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Many single nucleotide polymorphisms (SNPs) have been identified for Bipolar Disorder (BD), but association between SNPs and BD can vary depending on the population tested. SNPs rs10994336 and rs9804190 in ANK3 and rs1006737 in CACNA1C have emerged as the most highly replicated SNPs significantly associated with BD. The aim of the present study was to assess the association of these SNPs with BD in the Pakistani population, which has never before been examined. A total of 120 BD and 120 control individuals from Pakistan were examined in this analysis. Genotyping results indicated that rs1006737 in CACNA1C was significantly associated with BD, while rs10994336 or rs9804190 in ANK3 was not significant when examined individually. However, risk score assessment found that the presence of two or more risk alleles was significantly associated with disease, indicating that risk alleles from ANK3 and CACNA1C may additively contribute to BD. A protein-protein interaction network was generated using STRING to probe the relationship between ANK3 and CACNA1C interactors and their associations with BD. While none of the interactors are directly linked to BD, they play a role in pathways linked to BD, including oxytocin and dopamine signaling pathways. Collectively, these results reveal a significant association of CACNA1C with BD among the Pakistani population, extending results from other ethnic groups to the Pakistani population for the first time.

Background Diffusion tensor imaging (DTI) studies, which allow the in-vivo investigation of brain tissue integrity, have shown that bipolar disorder (BD) patients present signs of white matter dysconnectivity. In parallel, genome-wide association studies (GWAS) identified several risk genetic variants for BD. I Methods In this mini-review, we summarized DTI studies coupling tract-based spatial statistics (TBSS), a reliable technique exploring white matter axon bundles, and genetics in BD. We performed a bibliographic search on PUBMED, using the search terms “TBSS”, “genetics”, “genome”, “genes”, “polymorphism”, “bipolar disorder”. Results Ten studies met these inclusion criteria. ANK3 and ZNF804A polymorphisms have shown the most consistent results, with the risk alleles showing abnormal white matter integrity in patients with BD. Limitations Current studies are limited by the investigation of single SNPs in small and chronically treated samples. Conclusions Most considered TBSS-DTI studies found associations between decreased white matter integrity and genetic risk variants. These results suggest an involvement of dysmyelination in the pathogenesis of BD. The combination of TBSS with genotyping can be powerful to unveil the role of white matter in BD, in conjunction with risk genes. Future DTI studies should combine TBSS and GWAS in large populations of drug-free or minimally treated patients with BD at the onset of the disease.

Ankyrin 3 (ANK3) has been implicated as a genetic risk factor for bipolar disorder (BD), however the resulting pathophysiological and treatment implications remain elusive. In a preclinical systems biological approach, we aimed to characterize the behavioral and proteomic effects of Ank3 haploinsufficiency and chronic mood-stabilizer treatment in mice. Psychiatric-related behavior was evaluated with the novelty-suppressed feeding (NSF) paradigm, elevated plus maze (EPM) and a passive avoidance task (PAT). Tandem mass spectrometry (MSE) was employed for hippocampal proteome profiling. A functional enrichment approach based on protein-protein interactions (PPIs) was performed to outline which biological processes in the hippocampus were affected by Ank3 haploinsufficiency and lithium treatment. Proteomic abundance changes as detected by MSE or highlighted by PPI network modelling were followed up by targeted selected reaction monitoring (SRM). Increased psychiatric-related behavior in Ank3+/- mice was ameliorated by lithium in all assessments (NSF, EPM, PAT). MSE followed by modular PPI clustering and functional annotation enrichment pointed towards kinesin-related axonal transport and glutamate signaling as mediators of Ank3+/- pathophysiology and lithium treatment. SRM validated this hypothesis and further confirmed abundance changes of ANK3 interaction partners. We propose that psychiatric-related behavior in Ank3+/- mice is connected to a disturbance of the kinesin cargo system, resulting in a dysfunction of neuronal ion channel and glutamate receptor transport. Lithium reverses this molecular signature, suggesting the promotion of anterograde kinesin transport as part of its mechanism of action in ameliorating Ank3-related psychiatric-related behavior.

