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Depression in late-life is associated with increased risk of cognitive decline and development of all-cause dementia. The neurobiology of late-life depression (LLD) may involve both neurochemical and neurodegenerative mechanisms that are common to depression and dementia. Transgenic amyloid mouse models show evidence of early degeneration of monoamine systems. Informed by these preclinical data, the hypotheses were tested that a spatial covariance pattern of higher beta-amyloid (Aβ) and lower serotonin transporter availability (5-HTT) in frontal, temporal, and parietal cortical regions would distinguish LLD patients from healthy controls and the expression of this pattern would be associated with greater depressive symptoms. Twenty un-medicated LLD patients who met DSM-V criteria for major depression and 20 healthy controls underwent PET imaging with radiotracers for Aβ ([11C]-PiB) and 5-HTT ([11C]-DASB). A voxel-based multi-modal partial least squares (mmPLS) algorithm was applied to the parametric PET images to determine the spatial covariance pattern between the two radiotracers. A spatial covariance pattern was identified, including higher Aβ in temporal, parietal and occipital cortices associated with lower 5-HTT in putamen, thalamus, amygdala, hippocampus and raphe nuclei (dorsal, medial and pontine), which distinguished LLD patients from controls. Greater expression of this pattern, reflected in summary 5-HTT/Aβ mmPLS subject scores, was associated with higher levels of depressive symptoms. The mmPLS method is a powerful approach to evaluate the synaptic changes associated with AD pathology. This spatial covariance pattern should be evaluated further to determine whether it represents a biological marker of antidepressant treatment response and/or cognitive decline in LLD patients.

DPYSL2/CRMP2is a microtubule-stabilizing protein crucial for neurogenesis and associated with numerous psychiatric and neurodegenerative disorders.DPYSL2has multiple RNA and protein isoforms, but few studies have differentiated between them or explored their individual functions. We previously demonstrated in HEK293 cells that a schizophrenia -associated variant in theDPYSL2B isoform (DPYSL2-B) reduced the length of cellular projections, created a transcriptomic disturbance that captured schizophrenia etiology, and was acted upon by the mTOR pathway. In the present study, we follow up on these results by creating, to our knowledge, the first models of endogenousDPYSL2-Bknockout in human induced pluripotent stem cells and excitatory glutamatergic neurons. We use CRISPR/Cas9 to specifically knock outDPYSL2-Band observe corresponding reduction of its RNA and protein. The average length of dendrites in knockout neurons was reduced up to 58% compared to controls. Transcriptome analysis reveals disruptions in pathways highly relevant to psychiatric disease including mTOR signaling, cytoskeletal dynamics, immune function, calcium signaling, and cholesterol biosynthesis. We also observed a significant enrichment of our differentially expressed genes in schizophrenia GWAS-associated loci. Our findings clarify the functions of the humanDPYSL2-Bisoform and confirm its involvement in molecular pathologies shared between many psychiatric diseases.

Identifying genes and variants contributing to rare disease phenotypes and Mendelian conditions informs biology and medicine, yet potential phenotypic consequences for variation of >75% of the ~20,000 annotated genes in the human genome are lacking. Technical advances to assess rare variation genome-wide, particularly exome sequencing (ES), enabled establishment in the United States of the National Institutes of Health (NIH)-supported Centers for Mendelian Genomics (CMGs) and have facilitated collaborative studies resulting in novel “disease gene” discoveries. Pedigree-based genomic studies and rare variant analyses in families with suspected Mendelian conditions have led to the elucidation of hundreds of novel disease genes and highlighted the impact of de novo mutational events, somatic variation underlying nononcologic traits, incompletely penetrant alleles, phenotypes with high locus heterogeneity, and multilocus pathogenic variation. Herein, we highlight CMG collaborative discoveries that have contributed to understanding both rare and common diseases and discuss opportunities for future discovery in single-locus Mendelian disorder genomics. Phenotypic annotation of all human genes; development of bioinformatic tools and analytic methods; exploration of non-Mendelian modes of inheritance including reduced penetrance, multilocus variation, and oligogenic inheritance; construction of allelic series at a locus; enhanced data sharing worldwide; and integration with clinical genomics are explored. Realizing the full contribution of rare disease research to functional annotation of the human genome, and further illuminating human biology and health, will lay the foundation for the Precision Medicine Initiative.

