Calcium-dependent Cell Adhesion Research Articles

N-Cadherin — A Potential Target for Psychopharmacology

Glycoprotein N-cadherin (Neuronal cadherin) belongs to the family of calcium-dependent cell adhesion molecules, representing a key element that carries out intercellular contacts in brain neurons. However, it is involved not only in the mechanical connection of neurons, but also influences the specifics of the further development and functional state of the neuron. This is due to the active interaction of N-cadherin with many proteins at the pre- and post-synapse, initiating a cascade of reactions that provide such processes as long-term potentiation (underlying learning and memory), morphogenesis, neuronal recognition, activation of receptors (NMDA and AMPA types), regulation of cytoskeleton formation. This polyfunctionality is necessary for specific neurons to connect to each other in a certain way, and such adhesion leads to the coordination of cell behavior through intercellular signaling and spatio–temporal control of differential gene expression. Mutations in the genes responsible for the expression of N-cadherin lead to various disorders of the functional activity of the synapse and the processes of spatial orientation and memory. Thus, involvement in important neuroplastic processes regulating cognitive functions and behavior determines interest in studying the effect of drugs on N-cadherin. In particular, N-cadherin deserves closer consideration by pharmacologists as a potential target in the mechanism of action of various psychoactive substances.

Trafficking of adhesion and aggregation-modulating proteins during the early stages of Dictyostelium development

The social amoeba Dictyostelium discoideum has been studied for close to a century to better understand conserved cellular and developmental processes. The life cycle of this model eukaryote is composed of a unicellular growth phase and a multicellular developmental phase that is induced by starvation. When starved, individual cells undergo chemotactic aggregation to form multicellular mounds that develop into slugs. Terminal differentiation of cells within slugs forms fruiting bodies, each composed of a stalk that supports a mass of viable spores that germinate and restart the life cycle when nutrients become available. Calcium-dependent cell adhesion protein A (CadA) and countin (CtnA) are two proteins that regulate adhesion and aggregation, respectively, during the early stages of D. discoideum development. While the functions of these proteins have been well-studied, the mechanisms regulating their trafficking are not fully understood. In this study, we reveal pathways and cellular components that regulate the intracellular and extracellular amounts of CadA and CtnA during aggregation. During growth and starvation, CtnA localizes to cytoplasmic vesicles and punctae. We show that CtnA is glycosylated and this post-translational modification is required for its secretion. Upon autophagy induction, a signal peptide for secretion facilitates the release of CtnA from cells via a pathway involving the μ subunit of the AP3 complex (Apm3) and the WASP and SCAR homolog, WshA. Additionally, CtnA secretion is negatively regulated by the D. discoideum orthologs of the human non-selective cation channel mucolipin-1 (Mcln) and sorting receptor sortilin (Sort1). As for CadA, it localizes to the cell periphery in growth-phase and starved cells. The intracellular and extracellular amounts of CadA are modulated by autophagy genes (atg1, atg9), Apm3, WshA, and Mcln. We integrate these data with previously published findings to generate a comprehensive model summarizing the trafficking of CadA and CtnA in D. discoideum. Overall, this study enhances our understanding of protein trafficking during D. discoideum aggregation, and more broadly, provides insight into the multiple pathways that regulate protein trafficking and secretion in all eukaryotes.

Cadherin-17 as a target for the immunoPET of adenocarcinoma

PurposeCadherin-17 (CDH17) is a calcium-dependent cell adhesion protein that is overexpressed in several adenocarcinomas, including gastric, colorectal, and pancreatic adenocarcinoma. High levels of CDH17 have been linked to metastatic disease and poor prognoses in patients with these malignancies, fueling interest in the protein as a target for diagnostics and therapeutics. Herein, we report the synthesis, in vitro validation, and in vivo evaluation of a CDH17-targeted 89Zr-labeled immunoPET probe.MethodsThe CDH17-targeting mAb D2101 was modified with an isothiocyanate-bearing derivative of desferrioxamine (DFO) to produce a chelator-bearing immunoconjugate — DFO-D2101 — and flow cytometry and surface plasmon resonance (SPR) were used to interrogate its antigen-binding properties. The immunoconjugate was then radiolabeled with zirconium-89 (t1/2 ~ 3.3 days), and the serum stability and immunoreactive fraction of [89Zr]Zr-DFO-D2101 were determined. Finally, [89Zr]Zr-DFO-D2101’s performance was evaluated in a trio of murine models of pancreatic ductal adenocarcinoma (PDAC): subcutaneous, orthotopic, and patient-derived xenografts (PDX). PET images were acquired over the course of 5 days, and terminal biodistribution data were collected after the final imaging time point.ResultsDFO-D2101 was produced with a degree of labeling of ~ 1.1 DFO/mAb. Flow cytometry with CDH17-expressing AsPC-1 cells demonstrated that the immunoconjugate binds to its target in a manner similar to its parent mAb, while SPR with recombinant CDH17 revealed that D2101 and DFO-D2101 exhibit nearly identical KD values: 8.2 × 10−9 and 6.7 × 10−9 M, respectively. [89Zr]Zr-DFO-D2101 was produced with a specific activity of 185 MBq/mg (5.0 mCi/mg), remained >80% stable in human serum over the course of 5 days, and boasted an immunoreactive fraction of >0.85. In all three murine models of PDAC, the radioimmunoconjugate yielded high contrast images, with high activity concentrations in tumor tissue and low uptake in non-target organs. Tumoral activity concentrations reached as high as >60 %ID/g in two of the cohorts bearing PDXs.ConclusionTaken together, these data underscore that [89Zr]Zr-DFO-D2101 is a highly promising probe for the non-invasive visualization of CDH17 expression in PDAC. We contend that this radioimmunoconjugate could have a significant impact on the clinical management of patients with both PDAC and gastrointestinal adenocarcinoma, most likely as a theranostic imaging tool in support of CDH17-targeted therapies.

