Plucked hair follicles from patients with chronic discoid lupus erythematosus show a disease-specific molecular signature

Objective: When faced with clinical symptoms of scarring alopecia—the standard diagnostic pathway involves a scalp biopsy which is an invasive and expensive procedure. This project aimed to assess if plucked hair follicles (HFs) containing living epithelial cells can offer a non-invasive approach to diagnosing inflammatory scalp lesions. Methods: Lesional and non-lesional HFs were extracted from the scalp of patients with chronic discoid lupus erythematosus (CDLE), psoriasis and healthy controls. RNA was isolated from plucked anagen HFs and microarray, as well as quantitative real-time PCR was performed. Results: Here, we report that gene expression analysis of only a small number of HF plucked from lesional areas of the scalp is sufficient to differentiate CDLE from psoriasis lesions or healthy HF. The expression profile from CDLE HFs coincides with published profiles of CDLE from skin biopsy. Genes that were highly expressed in lesional CDLE corresponded to well-known histopathological diagnostic features of CDLE and included those related to apoptotic cell death, the interferon signature, complement components and CD8+ T-cell immune responses. Conclusions: We therefore propose that information obtained from this non-invasive approach are sufficient to diagnose scalp lupus erythematosus. Once validated in routine clinical settings and compared with other scarring alopecias, this rapid and non-invasive approach will have great potential for paving the way for future diagnosis of inflammatory scalp lesions

Control of harmonic generation by the time delay between two-color, bicircular few-cycle mid-IR laser pulses

We study control of high-order harmonic generation (HHG) driven by time-delayed, few-cycle ω and 2ω counterrotating mid-IR pulses. Our numerical and analytical study shows that the time delay between the two-color pulses allows control of the harmonic positions, both those allowed by angular momentum conservation and those seemingly forbidden by it. Moreover, the helicity of any particular harmonic is tunable from left to right circular without changing the driving pulse helicity. The highest HHG yield occurs for a time delay comparable to the fundamental period T=2π/ω

Potentials of Mean Force for Protein Structure Prediction Vindicated, Formalized and Generalized

Understanding protein structure is of crucial importance in science, medicine and biotechnology. For about two decades, knowledge based potentials based on pairwise distances -- so-called "potentials of mean force" (PMFs) -- have been center stage in the prediction and design of protein structure and the simulation of protein folding. However, the validity, scope and limitations of these potentials are still vigorously debated and disputed, and the optimal choice of the reference state -- a necessary component of these potentials -- is an unsolved problem. PMFs are loosely justified by analogy to the reversible work theorem in statistical physics, or by a statistical argument based on a likelihood function. Both justifications are insightful but leave many questions unanswered. Here, we show for the first time that PMFs can be seen as approximations to quantities that do have a rigorous probabilistic justification: they naturally arise when probability distributions over different features of proteins need to be combined. We call these quantities reference ratio distributions deriving from the application of the reference ratio method. This new view is not only of theoretical relevance, but leads to many insights that are of direct practical use: the reference state is uniquely defined and does not require external physical insights; the approach can be generalized beyond pairwise distances to arbitrary features of protein structure; and it becomes clear for which purposes the use of these quantities is justified. We illustrate these insights with two applications, involving the radius of gyration and hydrogen bonding. In the latter case, we also show how the reference ratio method can be iteratively applied to sculpt an energy funnel. Our results considerably increase the understanding and scope of energy functions derived from known biomolecular structures

Catalytic Function of PLA2G6 Is Impaired by Mutations Associated with Infantile Neuroaxonal Dystrophy but Not Dystonia-Parkinsonism