Background Ankyrin-3 ( ANK3 ) was one of the first genes to reach significance in a bipolar disorder genome-wide association study. Many subsequent association studies confirmed this finding and implicated this gene in schizophrenia. However, the exact nature of the role of ANK3 in the pathophysiology remains elusive. In particular, the specific isoforms involved and the nature of the imbalance are unknown. Methods We genotyped a Norwegian sample of 402 patients with bipolar disorder, 293 patients with schizophrenia, and 330 healthy control subjects genome-wide with the Illumina Human Exome BeadChip. We performed allelic association tests at the genome-wide and gene levels and found a significantly associated single nucleotide polymorphism in a splice site of ANK3 . We replicated this finding in two other samples and studied the functional effect of this single nucleotide polymorphism by performing quantitative polymerase chain reaction on the affected exon junction in complementary DNA from blood total RNA. Results The splice site single nucleotide polymorphism (rs41283526) is located in an alternatively spliced exon of ANK3 and has a strong and significant protective effect against bipolar disorder (odds ratio=.31) and schizophrenia (odds ratio=.21). The minor allele of rs41283526 is a loss-of-function variant that disables the correct splicing of the transcript. Data from the BrainSpan human developmental transcriptome show that the exon bearing this variant is expressed only in a minor isoform of ANK3 , the transcription of which is initiated in early adolescence. Conclusions Our results suggest that an elevated expression of this transcript starting in adolescence may be an important factor in the pathophysiology of schizophrenia and bipolar disorder.

Ankyrin-3 (ANK3) was one of the first genes to reach significance in a bipolar disorder genome-wide association study. Many subsequent association studies confirmed this finding and implicated this gene in schizophrenia. However, the exact nature of the role of ANK3 in the pathophysiology remains elusive. In particular, the specific isoforms involved and the nature of the imbalance are unknown.We genotyped a Norwegian sample of 402 patients with bipolar disorder, 293 patients with schizophrenia, and 330 healthy control subjects genome-wide with the Illumina Human Exome BeadChip. We performed allelic association tests at the genome-wide and gene levels and found a significantly associated single nucleotide polymorphism in a splice site of ANK3. We replicated this finding in two other samples and studied the functional effect of this single nucleotide polymorphism by performing quantitative polymerase chain reaction on the affected exon junction in complementary DNA from blood total RNA.The splice site single nucleotide polymorphism (rs41283526) is located in an alternatively spliced exon of ANK3 and has a strong and significant protective effect against bipolar disorder (odds ratio = .31) and schizophrenia (odds ratio = .21). The minor allele of rs41283526 is a loss-of-function variant that disables the correct splicing of the transcript. Data from the BrainSpan human developmental transcriptome show that the exon bearing this variant is expressed only in a minor isoform of ANK3, the transcription of which is initiated in early adolescence.Our results suggest that an elevated expression of this transcript starting in adolescence may be an important factor in the pathophysiology of schizophrenia and bipolar disorder.

Background Ankyrin3 has been identified as a risk gene for Bipolar Disorder (BD) and autism spectrum disorders by multiple genome-wide and targeted association studies as well as sequencing studies. Carriers of the BD risk alleles have reduced ankyrin3 expression in the brain, suggesting that ankyrin3 suppression contributes to disease. However, the mechanism through which ankyrin3 confers risk is unknown. Ankyrin3 encodes the ankyrinG protein that tethers integral membrane proteins to the cytoskeleton. We previously reported that Ank3+/- heterozygous mice, which have ~50% reduced ankyrinG in the brain, exhibit behavioral changes reminiscent of bipolar mania (impulsivity, increased motivation for reward), and altered expression of proteins involved in neuron axonal transport. Both the behavioral and protein expression changes in Ank3+/- mice are normalized by lithium treatment, supporting the disease relevance of these findings. This study sought to further examine the molecular mechanism underlying the behavioral and neuronal changes induced by ankyrin3 suppression in brain. Methods We performed RNA sequencing of hippocampus from Ank3+/- heterozygous and Ank3+/+ wildtype mice, followed by identification of differentially expressed genes using the Tuxedo package. Biological pathways implicated by RNAseq analysis were verified by Western blot of hippocampal protein and live cell imaging of forebrain primary neurons. CRISPR/dCas9 technology was used to repress ankyrin3 transcription in a mouse neuronal system in order to conduct in-depth biochemical analysis. Neurons were treated with lithium and CHIR-99021, a selective inhibitor of the lithium target GSK3B, as well as an inhibitor of CRMP2, an axon-specific substrate of GSK3B. Results RNAseq identified 283 differentially expressed genes between Ank3+/- and Ank3+/+ mouse hippocampus (fold change>1.2, p 20% increased EB3, p 30%, p Discussion Ankyrin3 disruption in mice is associated with altered neuronal microtubule dynamics. Lithium reversal of both the microtubule and behavioral changes of Ank3+/- mice suggest that microtubule defects underlie manic-like behaviors observed in these mice. Our results support the investigation of risk genes using mouse and neuronal models to elucidate the neural mechanisms underlying psychiatric illness.

