Michael R. Barnes

Disruption of the neurexin 1 gene is associated with schizophrenia

Deletions within the neurexin 1 gene (NRXN1; 2p16.3) are associated with autism and have also been reported in two families with schizophrenia. We examined NRXN1, and the closely related NRXN2 and NRXN3 genes, for copy number variants (CNVs) in 2977 schizophrenia patients and 33 746 controls from seven European populations (Iceland, Finland, Norway, Germany, The Netherlands, Italy and UK) using microarray data. We found 66 deletions and 5 duplications in NRXN1, including a de novo deletion: 12 deletions and 2 duplications occurred in schizophrenia cases (0.47%) compared to 49 and 3 (0.15%) in controls. There was no common breakpoint and the CNVs varied from 18 to 420 kb. No CNVs were found in NRXN2 or NRXN3. We performed a Cochran-Mantel-Haenszel exact test to estimate association between all CNVs and schizophrenia (P = 0.13; OR = 1.73; 95% CI 0.81-3.50). Because the penetrance of NRXN1 CNVs may vary according to the level of functional impact on the gene, we next restricted the association analysis to CNVs that disrupt exons (0.24% of cases and 0.015% of controls). These were significantly associated with a high odds ratio (P = 0.0027; OR 8.97, 95% CI 1.8-51.9). We conclude that NRXN1 deletions affecting exons confer risk of schizophrenia.

Pathway and network-based analysis of genome-wide association studies in multiple sclerosis

Genome-wide association studies (GWAS) testing several hundred thousand SNPs have been performed in multiple sclerosis (MS) and other complex diseases. Typically, the number of markers in which the evidence for association exceeds the genome-wide significance threshold is very small, and markers that do not exceed this threshold are generally neglected. Classical statistical analysis of these datasets in MS revealed genes with known immunological functions. However, many of the markers showing modest association may represent false negatives. We hypothesize that certain combinations of genes flagged by these markers can be identified if they belong to a common biological pathway. Here we conduct a pathway-oriented analysis of two GWAS in MS that takes into account all SNPs with nominal evidence of association (P < 0.05). Gene-wise P-values were superimposed on a human protein interaction network and searches were conducted to identify sub-networks containing a higher proportion of genes associated with MS than expected by chance. These sub-networks, and others generated at random as a control, were categorized for membership of biological pathways. GWAS from eight other diseases were analyzed to assess the specificity of the pathways identified. In the MS datasets, we identified sub-networks of genes from several immunological pathways including cell adhesion, communication and signaling. Remarkably, neural pathways, namely axon-guidance and synaptic potentiation, were also over-represented in MS. In addition to the immunological pathways previously identified, we report here for the first time the potential involvement of neural pathways in MS susceptibility.

