Binding pocket stabilization by high-throughput screening of yeast display libraries

Protein dynamics have a great influence on the binding pockets of some therapeutic targets. Flexible protein binding sites can result in transient binding pocket formation which might have a negative impact on drug screening efforts. Here, we describe a protein engineering strategy with FK506-binding protein 51 (FKBP51) as a model protein, which is a promising target for stress-related disorders. High-throughput screening of yeast display libraries of FKBP51 resulted in the identification of variants exhibiting higher affinity binding of conformation-specific FKBP51 selective inhibitors. The gene libraries of a random mutagenesis and site saturation mutagenesis of the FK1 domain of FKBP51 encoding sequence were used to create a yeast surface display library. Fluorescence-activated cell sorting for FKBP51 variants that bind conformation-specific fluorescently labeled ligands with high affinity allowed for the identification of 15 different protein variants with improved binding to either, or both FKBP51-specific ligands used in the screening, with improved affinities up to 34-fold compared to the wild type. These variants will pave the way to a better understanding of the conformational flexibility of the FKBP51 binding pocket and may enable the isolation of new selective ligands that preferably and selectively bind the active site of the protein in its open conformation state

Genome-wide association study identifies 30 Loci Associated with Bipolar Disorder

This paper is dedicated to the memory of Psychiatric Genomics Consortium (PGC) founding member and Bipolar disorder working group co-chair Pamela Sklar. We thank the participants who donated their time, experiences and DNA to this research, and to the clinical and scientific teams that worked with them. We are deeply indebted to the investigators who comprise the PGC. The views expressed are those of the authors and not necessarily those of any funding or regulatory body. Analyses were carried out on the NL Genetic Cluster Computer (http://www.geneticcluster.org ) hosted by SURFsara, and the Mount Sinai high performance computing cluster (http://hpc.mssm.edu).Bipolar disorder is a highly heritable psychiatric disorder. We performed a genome-wide association study including 20,352 cases and 31,358 controls of European descent, with follow-up analysis of 822 variants with P<1x10-4 in an additional 9,412 cases and 137,760 controls. Eight of the 19 variants that were genome-wide significant (GWS, p < 5x10-8) in the discovery GWAS were not GWS in the combined analysis, consistent with small effect sizes and limited power but also with genetic heterogeneity. In the combined analysis 30 loci were GWS including 20 novel loci. The significant loci contain genes encoding ion channels, neurotransmitter transporters and synaptic components. Pathway analysis revealed nine significantly enriched gene-sets including regulation of insulin secretion and endocannabinoid signaling. BDI is strongly genetically correlated with schizophrenia, driven by psychosis, whereas BDII is more strongly correlated with major depressive disorder. These findings address key clinical questions and provide potential new biological mechanisms for BD.This work was funded in part by the Brain and Behavior Research Foundation, Stanley Medical Research Institute, University of Michigan, Pritzker Neuropsychiatric Disorders Research Fund L.L.C., Marriot Foundation and the Mayo Clinic Center for Individualized Medicine, the NIMH Intramural Research Program; Canadian Institutes of Health Research; the UK Maudsley NHS Foundation Trust, NIHR, NRS, MRC, Wellcome Trust; European Research Council; German Ministry for Education and Research, German Research Foundation IZKF of Münster, Deutsche Forschungsgemeinschaft, ImmunoSensation, the Dr. Lisa-Oehler Foundation, University of Bonn; the Swiss National Science Foundation; French Foundation FondaMental and ANR; Spanish Ministerio de Economía, CIBERSAM, Industria y Competitividad, European Regional Development Fund (ERDF), Generalitat de Catalunya, EU Horizon 2020 Research and Innovation Programme; BBMRI-NL; South-East Norway Regional Health Authority and Mrs. Throne-Holst; Swedish Research Council, Stockholm County Council, Söderström Foundation; Lundbeck Foundation, Aarhus University; Australia NHMRC, NSW Ministry of Health, Janette M O'Neil and Betty C Lynch

Genomic Relationships, Novel Loci, and Pleiotropic Mechanisms across Eight Psychiatric Disorders