Summary Variants in the ANK3 gene encoding ankyrin-G are associated with neurodevelopmental disorders, including intellectual disability, autism, schizophrenia, and bipolar disorder. However, no upstream regulators of ankyrin-G at synapses are known. Here, we show that ankyrin-G interacts with Usp9X, a neurodevelopmental-disorder-associated deubiquitinase (DUB). Usp9X phosphorylation enhances their interaction, decreases ankyrin-G polyubiquitination, and stabilizes ankyrin-G to maintain dendritic spine development. In forebrain-specific Usp9X knockout mice (Usp9X–/Y), ankyrin-G as well as multiple ankyrin-repeat domain (ANKRD)-containing proteins are transiently reduced at 2 but recovered at 12 weeks postnatally. However, reduced cortical spine density in knockouts persists into adulthood. Usp9X–/Y mice display increase of ankyrin-G ubiquitination and aggregation and hyperactivity. USP9X mutations in patients with intellectual disability and autism ablate its catalytic activity or ankyrin-G interaction. Our data reveal a DUB-dependent mechanism of ANKRD protein homeostasis, the impairment of which only transiently affects ANKRD protein levels but leads to persistent neuronal, behavioral, and clinical abnormalities.

Brain ageing is thought to be related to geriatric depression, but the relationship between ageing and depression among middle aged individuals is unknown. The present study aimed to evaluate whether the age-related reduction of brain cortical thickness (CT) can be found in adult first-episode MDD patients, as well as to identify the possible genetic effect of the ANK3 gene polymorphism age-relates CT reduction. This study recruited 153 first-episode MDD patients with a disease duration < 2 years and 276 healthy controls (HC), and the CT of 68 whole brain regions and two ANK3 SNPs (rs1994336 and rs10994359) were analyzed. The results showed that although the CT of both groups was negative correlated with age, the MDD group had significant greater age-related decrease in CT than the HC group (–9.35 × 10−3 mm/year for MDD vs. –1.23 × 10−3 mm/year for HC in the left lateral orbitofrontal lobe). The multivariate analysis of covariance (MANCOVA) results yielded significant interactions of diagnosis × age, genotype × age and diagnosis × genotype interaction for rs10994359. In HC, the C allele showed a protective effect on age-related CT reduction. The reduction in CT with age was several times as greater in non-C carriers as in C carriers (–3.54 × 10−3 vs.–0.15 × 10−3 mm/year in left supramarginal gyrus) for HC. However, this protective effect disappeared in patients with MDD. We did not find a clear effect of rs1994336 on the age-related CT reduction. The findings indicate that the widespread accelerated brain ageing occurs early in adult-onset depression and this ageing may be a pathological mechanisms of depression rather than an outcome of the disease. The ANK3 rs10994359 polymorphism may partially affect regional cortical ageing in MDD.

Recent studies have shown that bipolar disorder (BPD) and schizophrenia (SZ) share some common genetic risk factors. This study aimed to examine the association between candidate single nucleotide polymorphisms (SNPs) identified from genome-wide association studies (GWAS) and risk of BPD and SZ. A total of 715 patients (244 BPD and 471 SZ) and 593 controls were genotyped using the Sequenom MassARRAY platform. We showed a positive association between LMAN2L (rs6746896) and risk of both BPD and SZ in a pooled population (P-value = 0.001 and 0.009, respectively). Following stratification by ethnicity, variants of the ANK3 gene (rs1938516 and rs10994336) were found to be associated with BPD in Malays (P-value = 0.001 and 0.006, respectively). Furthermore, an association exists between another variant of LMAN2L (rs2271893) and SZ in the Malay and Indian ethnic groups (P-value = 0.003 and 0.002, respectively). Gene–gene interaction analysis revealed a significant interaction between the ANK3 and LMAN2L genes (empirical P = 0.0107). Significant differences were shown between patients and controls for two haplotype frequencies of LMAN2L: GA (P = 0.015 and P = 0.010, for BPD and SZ, respectively) and GG (P = 0.013 for BPD). Our study showed a significant association between LMAN2L and risk of both BPD and SZ.