Background Exposure to Toxoplasma gondii (TOXO) has been consistently associated with the development of schizophrenia, but the neurobiological mechanism through which this occurs is not well elucidated. Emerging data has broadly implicated the adaptive immune system as a possible pathway from TOXO infection to schizophrenia. In order to explore the hypothesis that crossreactive T cells could help mediate this relationship, we built upon the genetic analysis from our psychiatrically-enriched Ashkenazi Jewish cohort using an in silico approach to predict HLA reactivity to TOXO epitopes, with the aim of identifying host proteins that could be susceptible to T-cell-mediated molecular mimicry. Methods We used netMHCpan v4.0 to generate a library of 2182 oligopeptides from the TOXO proteome that were predicted to be strongly antigenic for individuals with HLA-C*04:01, an allele of interest since our analysis indicated that the odds ratio for TOXO infection were in the opposite direction for those with schizophrenia compared to controls. A predicted binding affinity less than 500 nM was used to identify epitopes that were likely to be biologically relevant. Epitopes identified by this approach were compared with human peptides for local sequence similarity using BLAST optimized for short peptide sequences in order to identify host proteins that could mimic TOXO antigens. Ingenuity Pathway Analysis was then used to interpret and synthesize possible biological relationships between predicted autoantigens and schizophrenia. Results Our pipeline identified 38 candidate proteins for molecular mimicry at a threshold of P Discussion Using an in silico approach, we identified 38 human proteins that could be targeted by a crossreactive T cell autoimmune response after exposure to TOXO. Several of these candidate proteins are highly relevant for genetic risk of schizophrenia and participate in molecular pathways that mediate antigen processing. CD8+ T cells contribute to autoimmunity through cytokine release and have been implicated in the relationship between TOXO exposure and schizophrenia. Though these results will need experimental confirmation, we hope that they will spur further research on the role of the adaptive immune system in toxoplasmosis and schizophrenia.

Introduction Alzheimer's disease (AD) is a disabling, common cause of dementia, and agitation is one of the most common and distressing symptoms for patients with AD. Escitalopram for agitation in Alzheimer's disease (S-CitAD) tests a novel, clinically derived therapeutic approach to treat agitation in patients with AD. Methods S-CitAD is a NIH-funded, investigator-initiated, randomized, multicenter clinical trial. Participants receive a structured psychosocial intervention (PSI) as standard of care. Participants without sufficient response to PSI are randomized to receive 15 mg escitalopram/day or a matching placebo in addition to PSI. Primary outcome is the Modified Alzheimer's Disease Cooperative Study - Clinical Global Impression of Change (mADCS-CGIC). Discussion S-CitAD will provide information about a practical, immediately available approach to treating agitation in patients with AD. S-CitAD may become a model of how to evaluate and predict treatment response in patients with AD and agitation as a neuropsychiatric symptom (ClinicalTrials.gov Identifier: NCT03108846).

