl dichloride with 3,5-dimethylpyrazole and 1H-indazole, respectively, yield the tridentate ligands 2,6-bis(3,5-dimethylpyrazol-1-ylcarbonyl)pyridine (1) and 2,6-bis(indazol-1-ylcarbonyl)pyridine (2). The molecular structure of the new compound (2) was determined by single-crystal X-ray diffraction. These ligands react with CrCl₃(THF)₃ in THF to form neutral complexes of general formula [CrCl₃{2,6-bis(azolylcarbonyl)pyridine-N,N,N}] (3, 4) which were isolated in high yield as air stable green solids and characterized by elemental analysis, magnetic moment, IR, and mass spectroscopies. Theoretical calculations predict that the thermodynamically preferred structure of the complexes is the fac configuration. After reaction with methylaluminoxane (MAO) the chromium(III) complexes are active for the polymerization of ethylene.     

Altitudinal and climatic adaptation is mediated by flowering traits and FRI, FLC, and PHYC genes in Arabidopsis

Extensive natural variation has been described for the timing of flowering initiation in many annual plants, including the model wild species Arabidopsis (Arabidopsis thaliana), which is presumed to be involved in adaptation to different climates. However, the environmental factors that might shape this genetic variation, as well as the molecular bases of climatic adaptation by modifications of flowering time, remain mostly unknown. To approach both goals, we characterized the flowering behavior in relation to vernalization of 182 Arabidopsis wild genotypes collected in a native region spanning a broad climatic range. Phenotype-environment association analyses identified strong altitudinal clines (0-2600 m) in seven out of nine flowering-related traits. Altitudinal clines were dissected in terms of minimum winter temperature and precipitation, indicating that these are the main climatic factors that might act as selective pressures on flowering traits. In addition, we used an association analysis approach with four candidate genes, FRIGIDA (FRI), FLOWERING LOCUS C (FLC), PHYTOCHROME C (PHYC), and CRYPTOCHROME2, to decipher the genetic bases of this variation. Eleven different loss-of-function FRI alleles of low frequency accounted for up to 16% of the variation for most traits. Furthermore, an FLC allelic series of six novel putative loss- and change-of-function alleles, with low to moderate frequency, revealed that a broader FLC functional diversification might contribute to flowering variation. Finally, environment-genotype association analyses showed that the spatial patterns of FRI, FLC, and PHYC polymorphisms are significantly associated with winter temperatures and spring and winter precipitations, respectively. These results support that allelic variation in these genes is involved in climatic adaptation.     

Melatonin enhances hydrogen peroxide-induced apoptosis in human promyelocytic leukaemia HL-60 cells

Melatonin is an indoleamine secreted by the pineal gland that shows multiple tasks. This ubiquitously acting free radical scavenger has recently been shown to stimulate the production of reactive oxygen species (ROS) in tumour cells, making them undergo apoptosis, whilst it prevents apoptosis in healthy cells. The mechanisms by which melatonin exerts these dual actions are, however, not yet clearly understood. Thus, the aim of this study was to further investigate how melatonin can enhance oxidative stress-induced apoptosis in a leukaemia cell line. The results show that melatonin increased the apoptotic effects of H(2)O(2) in human myeloid HL-60 cells as assessed by cellular viability, mitochondrial permeability transition induction, mitochondrial membrane depolarization, ROS generation, caspases 3, 8 and 9 activity, phosphatidylserine externalization, and DNA fragmentation techniques. When healthy leucocytes were exposed to H(2)O(2), melatonin increased the viability of the cells. Taken together, the findings indicate that melatonin is a potential physiological tool capable of protecting healthy cells from chemotherapy-induced ROS production as well as inducing tumour cell death. Because cancer cells manifest increased oxidative stress as a result of their elevated metabolism, the use of melatonin may be useful in impairing their ROS buffering capacity.