Genetic influences on psychiatric disorders transcend diagnostic boundaries, suggesting substantial pleiotropy of contributing loci. However, the nature and mechanisms of these pleiotropic effects remain unclear. We performed analyses of 232,964 cases and 494,162 controls from genome-wide studies of anorexia nervosa, attention-deficit/hyper-activity disorder, autism spectrum disorder, bipolar disorder, major depression, obsessive-compulsive disorder, schizophrenia, and Tourette syndrome. Genetic correlation analyses revealed a meaningful structure within the eight disorders, identifying three groups of inter-related disorders. Meta-analysis across these eight disorders detected 109 loci associated with at least two psychiatric disorders, including 23 loci with pleiotropic effects on four or more disorders and 11 loci with antagonistic effects on multiple disorders. The pleiotropic loci are located within genes that show heightened expression in the brain throughout the lifespan, beginning prenatally in the second trimester, and play prominent roles in neurodevelopmental processes. These findings have important implications for psychiatric nosology, drug development, and risk prediction.Peer reviewe

Analysis of shared heritability in common disorders of the brain

ience, this issue p. eaap8757 Structured Abstract INTRODUCTION Brain disorders may exhibit shared symptoms and substantial epidemiological comorbidity, inciting debate about their etiologic overlap. However, detailed study of phenotypes with different ages of onset, severity, and presentation poses a considerable challenge. Recently developed heritability methods allow us to accurately measure correlation of genome-wide common variant risk between two phenotypes from pools of different individuals and assess how connected they, or at least their genetic risks, are on the genomic level. We used genome-wide association data for 265,218 patients and 784,643 control participants, as well as 17 phenotypes from a total of 1,191,588 individuals, to quantify the degree of overlap for genetic risk factors of 25 common brain disorders. RATIONALE Over the past century, the classification of brain disorders has evolved to reflect the medical and scientific communities' assessments of the presumed root causes of clinical phenomena such as behavioral change, loss of motor function, or alterations of consciousness. Directly observable phenomena (such as the presence of emboli, protein tangles, or unusual electrical activity patterns) generally define and separate neurological disorders from psychiatric disorders. Understanding the genetic underpinnings and categorical distinctions for brain disorders and related phenotypes may inform the search for their biological mechanisms. RESULTS Common variant risk for psychiatric disorders was shown to correlate significantly, especially among attention deficit hyperactivity disorder (ADHD), bipolar disorder, major depressive disorder (MDD), and schizophrenia. By contrast, neurological disorders appear more distinct from one another and from the psychiatric disorders, except for migraine, which was significantly correlated to ADHD, MDD, and Tourette syndrome. We demonstrate that, in the general population, the personality trait neuroticism is significantly correlated with almost every psychiatric disorder and migraine. We also identify significant genetic sharing between disorders and early life cognitive measures (e.g., years of education and college attainment) in the general population, demonstrating positive correlation with several psychiatric disorders (e.g., anorexia nervosa and bipolar disorder) and negative correlation with several neurological phenotypes (e.g., Alzheimer's disease and ischemic stroke), even though the latter are considered to result from specific processes that occur later in life. Extensive simulations were also performed to inform how statistical power, diagnostic misclassification, and phenotypic heterogeneity influence genetic correlations. CONCLUSION The high degree of genetic correlation among many of the psychiatric disorders adds further evidence that their current clinical boundaries do not reflect distinct underlying pathogenic processes, at least on the genetic level. This suggests a deeply interconnected nature for psychiatric disorders, in contrast to neurological disorders, and underscores the need to refine psychiatric diagnostics. Genetically informed analyses may provide important "scaffolding" to support such restructuring of psychiatric nosology, which likely requires incorporating many levels of information. By contrast, we find limited evidence for widespread common genetic risk sharing among neurological disorders or across neurological and psychiatric disorders. We show that both psychiatric and neurological disorders have robust correlations with cognitive and personality measures. Further study is needed to evaluate whether overlapping genetic contributions to psychiatric pathology may influence treatment choices. Ultimately, such developments may pave the way toward reduced heterogeneity and improved diagnosis and treatment of psychiatric disorders

The genetic architecture of the human cerebral cortex

The cerebral cortex underlies our complex cognitive capabilities, yet little is known about the specific genetic loci that influence human cortical structure. To identify genetic variants that affect cortical structure, we conducted a genome-wide association meta-analysis of brain magnetic resonance imaging data from 51,665 individuals. We analyzed the surface area and average thickness of the whole cortex and 34 regions with known functional specializations. We identified 199 significant loci and found significant enrichment for loci influencing total surface area within regulatory elements that are active during prenatal cortical development, supporting the radial unit hypothesis. Loci that affect regional surface area cluster near genes in Wnt signaling pathways, which influence progenitor expansion and areal identity. Variation in cortical structure is genetically correlated with cognitive function, Parkinson's disease, insomnia, depression, neuroticism, and attention deficit hyperactivity disorder

