Protein stability is measured by denaturation: When solvent conditions are changed (e.g., temperature, denaturant concentration, or pH) the protein population switches between thermodynamic states. ...The resulting denaturation curves have baselines. If the baselines are steep, nonlinear, or incomplete, it becomes difficult to characterize protein denaturation. Baselines arise because the chromophore probing denaturation is sensitive to solvent conditions, or because the thermodynamic states evolve structurally when solvent conditions are changed, or because the barriers are very low (downhill folding). Kinetics can largely eliminate such baselines: Relaxation of chromophores, or within thermodynamic states, is much faster than the transition over activation barriers separating states. This separation of time scales disentangles population switching between states (desired signal) from chromophore or population relaxation within states (baselines). We derive simple formulas to extract unfolding thermodynamics from kinetics. The formulas are tested with model data and with a difficult experimental test case: the apparent two-state folder PI3K SH3 domain. Its melting temperature T(m) can be extracted reliably by our "thermodynamics from kinetics approach," even when conventional fitting is unreliable.
Abstract
Background
Genomic dysregulation is likely to contribute to neuronal dysfunction in prefrontal cortex (PFC) and other brain regions affected in schizophrenia (SCZ), but genome-scale mapping ...of neuronal transcriptomes and epigenomes has not been conducted in larger cohorts. We have shown recently that the genomic landscape of open chromatin-associated histone modifications, including histone H3-acetyl-lysine 27 (H3K27ac), show in neurons significant associations with the genetic risk architecture of schizophrenia.
Methods
We conducted H3K4me3 (H3K27ac) chromatin immunoprecipitation sequencing (ChIP-seq) on fluorescence-activated cell sorted (FACS) nuclei from dorsolateral PFC of 250 postmortem samples from SCZ and 280 matched control brains, generating altogether ~7(11) billion reads representing H3K4me3 (H3K27ac)-tagged nucleosomal DNA from PFC neurons. In addition, genotyping for the entire cohort of 530 brains was performed.
Results
Enrichment of differentially modified regions across schizophrenia cases and controls in calcium and potassium signaling pathways. This finding was replicated in differentially modified regions across cases and controls obtained from independent cohort. In addition to this, enrichment of schizophrenia GWAS loci was observed in differentially modified regions across cases and controls.
Discussion
Our dataset will provide unique insights into non-coding variants associated with neuronal dysfunction in schizophrenia. Our PsychENCODE sponsored resource highlights the critical role of cell-type specific signatures at regulatory and disease-associated non-coding sequences in the human frontal lobe. This is the first histone methylation and acetylation ChIP-seq datasets generated from prefrontal cortex brain region of 250 schizophrenia cases and 280 controls, postmortem human brain samples. As of now, this is the largest, yet unpublished ChIP-seq dataset from schizophrenia and control brains.
Securing the tracheal tube (TT) at a fixed recommended depth of 21/23 cm in female and male patients, respectively, may result in inappropriate placement of the TT in some patients. The aim of the ...present study was to determine the vocal cord-carina distance (VCD) and tracheal length (TL) to ascertain the optimal depth of TT placement during orotracheal intubation in the adult Indian population.
A total of 92 adults undergoing elective surgery under general anaesthesia with orotracheal intubation were studied. Surface anatomy airway measurements were noted. A cuffed TT (female size 7 mm ID and male size 8 mm ID) was inserted with the intubation guide mark at level with the vocal cords (VCs). Fiberoptic bronchoscopy-guided measurements were obtained for VCD, TL, TT tip-carina distance, VC-cricoid distance and lip-carina (L-C) distance.
The mean±SD VCD was 12.82±2.05 and 12.02±1.44 cm, and TL was 10.14±2.04 and 9.37±1.28 cm in male and female patients, respectively. Statistically significant differences were observed between male and female patients in VCD (p=0.033), TL (p=0.032), L-C distance (p<0.001) and lip-TT tip distance (p<0.001); lip-TT tip distance was 19.50±1.39 cm in male patients and 18.17±1.28 cm in female patients. The L-C distance correlated with patient height, weight and neck length. L-C distance=7.214+0.049×Height+0.320×Neck length+0.033×Weight.
We recommend placing the TT with its proximal guide mark at the level of VCs in the Indian population. The 21/23 cm rule for tube placement depth in female and male patients, respectively, cannot be routinely followed in the Indian population.