Use of genome-wide association studies for drug repositioning

1. Weaver, T., Maurer, J. & Hayashizaki, Y. Nat. Rev. Genet. 5, 861–866 (2004). 2. Fan, M., Tsai, J., Chen, B., Fan, K. & LaBaer, J. Science 307, 1877 (2005). 3. Campbell, E.G. et al. J. Am. Med. Assoc. 287, 473– 480 (2002). control of reagents by the institution, they can often cause long delays for the researcher looking to obtain these reagents. Addgene has streamlined the technology transfer process by (i) using the universal biological material transfer agreement (UBMTA) as the basis for all transfers, (ii) making the agreements as consistent as possible across all institutions and (iii) allowing for electronic signatures from institutions that both contribute and request materials. This system has been used for >80,000 orders from >2,500 institutions worldwide. As more technology transfer offices have adapted to this system, the time required for MTA approval has been halved, with the median time now <36 h. Moving forward, it would be more efficient for institutions to implement a similar electronic MTA system for all academic resource transfers. Ultimately, BRCs like Addgene will be important for guiding academic laboratories into a new age of high-throughput research and corporate funding. We are seeing a paradigm shift in the pharmaceutical industry toward greater collaborations with academia self-sustaining and does not rely on outside funding. The most popular plasmids in the collection are empty backbones created for specific gene expression or knockdown experiments, control plasmids, and constructs used for generating lentiviruses and retroviruses. A quick look at Addgene’s most requested plasmids, according to laboratory (Table 1), reveals a collection of vectors that can be used in various applications across multiple disciplines. If a BRC like Addgene were not archiving and distributing these valuable reagents, they would be far less accessible to the scientific community3. Indeed, many researchers, especially those outside the discipline of the contributing laboratory, might not even realize that some of these powerful tools exist. Addgene has become a global repository, sending out approximately half of its requests to scientists outside the United States. Addgene now distributes genomic resources for large-scale projects, such as the Zinc Finger Consortium (http://www. zincfingers.org/), the Structural Genomics Consortium (http://www.thesgc.org/) and the Center for Genomic Engineering (http:// www.cge.umn.edu/). Moving forward, Addgene hopes to collaborate with additional groups to help support their archival and distribution efforts. In addition to archiving and distributing a physical reagent, Addgene also plays a crucial role by archiving information about these reagents and making it accessible to all potential users through an online database. Addgene’s website receives an average of 35,000 page views per weekday. Having clone information available helps with reproducibility and future use, especially because checking the accuracy of this information is often an onerous task for many laboratories. Similar to other BRCs, Addgene can handle large volumes of samples and data, which facilitates the development of efficient, large-scale processes for standardizing quality control and maintaining comprehensive databases of information. Currently, Addgene sequences key regions of all incoming constructs, which helps maintain a standardized bar for accuracy throughout the repository. Addgene has developed one of the first electronic material transfer agreement (MTA) systems, which has helped expedite the MTA process. Over the past few decades, there has been an increase in the use of MTAs for transferring reagents between academic and nonprofit organizations. Although MTAs may be a practical means of maintaining Use of genome-wide association studies for drug repositioning

Bioinformatics for geneticists.

List of Contributors. Foreword. SECTION I: AN INTRODUCTION TO BIOINFORMATICS FOR THE GENETICIST. Introduction: The Role of Genetic Bioinformatics (Michael R. Barnes and Ian C. Gray). Internet Resources for the Geneticist (Michael R. Barnes and Christopher Southan). Human Genetic Variation: Databases and Concepts (Michael R. Barnes). Finding, Delineating and Analysing Genes (Christopher Southan). SECTION II: THE IMPACT OF COMPLETE GENOME SEQUENCES ON GENETICS. Assembling a View of the Human Genome (Colin A. Semple). Mouse and Rat Genome Informatics (Judith A. Blake, Janan Eppig and Carol J. Bult). Genetic and Physical Map Resources -- An Integrated View (Michael R. Barnes). SECTION III: BIOINFORMATICS FOR GENETIC STUDY DESIGN. From Linkage Peak to Culprit Gene: Following Up Linkage Analysis of Complex Phenotypes with Population--based Association Studies (Ian C. Gray). Genetic Studies from Genomic Sequence (Michael R. Barnes). SNP Discovery and PCR--based Assay Design: From In Silico Data to the Laboratory Experiment (Ellen Vieux, Gabor Marth and Pui Kwok). Tools for Statistical Analysis of Genetic Data (Aruna Bansal, Peter R. Boyd and Ralph McGinnis). SECTION IV: BIOLOGICAL SEQUENCE ANALYSIS AND CHARACTERIZATION. Predictive Functional Analysis of Polymorphisms: An Overview (Michael R. Barnes). Functional In Silico Analysis of Non--coding SNPs (Thomas Werner). Amino Acid Properties and Consequences of Substitutions (Matthew J. Betts and Robert B. Russell). SECTION V: GENETICS/GENOMICS INTERFACES. Gene Expression Informatics and Analysis (Antoine H. C. van Kampen, Jan M. Ruijter, Barbera D. C. van Schaik, Huib N. Caron and Rogier Versteeg). Proteomic Informatics (Jerome Wojcik and Alexandre Hamburger). Concluding Remarks: Final Thoughts and Future Trends (Michael R. Barnes and Ian C. Gray). Appendix I. Appendix II. Glossary. Index.