Study of single nucleotide polymorphisms and serum level of E-cadherin and their relation to psoriasis severity

Backgroundpsoriasis is a persistent autoimmune-mediated systemic inflammatory illness that affects the skin and several organ systems. A family of calcium-dependent cell adhesion molecules called cadherins (CDH) works through homophilic contacts to keep cells attached to one another. In comparison to normal skin, several investigations found that the expression of E-cadherin was downregulated in the granular and basal layers of psoriasis lesional skin. ObjectivesThis study aimed to relate the serum level of E-cadherin and its genes (rs17715799 and rs13689) SNPs with the severity of psoriasis. MethodsIn the current study, 120 participants were divided into two groups: 60 patients with plaque psoriasis were in group I, and 60 healthy controls were in group II. The serum level of E-cadherin protein was measured using an enzyme linked immunosorbent assay (ELISA), and SNPs in the E-cadherin gene (rs17715799 and rs13689) were identified using real-time polymerase chain reaction (RT-PCR) technology. ResultsIn our study group I had a considerably higher serum level of the protein E-cadherin. It was confirmed that the E cadherin (rs13689) genotype frequency distribution revealed a significant difference between the two studied groups. The frequency of the CC genotype was higher in group I. While there was no significant variation in the genotype frequency distribution of E-cadherin (rs17715799) between the two study groups under the dominant, over-dominant, and recessive modes of inheritance (p 0.204) conclusion: patients with psoriasis had considerably higher serum levels of E-cadherin. The E-cadherin gene rs13689 polymorphism affects the chance of developing psoriasis. However, the E-cadherin gene rs17715799 polymorphism has no impact on the onset of psoriasis. As a result, the E-cadherin gene rs13689 but not rs17715799 might be considered as a marker for the risk of developing psoriasis.

Heterophilic and homophilic cadherin interactions in intestinal intermicrovillar links are species dependent.

Enterocytes are specialized epithelial cells lining the luminal surface of the small intestine that build densely packed arrays of microvilli known as brush borders. These microvilli drive nutrient absorption and are arranged in a hexagonal pattern maintained by intermicrovillar links formed by 2 nonclassical members of the cadherin superfamily of calcium-dependent cell adhesion proteins: protocadherin-24 (PCDH24, also known as CDHR2) and the mucin-like protocadherin (CDHR5). The extracellular domains of these proteins are involved in heterophilic and homophilic interactions important for intermicrovillar function, yet the structural determinants of these interactions remain unresolved. Here, we present X-ray crystal structures of the PCDH24 and CDHR5 extracellular tips and analyze their species-specific features relevant for adhesive interactions. In parallel, we use binding assays to identify the PCDH24 and CDHR5 domains involved in both heterophilic and homophilic adhesion for human and mouse proteins. Our results suggest that homophilic and heterophilic interactions involving PCDH24 and CDHR5 are species dependent with unique and distinct minimal adhesive units.

The effect of the content of ApoA-I in peripheral blood on the state of immune homeostasis in people living in extreme climatic conditions of the Arctic.