Objective: To assess potential genetic influences on citalopram treatment efficacy for agitation in individuals with Alzheimer dementia (AD). Six functional genetic variants were studied in the following genes: serotonin receptor 2A ( HTR2A-T102C), serotonin receptor 2C ( HTR2C-Cys23Ser), serotonin transporter ( 5HTT-LPR), brain-derived neurotropic factor (BDNF-Val66Met), apolipoprotein E (ε2, ε3, ε4 variants), and cytochrome P450 ( CYP2C19). Treatment response by genotype was measured by (1) the agitation domain of the Neurobehavioral Rating Scale, (2) the modified Alzheimer Disease Cooperative Study—Clinical Global Impression of Change scale (mADCS-CGIC), (3) the agitation domain of the Neuropsychiatric Inventory (NPI), and (4) the Cohen-Mansfield Agitation Inventory. Method: We utilized data from the Citalopram for Agitation in Alzheimer’s Disease (CitAD) database. CitAD was a 9-week randomized, double-blind, placebo-controlled multicenter clinical trial showing significant improvement in agitation and caregiver distress in patients treated with citalopram. Proportional odds logistic regression and mixed effects models were used to examine the above-mentioned outcome measures. Results: Significant interactions were noted on the NPI agitation domain for HTR2A (likelihood ratio [LR] = 6.19, df = 2, P = .04) and the mADCS-CGIC for HTR2C (LR = 4.33, df = 2, P = .02) over 9 weeks. Discussion: Treatment outcomes in CitAD showed modest, although statistically significant, influence of genetic variation at HTR2A and HTR2C loci. Future studies should continue to examine the interaction of known genetic variants with antidepressant treatment in patients with AD having agitation.

Discovering the genetic basis of a Mendelian phenotype establishes a causal link between genotype and phenotype, making possible carrier and population screening and direct diagnosis. Such discoveries also contribute to our knowledge of gene function, gene regulation, development, and biological mechanisms that can be used for developing new therapeutics. As of February 2015, 2,937 genes underlying 4,163 Mendelian phenotypes have been discovered, but the genes underlying ∼50% (i.e., 3,152) of all known Mendelian phenotypes are still unknown, and many more Mendelian conditions have yet to be recognized. This is a formidable gap in biomedical knowledge. Accordingly, in December 2011, the NIH established the Centers for Mendelian Genomics (CMGs) to provide the collaborative framework and infrastructure necessary for undertaking large-scale whole-exome sequencing and discovery of the genetic variants responsible for Mendelian phenotypes. In partnership with 529 investigators from 261 institutions in 36 countries, the CMGs assessed 18,863 samples from 8,838 families representing 579 known and 470 novel Mendelian phenotypes as of January 2015. This collaborative effort has identified 956 genes, including 375 not previously associated with human health, that underlie a Mendelian phenotype. These results provide insight into study design and analytical strategies, identify novel mechanisms of disease, and reveal the extensive clinical variability of Mendelian phenotypes. Discovering the gene underlying every Mendelian phenotype will require tackling challenges such as worldwide ascertainment and phenotypic characterization of families affected by Mendelian conditions, improvement in sequencing and analytical techniques, and pervasive sharing of phenotypic and genomic data among researchers, clinicians, and families.

Repetitive sequences comprise a large portion of the genome and are often thought of as “junk DNA.” They are a significant source of genetic variation, particularly Alu elements. Their functional consequence is frequently dismissed. Here, we test the hypothesis that Alu polymorphisms contribute to phenotypic differences between individuals. We identified an enrichment of Alu polymorphisms in regions of the genome associated with human disease risk. Further, we find 44 instances where the trait-associated SNP is a surrogate for presence or absence of an Alu insertion. This finding indicates that the Alu may be the variant effecting disease risk, an intriguing possibility given its size and regulatory potential. This work emphasizes the importance of considering repeat polymorphisms in functional variant analysis.

Significance Repetitive sequences comprise a large portion of the genome and are often thought of as “junk DNA.” They are a significant source of genetic variation, particularly Alu elements. Their functional consequence is frequently dismissed. Here, we test the hypothesis that Alu polymorphisms contribute to phenotypic differences between individuals. We identified an enrichment of Alu polymorphisms in regions of the genome associated with human disease risk. Further, we find 44 instances where the trait-associated SNP is a surrogate for presence or absence of an Alu insertion. This finding indicates that the Alu may be the variant effecting disease risk, an intriguing possibility given its size and regulatory potential. This work emphasizes the importance of considering repeat polymorphisms in functional variant analysis.