Post-Traumatic Stress Disorder (PTSD) is a debilitating psychiatric disorder occurring in individuals exposed to trauma. To date, little is known about the genetic aetiology of the disorder, although ...the latest GWAS (carried out by the PGC-PTSD working group) demonstrates that genetic heritability is in line with other psychiatric disorders. PTSD development involves multi-systemic dysregulation in multiple brain regions and diverse peripheral tissues. Some peripheral systems are particularly interesting since epidemiologic evidence suggests that PTSD patients commonly have cardiovascular, metabolic and immune dysregulation.
Transcriptomic Imputation approaches use machine-learning methods to impute gene expression from large genotype data using curated eQTL reference panels. These offer an exciting opportunity to compare gene associations across neurological and peripheral tissues. Here, we apply CommonMind Consortium (CMC) and Genotype-Tissue Expression (GTEx) derived gene expression prediction models to the PGC-PTSD data (9,245 cases/ 24,285 controls). Models included 12 brain regions, five cardiovascular tissues, 2 endocrine tissues, the tibial nerve, adipose tissue and whole blood.
We identified 24 significant gene-tissue associations, of which 5 were in peripheral tissues (adrenal gland, heart atrial appendage, tibial artery, tibial nerve). We stratified analyses according to trauma type (civilian vs. combat trauma), sex, and self-defined ancestry. Our three strongest associations were identified in military cohorts only, which supports the hypothesis that there is substantial genetic heterogeneity between civilian and combat PTSD risk.
We used the PsychENCODE neuronal and non-neuronal reference map for two histone marks associated with transcription and open chromatin (3-trimethyl-lysine 4 (H3K4me3) and H3-acetyl-lysine 27 (H3K27ac) to understand patterns of histone modification among our PTSD-associated genes. Preliminary analyses indicate a significant correlation between PTSD-association statistics and the presence of both histone marks (correlation with neuronal H3K4me3, Pearson R=0.87, p=3.99×10-5). We intend to expand this analysis to include a wider range of histone modification marks.
We will further expand these analyses to identify tissue specific gene clusters and enriched pathways across tissues or in specific tissues. Finally, we will use neuroimaging data and physiological cardiovascular data to functionally validate our results.
Recent progress in understanding the genetic basis of many psychiatric diseases has identified rare and common variants responsible for genetic risk. Integrating epigenomics data from ...disease-relevant cell types and tissues promises to enhance interpretation of these risk variants and the mechanisms by which they confer disease liability. A better understanding of the role of non-coding regulatory DNA and variation in histone modification in mediating the effects of genetic risk variants promises to give insights into the molecular mechanisms underlying psychiatric disease. With this goal in mind, the PsychENCODE Consortium generated a large-scale epigenomics resource for the human brain to serve as a foundation for integrative genomics in psychiatric research.
We performed ChIP-Seq on 17 subjects, 2 brain regions, 2 cell types and 2 histone marks that yielded 136 total ChIP-Seq samples. Of these 129 (n=63 H3K4me3; n=66 H3K27ac) passed quality control. For each combination of brain region, cell type and histone mark, reads from corresponding samples were combined into a consolidated subset of the data. This produced 8 consolidated subsets, corresponding to: ACC-neuronal-H3K4me3, PFC-neuronal-H3K4me3, etc. The entire data set was then processed and analyzed through the annotation, peak calling, covariate and pathway analyses pipeline.
We have generated the largest cell type-specific dataset of histone-modification in the human brain to date across two brain cortical regions, composed of reference maps from 129 samples collected from 17 healthy subjects. These reference maps consist of ChIP-Seq for two histone marks associated with promoters and active enhancers, H3-trimethyl-lysine 4 (H3K4me3) and H3-acetyl-lysine 27 (H3K27ac) from either neurons, or surrounding non-neuronal cells. Analysis of this data has revealed unexplored insights into cell-, brain region- and to a lesser extent, subject- and sex-specific histone methylation and acetylation landscapes shaping the epigenome of the adult human cerebral cortex. We identify multiple loci with robust region-specific differences separating prefrontal from cingulate neurons, and numerous genome loci with extraordinary high inter-individual variability in H3K27ac and H3K4me3. Finally, we found a significant enrichment of risk variants for schizophrenia, educational attainment and depressive symptoms only in neuronal markers, further indicating the importance of conducting cell type-specific epigenome studies.
Risk variants for schizophrenia, depressive symptoms, neuroticism and educational years are significantly enriched in genomic loci marked specifically in neuronal cells. The difference between neuronal and non-neuronal cells is the major axis of variation in histone modification, yet many genomic loci show substantial epigenetic variability across subjects and brain regions. These findings highlight the utility of this NIMH PsychENCODE-sponsored resource to elucidate epigenomic architectures of regulatory and disease-associated non-coding DNA in the human brain.
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