SummaryRecent evidence implicates glutamatergic synapses as key pathogenic sites in psychiatric disorders. Common and rare variants in the ANK3 gene, encoding ankyrin-G, have been associated with bipolar disorder, schizophrenia, and autism. Here we demonstrate that ankyrin-G is integral to AMPAR-mediated synaptic transmission and maintenance of spine morphology. Using superresolution microscopy we find that ankyrin-G forms distinct nanodomain structures within the spine head and neck. At these sites, it modulates mushroom spine structure and function, probably as a perisynaptic scaffold and barrier within the spine neck. Neuronal activity promotes ankyrin-G accumulation in distinct spine subdomains, where it differentially regulates NMDA receptor-dependent plasticity. These data implicate subsynaptic nanodomains containing a major psychiatric risk molecule, ankyrin-G, as having location-specific functions and open directions for basic and translational investigation of psychiatric risk molecules.

Background: The present study investigated associations between the strongest joint genetic risk variants for bipolar disorder (BD) and schizophrenia (SCZ) and a history of suicide attempt in patients with BD, SCZ and related psychiatric disorders. Methods: A history of suicide attempt was assessed in a sample of 1009 patients with BD, SCZ and related psychosis spectrum disorders, and associations with the joint genetic risk variants for BD and SCZ (rs2239547 (ITIH3/4-region), rs10994359 (ANK3) and rs4765905 (CACNA1 C)) were investigated. Previously reported susceptibility loci for suicide attempt in BD were also investigated. Associations were tested by logistic regression with Bonferroni correction for multiple testing. Results: The risk allele in rs2239547 (ITIH3/4-region) was significantly associated with a history of suicide attempt (p ¼ 0.01) after multiple testing correction (p threshold o 0.017). The previous suicide attempt susceptibility loci were only nominally associated, but had the same direction of risk in the replication sample (sign test, p ¼ 0.02). Limitations: Relatively small sample size and retrospective clinical assessment. Conclusions: We detected a novel association between suicide attempt and the ITIH3/4-region in a combined group of patients with BD, SCZ and related psychosis spectrum disorders. This may be useful in understanding molecular mechanisms of suicidal behaviour in severe mental disorders, although replication is warranted. Keywords: Suicidal behaviour; Bipolar disorder; Schizophrenia; Candidate gene (c) 2014 The Authors. Published by Elsevier B.V. Open access under CC BY-NC-ND license.

Background Major depressive disorder is a high-prevalence disease associated with a heavy burden on both personal well-being and socio-economical welfare, partly as a result of lacking tailored treatment options [1]. Common single nucleotide polymorphisms (SNPs) were estimated to account for 0.42 of the variance in antidepressant response [2], confirming the hypothesis that genetic polymorphisms may be used as effective markers to provide tailored antidepressant treatments. Candidate gene and genome-wide analyses can provide a complementary strategy, since the former can be applied to clarify the role of SNPs with high pre-test probability of association with the trait and the latter is useful to study the joint effects of a number of SNPs in a gene or a set of genes [3]. Aim We applied such a complementary strategy to the study of eight genes that are very strong candidates with previous evidence of pleiotropic effect across psychiatric traits. The genes of interest are involved in the regulation of neurotransmission (CACNA1C, CACNB2, ANK3), neural differentiation, synaptic plasticity, adhesion processes and structural organization (GRM7, TCF4, ITIH3, SYNE1) and glucocorticoid signaling (FKBP5). Methods Three samples with major depressive disorder (total n=671) were genotyped for 44 SNPs in strong candidate genes based on biological function and previous genome-wide association studies (CACNA1C, CACNB2, ANK3, GRM7, TCF4, ITIH3, SYNE1, FKBP5). Phenotypes were response/remission after 4 weeks of treatment and treatment-resistant depression (TRD: non response/non remission to at least two antidepressant treatments). Genome-wide data from STAR*D were used to replicate findings for response/remission (Level 1, n=1409) and TRD (Level 2, n=620). Pathways including the most promising candidate genes for involvement in TRD were investigated in STAR*D Level 2. Top pathway(s) were investigated using machine learning models. Results FKBP5 rs3800373, rs1360780 and rs9470080 showed replicated associations with response, remission or TRD. CACNA1C SNPs showed contradictory direction of association across samples. ANK3 rs1049862 AA genotype showed a replicated association with better outcome. In STAR*D the best pathway associated with TRD included CACNA1C (GO:0006942, permutated p=0.15). Neural networks and gradient boosted machine showed that independent SNPs in this pathway predicted TRD with a mean sensitivity of 0.83 and specificity of 0.56 after 10-fold cross validation repeated 100 times. Conclusions FKBP5 polymorphisms should be considered for inclusion in antidepressant pharmacogenetic tests. CACNA1C is a good candidate and GO:0006942 includes several genes coding for ion channels expressed in the central nervous system and other genes relevant for excitatory mechanisms. CACNB2 and ANK3 showed replicated associations with phenotypes and further investigations could help in clarifying their role. This study may pave the way to the identification of sets of genetic predictors in specific pathways able to predict the risk of TRD. It is reasonable to hypothesize a certain degree of variability in the genetic variants involved in TRD across different patients, but the involved pathways are expected to be more stable. Validated genetic markers of TRD could have a pivotal role in the implementation of targeted antidepressant treatments.