Mutations in the PLA2G6 gene have been identified in autosomal recessive neurodegenerative diseases classified as infantile neuroaxonal dystrophy (INAD), neurodegeneration with brain iron accumulation (NBIA), and dystonia-parkinsonism. These clinical syndromes display two significantly different disease phenotypes. NBIA and INAD are very similar, involving widespread neurodegeneration that begins within the first 1-2 years of life. In contrast, patients with dystonia-parkinsonism present with a parkinsonian movement disorder beginning at 15 to 30 years of age. The PLA2G6 gene encodes the PLA2G6 enzyme, also known as group VIA calcium-independent phospholipase A(2), which has previously been shown to hydrolyze the sn-2 acyl chain of phospholipids, generating free fatty acids and lysophospholipids.We produced purified recombinant wildtype (WT) and mutant human PLA2G6 proteins and examined their catalytic function using in vitro assays with radiolabeled lipid substrates. We find that human PLA2G6 enzyme hydrolyzes both phospholipids and lysophospholipids, releasing free fatty acids. Mutations associated with different disease phenotypes have different effects on catalytic activity. Mutations associated with INAD/NBIA cause loss of enzyme activity, with mutant proteins exhibiting less than 20% of the specific activity of WT protein in both lysophospholipase and phospholipase assays. In contrast, mutations associated with dystonia-parkinsonism do not impair catalytic activity, and two mutations produce a significant increase in specific activity for phospholipid but not lysophospholipid substrates.These results indicate that different alterations in PLA2G6 function produce the different disease phenotypes of NBIA/INAD and dystonia-parkinsonism. INAD/NBIA is caused by loss of the ability of PLA2G6 to catalyze fatty acid release from phospholipids, which predicts accumulation of PLA2G6 phospholipid substrates and provides a mechanistic explanation for the accumulation of membranes in neuroaxonal spheroids previously observed in histopathological studies of INAD/NBIA. In contrast, dystonia-parkinsonism mutations do not appear to directly impair catalytic function, but may modify substrate preferences or regulatory mechanisms for PLA2G6

Reprogramming of orientation columns in visual cortex : a domino effect

Abstract : Cortical organization rests upon the fundamental principle that neurons sharing similar properties are co-located. In the visual cortex, neurons are organized into orientation columns. In a column, most neurons respond optimally to the same axis of an oriented edge, that is, the preferred orientation. This orientation selectivity is believed to be absolute in adulthood. However, in a fully mature brain, it has been established that neurons change their selectivity following sensory experience or visual adaptation. Here, we show that after applying an adapter away from the tested cells, neurons whose receptive fields were located remotely from the adapted site also exhibit a novel selectivity in spite of the fact that they were not adapted. These results indicate a robust reconfiguration and remapping of the orientation domains with respect to each other thus removing the possibility of an orientation hole in the new hypercolumn. These data suggest that orientation columns transcend anatomy, and are almost strictly functionally dynamic

Allostery in Its Many Disguises: From Theory to Applications.

Allosteric regulation plays an important role in many biological processes, such as signal transduction, transcriptional regulation, and metabolism. Allostery is rooted in the fundamental physical properties of macromolecular systems, but its underlying mechanisms are still poorly understood. A collection of contributions to a recent interdisciplinary CECAM (Center Européen de Calcul Atomique et Moléculaire) workshop is used here to provide an overview of the progress and remaining limitations in the understanding of the mechanistic foundations of allostery gained from computational and experimental analyses of real protein systems and model systems. The main conceptual frameworks instrumental in driving the field are discussed. We illustrate the role of these frameworks in illuminating molecular mechanisms and explaining cellular processes, and describe some of their promising practical applications in engineering molecular sensors and informing drug design efforts

The Clinical Impact of Copy Number Variants in Inherited Bone Marrow Failure Syndromes