The paper presents data on the impacts of the content of ApoA-I in peripheral blood on the state of immune homeostasis in people living in extreme climatic conditions of the Arctic. From the village of Revda, Murmansk region, 191 people were examined, 160 women and 31 men, aged from 21 to 55 years. The analysis of the results was carried out depending on the level of ApoA-I content: 111 examined people had a low content of ApoA-I (< 115 mg/dl) and 80 people had a concentration of ApoA-I - within the physiological norm (115-220 mg/dl). Deficiency of ApoA-I is associated with an increase in plasma concentrations of total cholesterol in 37.5% and triglycerides in 62.5% of cases. Low content of ApoA-I is associated with a decrease in the content of neutrophilic granulocytes and an increase in the concentration of small lymphocytes, IL-10 and IgE. Deficiency of ApoA-I is associated with an increase in the content of CD45RA+, CD16+, CD56+ lymphocyte phenotypes in blood. Low concentrations of ApoA-I are associated with low expression activity of L-selectin gene and free L-selectin ligand. Due to deficiency of ApoA-I, no significant differences were detected in the content of transferrin, free transferrin receptor, haptoglobin, free calcium-dependent cell adhesion protein, and free pool of receptors involved in apoptosis. Conclusion: deficiency of ApoA-I in blood plasma is associated with increased migration of neutrophils into the tissue and cell-mediated cytotoxicity of lymphocytes and occurs because of the effect of decreased activity of the level of antioxidant defense, changes in the structure of cell membranes and the participation of adhesion molecules.

PCDH19 interplay with GABA(A) receptors: a window to DEE9 pathogenetic mechanisms.

PCDH19 interplay with GABA(A) receptors: a window to DEE9 pathogenetic mechanisms.

Crystal structure of the nonclassical cadherin-17 N-terminus and implications for its adhesive binding mechanism.

The cadherin superfamily of calcium-dependent cell-adhesion proteins has over 100 members in the human genome. All members of the superfamily feature at least a pair of extracellular cadherin (EC) repeats with calcium-binding sites in the EC linker region. The EC repeats across family members form distinct complexes that mediate cellular adhesion. For instance, classical cadherins (five EC repeats) strand-swap their N-termini and exchange tryptophan residues in EC1, while the clustered protocadherins (six EC repeats) use an extended antiparallel `forearm handshake' involving repeats EC1-EC4. The 7D-cadherins, cadherin-16 (CDH16) and cadherin-17 (CDH17), are the most similar to classical cadherins and have seven EC repeats, two of which are likely to have arisen from gene duplication of EC1-2 from a classical ancestor. However, CDH16 and CDH17 lack the EC1 tryptophan residue used by classical cadherins to mediate adhesion. The structure of human CDH17 EC1-2 presented here reveals features that are not seen in classical cadherins and that are incompatible with the EC1 strand-swap mechanism for adhesion. Analyses of crystal contacts, predicted glycosylation and disease-related mutations are presented along with sequence alignments suggesting that the novel features in the CDH17 EC1-2 structure are well conserved. These results hint at distinct adhesive properties for 7D-cadherins.

Clinicopathological significance and prognostic value of E-cadherin expression in non-small cell lung cancer: A protocol for systematic review and meta-analysis.

Background:E-cadherin, a calcium-dependent cell adhesion molecule, as an important mediator of adhesion and signaling pathway, plays a key role in maintaining tissue integrity. However, the association of E-cadherin expression with clinicopathological features and prognostic value in non-small cell lung cancer (NSCLC) is still controversial. Therefore, the purpose of the study is to explore the clinicopathological features and prognostic value of E-cadherin expression in non-small cell lung cancer by meta-analysis.Methods:PubMed, EMBASE, Cochrane Library, and Web of Science were searched to collect the studies about expression of E-cadherin and clinicopathological features and prognosis of non-small cell lung cancer. The last search time was May 2020. Stata 15.0 software was used for statistical analysis.Results:A total of 35 studies were included, of which the results showed that high expression of E-cadherin compared with its low expression, for overall survival, HR = 0.68 (95% CI:0.64–0.73, P < .05); for disease-free survival or progression-free survival, HR = 0.54 (95% CI: 0.44–0.67); low differentiation of lung cancer compared with moderate and high differentiation, OR = 0.40 (95% CI: 0.27–0.58, P < .05); Advanced lung cancer compared with early stage, OR = 0.54 (95% CI: 0.44–0.66, P < .05); lymph node metastasis compared with non-lymph node metastasis, OR = 0.49 (95% CI: 0.31∼0.77).Conclusion:Low expression of E-cadherin is closely related to poor prognosis of patients with NSCLC, promoting tumor staging and lymph node metastasis, inhibiting tumor differentiation as well.