Summary Background The ankyrin 3 gene ( ANK3 ) produces the ankyrin G protein that plays an integral role in regulating neuronal activity. Previous studies have linked ANK3 to bipolar disorder and schizophrenia. A recent mouse study suggests that ANK3 may regulate behavioral disinhibition and stress reactivity. This led us to hypothesize that ANK3 might also be associated with stress-related psychopathology such as posttraumatic stress disorder (PTSD), as well as disorders of the externalizing spectrum such as antisocial personality disorder and substance-related disorders that are etiologically linked to impulsivity and temperamental disinhibition. Methods We examined the possibility of association between ANK3 SNPs and both PTSD and externalizing (defined by a factor score representing a composite of adult antisociality and substance abuse) in a cohort of white non-Hispanic combat veterans and their intimate partners ( n = 554). Initially, we focused on rs9804190—a SNP previously reported to be associated with bipolar disorder, schizophrenia, and ankyrin G expression in brain. Then we examined 358 additional ANK3 SNPs utilizing a multiple-testing correction. Results rs9804190 was associated with both externalizing and PTSD ( p = 0.028 and p = 0.042 respectively). Analysis of other ANK3 SNPs identified several that were more strongly associated with either trait. The most significant association with externalizing was observed at rs1049862 ( p = 0.00040, p corrected = 0.60). The most significant association with PTSD ( p = 0.00060, p corrected = 0.045) was found with three SNPs in complete linkage disequilibrium (LD)—rs28932171, rs11599164, and rs17208576. Conclusions These findings support a role of ANK3 in risk of stress-related and externalizing disorders, beyond its previous associations with bipolar disorder and schizophrenia.

Background Ankyrin 3 ( ANK3 ) has been strongly implicated as a risk gene for bipolar disorder (BD) by recent genome-wide association studies of patient populations. However, the genetic variants of ANK3 contributing to BD risk and their pathological function are unknown. Methods To gain insight into the potential disease relevance of ANK3 , we examined the function of mouse Ank3 in the regulation of psychiatric-related behaviors using genetic, neurobiological, pharmacological, and gene-environment interaction (G×E) approaches. Ank3 expression was reduced in mouse brain either by viral-mediated RNA interference or through disruption of brain-specific Ank3 in a heterozygous knockout mouse. Results RNA interference of Ank3 in hippocampus dentate gyrus induced a highly specific and consistent phenotype marked by decreased anxiety-related behaviors and increased activity during the light phase, which were attenuated by chronic treatment with the mood stabilizer lithium. Similar behavioral alterations of reduced anxiety and increased motivation for reward were also exhibited by Ank3+/– heterozygous mice compared with wild-type Ank3+/+ mice. Remarkably, the behavioral traits of Ank3 +/– mice transitioned to depression-related features after chronic stress, a trigger of mood episodes in BD. Ank3 +/– mice also exhibited elevated serum corticosterone, suggesting that reduced Ank3 expression is associated with elevated stress reactivity. Conclusions This study defines a new role for Ank3 in the regulation of psychiatric-related behaviors and stress reactivity that lends support for its involvement in BD and establishes a general framework for determining the disease relevance of genes implicated by patient genome-wide association studies.