Genetic Overlap Between Alzheimer’s Disease and Bipolar Disorder Implicates the MARK2 and VAC14 Genes

Background: Alzheimer's disease (AD) and bipolar disorder (BIP) are complex traits influenced by numerous common genetic variants, most of which remain to be detected. Clinical and epidemiological evidence suggest that AD and BIP are related. However, it is not established if this relation is of genetic origin. Here, we applied statistical methods based on the conditional false discovery rate (FDR) framework to detect genetic overlap between AD and BIP and utilized this overlap to increase the power to identify common genetic variants associated with either or both traits. Methods: We obtained genome wide association studies data from the International Genomics of Alzheimer's Project part 1 (17,008 AD cases and 37,154 controls) and the Psychiatric Genetic Consortium Bipolar Disorder Working Group (20,352 BIP cases and 31,358 controls). We used conditional QQ-plots to assess overlap in common genetic variants between AD and BIP. We exploited the genetic overlap to re-rank test-statistics for AD and BIP and improve detection of genetic variants using the conditional FDR framework. Results: Conditional QQ-plots demonstrated a polygenic overlap between AD and BIP. Using conditional FDR, we identified one novel genomic locus associated with AD, and nine novel loci associated with BIP. Further, we identified two novel loci jointly associated with AD and BIP implicating the MARK2 gene (lead SNP rs10792421, conjunctional FDR=0.030, same direction of effect) and the VAC14 gene (lead SNP rs11649476, conjunctional FDR=0.022, opposite direction of effect). Conclusions: We found polygenic overlap between AD and BIP and identified novel loci for each trait and two jointly associated loci. Further studies should examine if the shared loci implicating the MARK2 and VAC14 genes could explain parts of the shared and distinct features of AD and BIP

Bipolar multiplex families have an increased burden of common risk variants for psychiatric disorders.

Multiplex families with a high prevalence of a psychiatric disorder are often examined to identify rare genetic variants with large effect sizes. In the present study, we analysed whether the risk for bipolar disorder (BD) in BD multiplex families is influenced by common genetic variants. Furthermore, we investigated whether this risk is conferred mainly by BD-specific risk variants or by variants also associated with the susceptibility to schizophrenia or major depression. In total, 395 individuals from 33 Andalusian BD multiplex families (166 BD, 78 major depressive disorder, 151 unaffected) as well as 438 subjects from an independent, BD case/control cohort (161 unrelated BD, 277 unrelated controls) were analysed. Polygenic risk scores (PRS) for BD, schizophrenia (SCZ), and major depression were calculated and compared between the cohorts. Both the familial BD cases and unaffected family members had higher PRS for all three psychiatric disorders than the independent controls, with BD and SCZ being significant after correction for multiple testing, suggesting a high baseline risk for several psychiatric disorders in the families. Moreover, familial BD cases showed significantly higher BD PRS than unaffected family members and unrelated BD cases. A plausible hypothesis is that, in multiplex families with a general increase in risk for psychiatric disease, BD development is attributable to a high burden of common variants that confer a specific risk for BD. The present analyses demonstrated that common genetic risk variants for psychiatric disorders are likely to contribute to the high incidence of affective psychiatric disorders in the multiplex families. However, the PRS explained only part of the observed phenotypic variance, and rare variants might have also contributed to disease development

The genetics of the mood disorder spectrum:genome-wide association analyses of over 185,000 cases and 439,000 controls

Background Mood disorders (including major depressive disorder and bipolar disorder) affect 10-20% of the population. They range from brief, mild episodes to severe, incapacitating conditions that markedly impact lives. Despite their diagnostic distinction, multiple approaches have shown considerable sharing of risk factors across the mood disorders. Methods To clarify their shared molecular genetic basis, and to highlight disorder-specific associations, we meta-analysed data from the latest Psychiatric Genomics Consortium (PGC) genome-wide association studies of major depression (including data from 23andMe) and bipolar disorder, and an additional major depressive disorder cohort from UK Biobank (total: 185,285 cases, 439,741 controls; non-overlapping N = 609,424). Results Seventy-three loci reached genome-wide significance in the meta-analysis, including 15 that are novel for mood disorders. More genome-wide significant loci from the PGC analysis of major depression than bipolar disorder reached genome-wide significance. Genetic correlations revealed that type 2 bipolar disorder correlates strongly with recurrent and single episode major depressive disorder. Systems biology analyses highlight both similarities and differences between the mood disorders, particularly in the mouse brain cell-types implicated by the expression patterns of associated genes. The mood disorders also differ in their genetic correlation with educational attainment – positive in bipolar disorder but negative in major depressive disorder. Conclusions The mood disorders share several genetic associations, and can be combined effectively to increase variant discovery. However, we demonstrate several differences between these disorders. Analysing subtypes of major depressive disorder and bipolar disorder provides evidence for a genetic mood disorders spectrum