Inherited bone marrow failure syndromes (IBMFSs) comprise a genetically heterogeneous group of diseases with hematopoietic failure and a wide array of physical malformations. Copy number variants (CNVs) were reported in some IBMFSs. It is unclear what impact CNVs play in patients evaluated for a suspected diagnosis of IBMFS. Clinical and genetic data of 323 patients from the Canadian Inherited Marrow Failure Registry from 2001 to 2014, who had a documented genetic work-up, were analyzed. Cases with pathogenic CNVs (at least 1 kilobasepairs) were compared to cases with other mutations. Genotype-phenotype correlations were performed to assess the impact of CNVs. Pathogenic nucleotide-level mutations were found in 157 of 303 tested patients (51.8%). Genome-wide CNV analysis by single nucleotide polymorphism arrays or comparative genomic hybridization arrays revealed pathogenic CNVs in 11 of 67 patients tested (16.4%). In four of these patients, identification of CNV was crucial for establishing the correct diagnosis as their clinical presentation was ambiguous. Eight additional patients were identified to harbor pathogenic CNVs by other methods. Of the 19 patients with pathogenic CNVs, four had compound-heterozygosity of a CNV with a nucleotide-level mutation. Pathogenic CNVs were associated with more extensive non-hematological organ system involvement

New statistical potential for quality assessment of protein models and a survey of energy functions

Abstract Background Scoring functions, such as molecular mechanic forcefields and statistical potentials are fundamentally important tools in protein structure modeling and quality assessment. Results The performances of a number of publicly available scoring functions are compared with a statistical rigor, with an emphasis on knowledge-based potentials. We explored the effect on accuracy of alternative choices for representing interaction center types and other features of scoring functions, such as using information on solvent accessibility, on torsion angles, accounting for secondary structure preferences and side chain orientation. Partially based on the observations made, we present a novel residue based statistical potential, which employs a shuffled reference state definition and takes into account the mutual orientation of residue side chains. Atom- and residue-level statistical potentials and Linux executables to calculate the energy of a given protein proposed in this work can be downloaded from http://www.fiserlab.org/potentials. Conclusions Among the most influential terms we observed a critical role of a proper reference state definition and the benefits of including information about the microenvironment of interaction centers. Molecular mechanical potentials were also tested and found to be over-sensitive to small local imperfections in a structure, requiring unfeasible long energy relaxation before energy scores started to correlate with model quality.</p

Four Distances between Pairs of Amino Acids Provide a Precise Description of their Interaction

The three-dimensional structures of proteins are stabilized by the interactions between amino acid residues. Here we report a method where four distances are calculated between any two side chains to provide an exact spatial definition of their bonds. The data were binned into a four-dimensional grid and compared to a random model, from which the preference for specific four-distances was calculated. A clear relation between the quality of the experimental data and the tightness of the distance distribution was observed, with crystal structure data providing far tighter distance distributions than NMR data. Since the four-distance data have higher information content than classical bond descriptions, we were able to identify many unique inter-residue features not found previously in proteins. For example, we found that the side chains of Arg, Glu, Val and Leu are not symmetrical in respect to the interactions of their head groups. The described method may be developed into a function, which computationally models accurately protein structures

Ablation of the androgen receptor from vascular smooth muscle cells demonstrates a role for testosterone in vascular calcification

Vascular calcification powerfully predicts mortality and morbidity from cardiovascular disease. Men have a greater risk of cardiovascular disease, compared to women of a similar age. These gender disparities suggest an influence of sex hormones. Testosterone is the primary and most well-recognised androgen in men. Therefore, we addressed the hypothesis that exogenous androgen treatment induces vascular calcification. Immunohistochemical analysis revealed expression of androgen receptor (AR) in the calcified media of human femoral artery tissue and calcified human valves. Furthermore, in vitro studies revealed increased phosphate (Pi)-induced mouse vascular smooth muscle cell (VSMC) calcification following either testosterone or dihydrotestosterone (DHT) treatment for 9 days. Testosterone and DHT treatment increased tissue non-specific alkaline phosphatase (Alpl) mRNA expression. Testosterone-induced calcification was blunted in VSMC-specific AR-ablated (SM-ARKO) VSMCs compared to WT. Consistent with these data, SM-ARKO VSMCs showed a reduction in Osterix mRNA expression. However, intriguingly, a counter-intuitive increase in Alpl was observed. These novel data demonstrate that androgens play a role in inducing vascular calcification through the AR. Androgen signalling may represent a novel potential therapeutic target for clinical intervention