Schizophrenia (SZ) and bipolar disorder (BPD) have high heritabilities and are clinically and genetically complex. Genome wide association studies (GWAS) and studies of copy number variations (CNV) in SZ and BPD have allowed probing of their underlying genetic risks. In this systematic review, we assess extant genetic signals from published GWAS and CNV studies of SZ and BPD up till March 2011. Risk genes associated with SZ at genome wide significance level (p value

Background The substantial contribution of genetic variants to inter-individual variance in antidepressant response makes genetic polymorphisms a unique opportunity to provide tailored antidepressant treatments. The present study combined a candidate approach of variants with strong previous evidence and a genome-wide approach in order to investigate the role of eight genes that appear optimal candidates for involvement in antidepressant response on the basis of their biological function and previous literature suggesting a pleiotropic effect across several psychiatric traits. Methods Three samples of patients with major depressive disorder (n=357, 218 and 96) were genotyped for 44 SNPs in genes involved in neurotransmission (CACNA1C, CACNB2, ANK3), neural differentiation, synaptic plasticity, adhesion processes, structural organization (GRM7, TCF4, ITIH3, SYNE1) and glucocorticoid signaling (FKBP5). Phenotypes were response/remission at week 4 and Treatment-Resistant Depression (TRD: non response/non remission to at least two antidepressants). STAR⁎D genome-wide data were used to replicate findings for response/remission (Level 1, n=1409) and TRD (Level 2, n=620). Standard quality control and genotype imputation were performed and pathways including the most promising candidate genes for involvement in TRD were investigated in STAR⁎D Level 2. Top pathway(s) were investigated also using machine learning (R cran Caret package). At SNP-level results with p Results Several FKBP5 SNPs (rs3800373, rs1360780, rs9470080) showed consistent associations with response, remission or TRD across at least two samples. Results involving CACNA1C SNPs (rs2283326, rs10848635, rs1006737) had contradictory direction of association across samples. ANK3 rs1049862 AA genotype showed a consistent association with a more favorable outcome in two samples. In STAR⁎D pathway analysis for the top candidate genes did not identify significant results after permutation. The best pathway associated with TRD included the CACNA1C gene (GO:0006942, regulation of striated muscle contraction, corrected p=0.15). Neural networks and gradient boosted machine showed that independent SNPs in this pathway predicted TRD with a mean accuracy of 0.73, sensitivity of 0.83 and specificity of 0.56. Discussion According to the present results and previous literature, FKBP5 polymorphisms should be considered for inclusion in pharmacogenetic tests for the prediction of antidepressant response and TRD. The contribution of CACNA1C polymorphisms was unclear, but further investigation is supported by previous literature and interesting findings for GO:0006942 pathway that includes several genes coding for ion channels that are expressed in the central nervous system as well as other genes relevant for excitatory mechanisms.

Background Bipolar Disorder (BD) is highly heterogeneous contributing to challenges in identifying genetic contributions to risk of illness. Narrowing the clinical phenotype may help identify risk genes. This study was designed to identify genetic components of psychotic mania in bipolar disorder, and potential genetic overlap between bipolar disorder with psychosis and schizophrenia. Methods We studied a Mayo Clinic Bipolar Disorder Biobank cohort of 695 BD type I (BD-I) cases and 777 controls. Of the 695 BD-I cases, 336 had a history of manic psychosis, and 309 had no history of psychosis. A Polygenic Risk Score (PRS) analysis was performed by using Psychiatric Genomics Consortium Schizophrenia genome-wide summary statistics as training dataset to create PRS. Schizophrenia PRS was then used to predict BD-I vs controls, BD-I with manic psychosis vs controls, BD-I without psychosis vs controls, and BD-I with manic psychosis vs BD-I without psychosis. A genome-wide analysis was performed comparing BD-I patients with a history of manic psychosis with BD-I cases with no history of psychosis. We also performed gene-level and pathway-level analyses using MAGMA in the same case-only sample. Results PRS analysis showed that the schizophrenia genetic risk score was more predictive of BD-I with manic psychosis than of BD-I without psychosis (BD-I with manic psychosis vs controls: p=1.1×10e-12, R2=6.6%; BD-I without psychosis: p=0.0001, R2=1.9%). Our GWAS showed no Single Nucleotide Polymorphisms (SNPs) with genome-wide significance; the top SNP was rs11126581 from the LRRTM4 gene (OR=2.17, p=1.08×10-6). LRRTM4 was also among the top genes in the gene-level analysis. The pathway analysis identified one significant pathway: the GO paranode region of the axon (p=0.008). Discussion Our results show that patients with schizophrenia are genetically more similar to bipolar patients who develop psychotic mania than those who never develop psychosis. This study was underpowered to detect SNPs associated with psychotic mania; however, the marginally significant association with LRRTM4 and the significant pathway, which contains ANK3, are potentially interesting findings. The LRRTM family and ANK3 have been previously implicated in schizophrenia and therefore could be involved in development of psychosis, regardless of schizophrenia or bipolar disorder distinction.