X-linked adrenoleukodystrophy (XALD), a neurological disorder caused by mutations in the peroxisomal membrane protein gene ABCD1, presents as a rapidly progressing, inflammatory cerebral demyelination (cerebral cases) or a slowly progressing, distal axonopathy (non-cerebral cases). Specific ABCD1 defects do not explain this significant phenotypic variation. Patients have increased plasma and tissue very long chain fatty acid levels and increased cellular oxidative stress and oxidative damage. Superoxide dismutase 2 (SOD2), at candidate modifier locus 6q25.3, detoxifies superoxide radicals protecting against oxidative stress and damage. We tested an SOD2 variant C47T (Ala16Val) associated with reduced enzymatic activity as a potential modifier gene of cerebral demyelinating disease by comparing 117 cerebral XALD cases with 105 non-cerebral XALD cases. The hypoactive valine allele of the variant was associated with cerebral disease under a dominant model in the full data set (p = 0.04; ORT* = 1.90, 95% CI 1.01-3.56) and the non-childhood cerebral disease subset (p = 0.03; ORT* = 2.47, 95% CI 1.08-5.61). Three tag SNPs were genotyped to test for additional SNP or haplotype associations. A common haplotype, GTAC, which included the SOD2 valine allele, was associated with cerebral disease in the full data set (p = 0.03; OR = 1.75, 95% CI 1.11-2.75) and the non-childhood cerebral disease subset (p = 0.008; OR = 2.20, 95% CI 1.27-3.83). There was no association between childhood cerebral XALD and the C47T variant or the GTAC haplotype. Thus, reduced SOD2 activity may contribute to the development of cerebral demyelination in adolescent and adult XALD patients.

Schizophrenia (SZ) is a heritable and complex psychiatric disorder with an estimated worldwide prevalence of about 1%. Research on the risk factors for SZ has thus far yielded few clues to causes, but has pointed to a heterogeneous etiology that likely involves multiple genes and gene-environment interactions. In this manuscript, we apply a novel method (trio logic regression, Li et al., 2009) to case-parent trio data from a SZ candidate gene study conducted on families of Ashkenazi Jewish descent, and demonstrate the method's ability to detect multi-gene models for SZ risk in the family-based design. In particular, we demonstrate how this method revealed a genotype-phenotype association that includes an allele without marginal effect. Genet. Epidemiol. 34: 396–406, 2010. © 2010 Wiley-Liss, Inc.

Schizophrenia (SCZ) is a severe, debilitating mental illness which has a significant genetic component. The identification of genetic factors related to SCZ has been challenging and these factors remain largely unknown. To evaluate the contribution of de novo variants (DNVs) to SCZ, we sequenced the exomes of 53 individuals with sporadic SCZ and of their non-affected parents. We identified 49 DNVs, 18 of which were predicted to alter gene function, including 13 damaging missense mutations, 2 conserved splice site mutations, 2 nonsense mutations, and 1 frameshift deletion. The average number of exonic DNV per proband was 0.88, which corresponds to an exonic point mutation rate of 1.7×10(-8) per nucleotide per generation. The non-synonymous-to-synonymous mutation ratio of 2.06 did not differ from neutral expectations. Overall, this study provides a list of 18 putative candidate genes for sporadic SCZ, and when combined with the results of similar reports, identifies a second proband carrying a non-synonymous DNV in the RGS12 gene.