Genome-Wide Association Study of Major Recurrent Depression in the U.K. Population

OBJECTIVE Studies of major depression in twins and families have shown moderate to high heritability, but extensive molecular studies have failed to identify susceptibility genes convincingly. To detect genetic variants contributing to major depression, the authors performed a genome-wide association study using 1,636 cases of depression ascertained in the U.K. and 1,594 comparison subjects screened negative for psychiatric disorders. METHOD Cases were collected from 1) a case-control study of recurrent depression (the Depression Case Control [DeCC] study; N=1346), 2) an affected sibling pair linkage study of recurrent depression (probands from the Depression Network [DeNT] study; N=332), and 3) a pharmacogenetic study (the Genome-Based Therapeutic Drugs for Depression [GENDEP] study; N=88). Depression cases and comparison subjects were genotyped at Centre National de Génotypage on the Illumina Human610-Quad BeadChip. After applying stringent quality control criteria for missing genotypes, departure from Hardy-Weinberg equilibrium, and low minor allele frequency, the authors tested for association to depression using logistic regression, correcting for population ancestry. RESULTS Single nucleotide polymorphisms (SNPs) in BICC1 achieved suggestive evidence for association, which strengthened after imputation of ungenotyped markers, and in analysis of female depression cases. A meta-analysis of U.K. data with previously published results from studies in Munich and Lausanne showed some evidence for association near neuroligin 1 (NLGN1) on chromosome 3, but did not support findings at BICC1. CONCLUSIONS This study identifies several signals for association worthy of further investigation but, as in previous genome-wide studies, suggests that individual gene contributions to depression are likely to have only minor effects, and very large pooled analyses will be required to identify them.

Health and population effects of rare gene knockouts in adult humans with related parents

Rare gene knockouts in adult humans On average, most peoples genomes contain approximately 100 completely nonfunctional genes. These loss-of-function (LOF) mutations tend to be rare and/or occur only as a single copy within individuals. Narasimhan et al. investigated LOF in a Pakistani population with high levels of consanguinity. Examining LOF alleles that were identical by descent, they found, as expected, an absence of homozygote LOF for certain protein-coding genes. However, they also identified many homozygote LOF alleles with no apparent deleterious phenotype, including some that were expected to confer genetic disease. Indeed, one family had lost the recombination-associated gene PRDM9. Science, this issue p. 474 The total loss of protein-coding genes, even those with the potential to confer genetic diseases, can be tolerated. Examining complete gene knockouts within a viable organism can inform on gene function. We sequenced the exomes of 3222 British adults of Pakistani heritage with high parental relatedness, discovering 1111 rare-variant homozygous genotypes with predicted loss of function (knockouts) in 781 genes. We observed 13.7% fewer homozygous knockout genotypes than we expected, implying an average load of 1.6 recessive-lethal-equivalent loss-of-function (LOF) variants per adult. When genetic data were linked to the individuals’ lifelong health records, we observed no significant relationship between gene knockouts and clinical consultation or prescription rate. In this data set, we identified a healthy PRDM9-knockout mother and performed phased genome sequencing on her, her child, and control individuals. Our results show that meiotic recombination sites are localized away from PRDM9-dependent hotspots. Thus, natural LOF variants inform on essential genetic loci and demonstrate PRDM9 redundancy in humans.