Background Over the last decade, a growing number of Genome-Wide Association Studies (GWAS) have been performed to identify common genetic variation that contributes to Bipolar Disorder (BD). These studies have identified a number of genes harboring common risk variants, and also demonstrated that BD has a highly polygenic genetic architecture, with many loci with small effects contributing to the disease. The contribution to risk of BD from rare variants is still not well understood, and whole exome and Whole Genome Sequencing (WGS) studies of BD are only beginning to emerge. Here, to investigate the role of rare variants with potentially greater effect sizes, we used WGS. Methods WGS was performed for 100 BD cases from the Mayo Clinic Bipolar Disorder Biobank and 1000 healthy controls from the Mayo Clinic Biobank. Following bioinformatics processing and genotype calling, Quality Control (QC) was performed. We excluded samples with low coverage or call rate, failed sex check, sample contamination or relatedness. We excluded variants with low call rate, failed HWE test, or failed Variant Quality Score Recalibration (VQSR). After QC, 99 cases and 961 controls with >26 million Single Nucleotide Variants (SNVs) with Minor Allele Frequency (MAF) ≤0.05 remained; sub-setting to annotated non-multiallelic variants within genes resulted in >11million SNVs for analysis. Rare variant gene-level tests were performed using two approaches: (1) burden test of nonsense mutations and missense mutations predicted to be deleterious by SIFT and PolyPhen2; (2) SNP-set kernel association test (SKAT) of all variants with MAF weights. We a priori selected 7 genes implicated in BD by GWAS for prioritized analysis: CACNA1C, ODZ4, ANK3, TRANK1, LMAN2L, NCAN, and SYNE1. Results After restricting analyses to genes in which ≥4 subjects carried variant alleles, 8,242 genes were analyzed by the burden test, and 25,833 genes were analyzed using SKAT. Neither of these gene-level testing approaches identified genome-wide significant associations. Among the previously implicated candidate genes, only ANK3 (p=0.011) and SYNE1 (p=0.006) were associated with BD at a nominally significant level in the burden and SKAT analyses, respectively. In ANK3, the 9 SNVs analyzed by the burden test were observed in 6/99 cases and 18/961 controls; the association result was driven primarily by rs148904927, a SNV that was observed in 5 (5%) cases and 7 (0.7%) controls [OR (95%CI)=7.2 (2.3–23.3)]. Discussion This pilot WGS study suggests that rare variants in ANK3 and SYNE1 may contribute to BD. The ANK3 SNV (rs148904927) that we observed in 5% of the cases (and 0.7% of controls) is a non-synonymous variant (Ser2409Pro) that may function by modifying the isoforms of ANK3 (Ankyrin G), an integral membrane protein that is modulated during developmental stages and is associated with several neuronal dysfunction disorders. According to HaploReg v4.1, Regulome DB, and the UCSC genome browser, this SNV is highly conserved and disrupts multiple binding factor motifs. This SNV is located in the most prominent candidate regulatory element of ANK3 in a DNAse hypersensitivity site enriched with regulatory histone mark (H3K27Ac), both features often associated with regulatory elements. This regulatory element overlaps a large alternative splice coding exon, suggesting that the variant may operate via regulation of transcription levels or isoform. Replication and functional studies are warranted to further investigate the role of this variant.