In March 2013 the American College of Medical Genetics and Genomics published a list of 56 genes with the recommendation that pathogenic and likely pathogenic variants detected incidentally by clinical sequencing be reported to patients. As an initial step in determining the practical consequences of this recommendation in the research setting, we searched for variants in these genes in 232 whole-exome sequences from the Baylor-Hopkins Center for Mendelian Genomics.We identified rare, nonsynonymous, and splicing single-nucleotide variants and insertions/deletions and assessed variant classification using the Human Gene Mutation, Emory, and ClinVar databases. We analyzed the burden of mutation in each of the 56 genes and determined which variants should be reported to patients.Our filtering resulted in 249 distinct variants, with a mean of 1.69 variants per individual. Half of these were novel missense mutations not classified by any of the three reference databases. Of 101 variants listed in the Human Gene Mutation Database, 48 were also in ClinVar and 3 were also in Emory; half of these shared variants were classified discordantly between databases. Some genes consistently had greater variation than others. In total, 0.86% of individuals had a reportable incidental variant.These observations demonstrate some current challenges of assessing phenotypic consequences of incidental variants for counseling patients.

Neuroglobin (NGB) is a neuron-specific vertebrate globin shown to protect against hypoxia, ischemia, oxidative stress and the toxic effects of Amyloid-beta. Following on our and others' results highlighting the importance of NGB expression in disease, we searched for genetic determinants of its expression. We found that a microRNA expressed with the NGB transcript shows significant target enrichments in the angiogenesis pathway and the Alzheimer disease/presenilin pathway. Using reporter constructs we identified potential promoter/enhancer elements between the transcription start site and 1,142 bp upstream. Using 184 post-mortem temporal lobe samples we replicated the reported negative effect of age, and after genotyping tagging SNPs we found one (rs981471) showing a significant correlation with the gene's expression and another (rs8014408) showing an interaction with age, the rare C allele being correlated with higher expression and faster decline. The two SNPs are towards the 3′ end of NGB within the same LD block, 52 Kb apart and modestly correlated (r 2 = 0.5). Next generation sequencing of the same 184 temporal lobe samples and 79 confirmed AD patients across the entire gene region (including >12 Kb on the 3′ and 5′ flank) revealed limited coding variation, suggesting purifying selection of NGB, but did not identify regulatory or disease associated rare variants. A dinucleotide repeat in intron 1 with extensive evidence of functionality showed interesting but inconclusive results, as it was not amenable to further molecular analysis.

Despite the strongly held view that schizophrenia (SZ) shows substantial genetic heterogeneity, pathway heterogeneity, as seen in cancer where different pathways are affected in similar tumors, has not been explored. We explore this possibility in a case-only study of the neuregulin signaling pathway (NSP), which has been prominently implicated in SZ and for which there is detailed knowledge on the ligand- and receptor-processing steps through β- and γ-secretase cleavage. We hypothesize that more than one damaging variants in the NSP genes might be necessary to cause disease, leading to an apparent clustering of such variants in only the few patients with affected NSP. We analyze linkage and next-generation sequencing results for the genes encoding components of the pathway, including NRG1, NRG3, ERBB4, β-secretase and the γ-secretase complex. We find multiple independent examples of supporting evidence for this hypothesis: (i) increased linkage scores over NSP genes, (ii) multiple positive interlocus correlations of linkage scores across families suggesting each family is linked to either many or none of the genes, (iii) aggregation of predicted damaging variants in a subset of individuals and (iv) significant phenotypic differences of the subset of patients carrying such variants. Collectively, our data strongly support the hypothesis that the NSP is affected by multiple damaging variants in a subset of phenotypically distinct patients. On the basis of this, we propose a general model of pathway heterogeneity in SZ, which, in part, may explain its phenotypic variability and genetic complexity.

We previously reported a schizophrenia-associated polymorphic CT di-nucleotide repeat (DNR) at the 5′-untranslated repeat (UTR) of DPYSL2, which responds to mammalian target of Rapamycin (mTOR) signaling with allelic differences in reporter assays. Now using microarray analysis, we show that the DNR alleles interact differentially with specific proteins, including the mTOR-related protein HuD/ELAVL4. We confirm the differential binding to HuD and other known mTOR effectors by electrophoretic mobility shift assays. We edit HEK293 cells by CRISPR/Cas9 to carry the schizophrenia risk variant (13DNR) and observe a significant reduction of the corresponding CRMP2 isoform. These edited cells confirm the response to mTOR inhibitors and show a twofold shortening of the cellular projections. Transcriptome analysis of these modified cells by RNA-seq shows changes in 12.7% of expressed transcripts at a false discovery rate of 0.05. These transcripts are enriched in immunity-related genes, overlap significantly with those modified by the schizophrenia-associated gene, ZNF804A, and have a reverse expression signature from that seen with antipsychotic drugs. Our results support the functional importance of the DPYSL2 DNR and a role for mTOR signaling in schizophrenia.