Table1_Binding pocket stabilization by high-throughput screening of yeast display libraries.DOCX

Protein dynamics have a great influence on the binding pockets of some therapeutic targets. Flexible protein binding sites can result in transient binding pocket formation which might have a negative impact on drug screening efforts. Here, we describe a protein engineering strategy with FK506-binding protein 51 (FKBP51) as a model protein, which is a promising target for stress-related disorders. High-throughput screening of yeast display libraries of FKBP51 resulted in the identification of variants exhibiting higher affinity binding of conformation-specific FKBP51 selective inhibitors. The gene libraries of a random mutagenesis and site saturation mutagenesis of the FK1 domain of FKBP51 encoding sequence were used to create a yeast surface display library. Fluorescence-activated cell sorting for FKBP51 variants that bind conformation-specific fluorescently labeled ligands with high affinity allowed for the identification of 15 different protein variants with improved binding to either, or both FKBP51-specific ligands used in the screening, with improved affinities up to 34-fold compared to the wild type. These variants will pave the way to a better understanding of the conformational flexibility of the FKBP51 binding pocket and may enable the isolation of new selective ligands that preferably and selectively bind the active site of the protein in its open conformation state.</p

Sotakarjut : Roolipelin visuaalinen taide ja ammatillinen vuorovaikutus projektityöympäristössä.

Opinnäytetyöni pohjautuu vuonna 2012 julkaistavaan roolipelikirjaan nimeltä Sotakarjut. TAMKista valmistuvana visuaalisena suunnittelijana olin mukana piirtämässä sekä konseptitaiteidetta, että kuvituskuvia kyseistä teosta varten. Kyseisen projektin aikana työskentelin sekä taiteellisen johdon alaisuudessa, että osana useamman kuvittajan/konseptitaiteilijan muodostamaa työryhmää, jolloin taiteellisesta ja ammatillisesta vuorovaikutuksesta tuli selkeä osa projektin kulkua. Näistä lähtökohdista päätin myös kirjoittaa opinnäytetyöni kirjallisen osuuden. Opinnäytetyöni tarkoitus on valaista monin tavoin vuorovaikutteiseksi muodostuvien projektityöympäristöjen vaikutusta taiteilijaan niin kuvittajan, kuin konseptitaiteilijan ammattissa. Valitsemaani aihetta lähestyn tekemäni projektin rajaaman kontekstin sisällä käymällä aluksi läpi nämä kaksi visuaalisen suunnittelun ammattihaaraa kartoittamalla taiteilijan roolia osana laajempaa projektityöympäristöä, ja tarjoamalla käytännön kautta lähestyvää näkökantaa esittelemilleni asioille. Tutkin sekä ulkoisen, että sisäisen vuorovaikutuksen merkitystä ja niiden luomia mahdollisuuksia luovalla alalla toimivan taiteilijan näkökulmasta. Esitän myös käytännön tapoja siitä, miten nämä vuorovaikutukselliset elementit voidaan valjastaa palvelemaan yksilön omaa ammatillista ja taiteellista kehitystä.My thesis work is based on a traditional role-playing-game that is set to be published in the year 2012, a book titled Sotakarjut. About to graduate from TAMK, Tampere University of Applied Sciences, as a visual designer I took part in this project, in a role that covered two distinct fields of profession: concept art and a illustration. As an single artist I worked under a artistic supervision and within a project that had several illustrators/consept artist in its ranks, thus making it essential to work as a singular creative team. In this thesis I will try to shed some light on how project-related work enviroments involves a specific kind of artististic approach from professional consept artists and illustrators. Having my project part work as my general context, I will start by making a general introduction on both of these creative professions, giving an overview on the subject of artististic work within project enviroments. I will also present a practical approach by adding examples from my actual project to support the theory based information I am presenting. In addition I will write about the many effects that professional interaction has within a working eviroment, from the artist view-point. Through this I will also explain how it is possible to assert yourself as a professional artist, so that this interaction can be made beneficial to the indivudual, both artistically and professional