Background Bipolar Disorder (BD) is a common, complex and severe mood disorder with high heritability. Genetic analyses of bipolar disorder continue to benefit from large international cohort collections amassed by members of the Psychiatric Genomics Consortium. Here we describe insights into disease risk from new and existing BD risk loci and from polygenic analyses of BD and related diseases and traits. Methods We performed GWAS meta-analysis of 32 cohorts from 14 countries in Europe and North America, totaling 20,352 cases and 31,358 controls of European descent, including 6,328 case and 7,963 control samples not previously reported. We tested suggestive SNPs (P Results Bipolar disorder GWAS identified 31 genome-wide significant (P BD2, P BD1, P Discussion The PGC2 Bipolar Disorder GWAS identified twenty novel risk loci. Single gene loci include the synaptic genes ANK3, GRIN2A, SHANK2 and RIMS1. Polygenic analyses reaffirm the striking polygenicity of bipolar disorder and close relationships with schizophrenia and other psychiatric disorders, and evince an affective-to-mood disorder gradient of genetic overlap from schizophrenia, to BDI, to BDII, to major depression. Bipolar disorder's positive genetic correlation with cognitive/education phenotypes stands in contrast to schizophrenia and major depression, suggesting an inroad to further dissection of genetic components specific to these major psychiatric disorders. Genetic overlaps of BD with lipid-, osteoarthritis- and immune-related traits, provide additional intriguing clinical and physiological implications.

We have characterized a new ankyrin gene, expressed in brain and other tissues, that is subject to extensive tissue-specific alternative mRNA processing. The full-length polypeptide has a molecular mass of 480 kDa and includes a predicted globular head domain, with membrane- and spectrin-binding activities, as well as an extended "tail" domain. We term this gene ankyrinG based on its giant size and general expression. Two brain-specific isoforms of 480 kDa and 270 kDa were identified that contain a unique stretch of sequence highly enriched in serine and threonine residues immediately following the globular head domain. Antibodies against the serine-rich domain and spectrin-binding domain revealed labeling of nodes of Ranvier and axonal initial segments. Ankyrin-binding proteins also known to be localized in these specialized membrane domains include the voltage-dependent sodium channel, the sodium/potassium ATPase, sodium/calcium exchanger, and members of the neurofascin/L1 family of cell adhesion molecules. The neural-specific ankyrinG polypeptides are candidates to participate in maintenance/targeting of ion channels and cell adhesion molecules to nodes of Ranvier and axonal initial segments.

Background: The gene ANK3 is implicated in bipolar disorder and schizophrenia. The present study investigated the influence of this gene on cognitive performance and brain structure among individuals with first-episode psychosis (FEP). The brief illness duration of an FEP sample makes it well suited for studying the effects of genetic variation. Methods: We genotyped 2 single nucleotide polymor phisms (SNPs; rs1938526 and rs10994336) in ANK3 in patients with FEP. Multivariate analysis of variance compared risk allele carriers and noncarriers on 6 domains of cognition consistent with MATRICS consensus. A subsample of 82 patients was assessed using mag netic resonance imaging. We compared brain structure between carriers and noncarriers using cortical thickness analysis and voxelbased morphometry on white matter. Results: In the 173 patients with FEP included in our study, rs1938526 and rs10994336 were in very high linkage disequilibrium (d' = 0.95), and analyses were therefore only carried out on the SNP (rs1938526) with the highest minor allele frequency (G). Allele G of rs1938526, was associated with lower cognitive performance across domains ( F6,164 = 2.38, p = 0.030) and significantly lower scores on the domains of verbal memory ( p = 0.015), working memory ( p = 0.006) and attention ( p = 0.019). The significant effects of this SNP on cognition were not maintained when controlling for IQ. Cortical thinning was observed in risk allele carri ers at diverse sites across cortical lobes bilaterally at a threshold of p < 0.01, false discovery rate‐corrected. Risk-allele carriers did not show any regions of reduced white matter volume. Limitations: The sample size is modest given that a low-frequency variant was being examined. Conclusion: The ANK3 risk allele rs1938526 appears to be associated with general cognitive impairment and widespread cortical thinning in patients with FEP.

The ankyrin3 gene encodes a novel form of ankyrin, AnkyrinG, expressed in multiple tissues but characteristically present at the axonal initial segment and nodes of Ranvier of neurons in the central and peripheral nervous systems. We have localized ANK3 to human Chromosome 10q21 by fluorescence in situ hybridization. The position of the murine homologue was determined by intersubspecific backcross analysis, mapping Ank3 to mouse Chromosome 10, between microsatellite marker D10Mit31 and the Bcr gene. This interval is known to comprise a region syntenic to human 10q. The localization of ANK3 is a preliminary step in identifying neurological disorders potentially associated with the gene.