Six novel polymorphic short sequence repeats were identified and localized on the linkage map of human chromosome 21 by genotyping the CEPH reference pedigrees. One of these markers, the tetrameric (AAAG)n repeat D21S1245, was found to be hypermutable. In the DNAs from lymphoblastoid cell lines of members of the 40 CEPH families a total of 18 new alleles were detected. These new alleles, sometimes appearing in mosaic forms, arose equally in paternal and maternal DNAs, and could be equally larger or smaller than the alleles from which they were derived. The larger alleles of D21S1245 are more prone to be converted to new alleles. None of the new alleles with mosaicism were present in the corresponding genomic blood DNA, and therefore originated during or after the establishment of the lymphoblastoid cell lines; half of the new alleles without mosaicism were also found in genomic blood DNA of the appropriate CEPH individuals. The range of germline mutation rate observed in the 716 meioses examined was 0.56-1.4 x 10(-2); the range of somatic mutations observed in the 405 cell lines examined was 1.96-3.46 x 10(-2). This is one of the most hypermutable microsatellite repeat polymorphism in the human genome detected to date. D21S1245, is highly polymorphic (heterozygosity of 0.96) and maps between D21S231 and D21S198.

Genetic variation in CLU encoding clusterin has been associated with Alzheimer's disease (AD) through replicated genome-wide studies, but the underlying mechanisms remain unknown. Following earlier reports that tightly regulated CLU alternative transcripts have different functions, we tested CLU single-nucleotide polymorphisms (SNPs), including those associated with AD for quantitative effects on individual alternative transcripts. In 190 temporal lobe samples without pathology, we found that the risk allele of the AD-associated SNP rs9331888 increases the relative abundance of transcript NM_203339 (P=4.3 × 10−12). Using an independent set of 115 AD and control samples, we replicated this result (P=0.0014) and further observed that multiple CLU transcripts are at higher levels in AD compared with controls. The AD SNP rs9331888 is located in the first exon of NM_203339 and therefore, it is a functional candidate for the observed effects. We tested this hypothesis by in vitro dual luciferase assays using SK-N-SH cells and mouse primary cortical neurons and found allelic effects on enhancer function, consistent with our results on post-mortem human brain. These results suggest a biological mechanism for the genetic association of CLU with AD risk and indicate that rs9331888 is one of the functional DNA variants underlying this association.

Low-copy repeats (LCRs) are genomic features that affect chromosome stability and can produce disease-associated rearrangements. We describe members of three families with deletions in 10q22.3-q23.31, a region harboring a complex set of LCRs, and demonstrate that rearrangements in this region are associated with behavioral and neurodevelopmental abnormalities, including cognitive impairment, autism, hyperactivity, and possibly psychiatric disease. Fine mapping of the deletions in members of all three families by use of a custom 10q oligonucleotide array-based comparative genomic hybridization (NimbleGen) and polymerase chain reaction-based methods demonstrated a different deletion in each family. In one proband, the deletion breakpoints are associated with DNA fragments containing noncontiguous sequences of chromosome 10, whereas, in the other two families, the breakpoints are within paralogous LCRs, removing approximately 7.2 Mb and 32 genes. Our data provide evidence that the 10q22-q23 genomic region harbors one or more genes important for cognitive and behavioral development and that recurrent deletions affecting this interval define a novel genomic disorder.