Analysis of gene expression in two large schizophrenia cohorts identifies multiple changes associated with nerve terminal function

Schizophrenia is a severe psychiatric disorder with a world-wide prevalence of 1%. The pathophysiology of the illness is not understood, but is thought to have a strong genetic component with some environmental influences on aetiology. To gain further insight into disease mechanism, we used microarray technology to determine the expression of over 30 000 mRNA transcripts in post-mortem tissue from a brain region associated with the pathophysiology of the disease (Brodmann area 10: anterior prefrontal cortex) in 28 schizophrenic and 23 control patients. We then compared our study (Charing Cross Hospital prospective collection) with that of an independent prefrontal cortex dataset from the Harvard Brain Bank. We report the first direct comparison between two independent studies. A total of 51 gene expression changes have been identified that are common between the schizophrenia cohorts, and 49 show the same direction of disease-associated regulation. In particular, changes were observed in gene sets associated with synaptic vesicle recycling, transmitter release and cytoskeletal dynamics. This strongly suggests multiple, small but synergistic changes in gene expression that affect nerve terminal function.

Lowering industry firewalls: pre-competitive informatics initiatives in drug discovery

Pharmaceutical research and development is facing substantial challenges that have prompted the industry to shift funding from early- to late-stage projects. Among the effects is a major change in the attitude of many companies to their internal bioinformatics resources: the focus has moved from the vigorous pursuit of intellectual property towards exploration of pre-competitive cross-industry collaborations and engagement with the public domain. High-quality, open and accessible data are the foundation of pre-competitive research, and strong public–private partnerships have considerable potential to enhance public data resources, which would benefit everyone engaged in drug discovery. In this article, we discuss the background to these changes and propose new areas of collaboration in computational biology and chemistry between the public domain and the pharmaceutical industry.

Advances in Blood Pressure Genomics

The elucidation of genes implicated in Mendelian forms of hypertension demonstrates rare variants with substantial effects are responsible, and often these genes lie within pathways managing sodium homeostasis. More recently with advances in affordable high-throughput genotyping strategies, multiple common genetic variants with modest effects on blood pressure (<1 mmHg systolic) have been discovered in the population. In aggregate, these common variants explain <3% of the variance of blood pressure. Although these findings may offer new mechanistic insights into the biology of blood pressure, a key question is can these findings translate into patient benefit? It is timely to reflect on recent advances in genomics, and the use of new resources, such as the 1000 Genomes Project and the Encyclopedia of DNA Elements, to annotate likely causal variants, and their relevance to cardiovascular disease. In this review, we discuss the advances in relation to our knowledge of the genetic architecture of blood pressure, and whether gene discoveries might influence cardiovascular risk assessment, help to stratify patient response to medicine, or identify new biological pathways for novel therapeutic targets.

Role for WNT16B in human epidermal keratinocyte proliferation and differentiation

WNT signalling regulates a variety of cell functions including cell fate, polarity, and differentiation via the canonical or β-catenin stabilisation pathway and/or the planar cell polarity or non-canonical pathway. We have previously demonstrated that two isoforms (A and B) from the WNT16 locus have differential expression in various adult human tissues. In this study we show that WNT16B but not WNT16A isoform was upregulated in basal cell carcinomas compared with normal skin. We further investigated the cellular and molecular functions of WNT16B in primary human epidermal keratinocytes and a keratinocyte cell line. Cellular expression of WNT16B neither stabilised β-catenin nor activated the lymphoid enhancer factor or T-cell factor transcriptional reporter in primary keratinocytes. WNT16B activated the Jun-N-terminal kinase cascade suggesting the activation of a non-canonical WNT signalling pathway. Constitutive expression of WNT16B significantly enhanced the rate of cell proliferation and prolonged clonogenicity in primary keratinocytes. Silencing WNT16B by RNA interference reduced keratinocyte proliferation. Furthermore, overexpression of WNT16B induced a hyperproliferation phenotype in an organotypical culture system. This work presents the first evidence that WNT16B activates human keratinocyte proliferation possibly via a β-catenin-independent non-canonical WNT transduction pathway.