E-cadherin immunohistochemical expression in gastric carcinoma in relation to different clinicopathological parameters

Background Gastric cancer (GC) is an aggressive malignancy and a common cause of cancer-related deaths worldwide. E-cadherin is a transmembrane calcium-dependent epithelial cell adhesion molecule. Loss of E-cadherin-mediated cell–cell adhesion is needed for tumor cell invasion and metastasis. Cells with mutated E-cadherin demonstrate increased motility, show altered organization of their actin cytoskeleton, and are associated with metastases, thereby providing evidence for its role as an invasion suppressor. Objectives To assess the prognostic value of E-cadherin expression in GC and to evaluate its correlations with other prognostic parameters. Materials and methods Immunohistochemical E-cadherin expression was studied in 36 cases of GC (24 cases of endoscopic biopsies and 12 cases of total gastrectomy), and the results were correlated with clinicopathological parameters. Results E-cadherin was positive in 15/36 (41.7%) cases of GC. A significant inverse correlation was found between E-cadherin expression and tumor grade, number of lymph nodes involved by metastasis, and Helicobacter pylori positivity, with P values of 0.02, 0.04, and 0.03, respectively. Conclusion E-cadherin loss plays an important role in invasion and metastasis potential in GC, and decrease or loss of its expression is predictive of poor prognosis. H. pylori plays an important role in gastric carcinogenesis, and it induces a statistically significant decrease in E-cadherin expression, so eradication of H. pylori infection can stop or decrease the progression of carcinogenesis.

Immunohistochemical Expression of E-cadherin in Invasive Duct Carcinoma of the Breast

Breast cancer is the most frequent malignant tumor in women worldwide. E-Cadherin is a calcium-dependent epithelial cell adhesion molecule that is expressed at adherens junctions. Its loss may result in a poorly differentiated tumorphenotype. Cells with mutated E-cadherin demonstrate increased motility, altered organization of their actin cytoskeletonand associated with metastases, thereby providing evidence for its role as an invasion suppressor.Objectives: Assess the prognostic value of E-cadherin expression in invasive duct carcinoma (IDC) of the breastand to evaluate its correlations with other prognostic parameters.Materials and Methods: Immunohistochemical (IHC) E-Cadherin expression was studied in 60 IDC and the results were correlated with the clinico-pathological parameters.Results: E-cadherin was positive in 46/60 (76.7%) of IDC; 15/60 (25%) showed strong membranous staining. A significant inverse correlation was found between E-cadherin expression and number of lymph node metastasis.Conclusions: Loss of E-cadherin expression is a predictive of invasion and metastasis in IDC.

Cell Migration Guided by Cell-Cell Contacts in Innate Immunity.

The directed migration of leukocytes to sites of damage or infection is necessary for a productive immune response. There is substantial evidence supporting a key role for chemoattractants in directed migration, however, less is known about how cell-cell contacts affect the migratory behavior of leukocytes in innate immunity. Here, we explore how cell-cell contacts can affect the directed migration of innate immune cells, including their role in attracting, repelling, or stopping cell motility. Further investigation of cell contact dynamics as guidance cues may yield new insights into the regulation of innate immunity.

Quantitative MRI-Based Analysis Identifies Developmental Limbic Abnormalities in PCDH19 Encephalopathy.

Protocadherin-19 (PCDH19) is a calcium dependent cell-adhesion molecule involved in neuronal circuit formation with prevalent expression in the limbic structures. PCDH19-gene mutations cause a developmental encephalopathy with prominent infantile onset focal seizures, variably associated with intellectual disability and autistic features. Diagnostic neuroimaging is usually unrevealing. We used quantitative MRI to investigate the cortex and white matter in a group of 20 PCDH19-mutated patients. By a statistical comparison between quantitative features in PCDH19 brains and in a group of age and sex matched controls, we found that patients exhibited bilateral reductions of local gyrification index (lGI) in limbic cortical areas, including the parahippocampal and entorhinal cortex and the fusiform and lingual gyri, and altered diffusivity features in the underlying white matter. In patients with an earlier onset of seizures, worse psychiatric manifestations and cognitive impairment, reductions of lGI and diffusivity abnormalities in the limbic areas were more pronounced. Developmental abnormalities involving the limbic structures likely represent a measurable anatomic counterpart of the reduced contribution of the PCDH19 protein to local cortical folding and white matter organization and are functionally reflected in the phenotypic features involving cognitive and communicative skills as well as local epileptogenesis.

Farelerde erken embriyo gelişiminde e-kadherin ekspresyonu

E-cadherin is a member of the cadherin superfamily of calcium-dependent cell adhesion molecules, and has critical roles in cell adhesion during early embryo development. To determine the expression and distribution of E-cadherin, we applied immunofluorescent staining on late 8 cell, morula and blastocyst stage embryos during the experiments. Firstly, we superovulated the females and sacrificed them to isolate oviducts. We flushed 2 cell embryos by using FHM HEPES-buffered medium from the CD1 female mouse oviducts and cultured them in KSOM medium until the blastocyst stage. Embryos were fixed and immunostained with E-cadherin, then images were assessed using a fluorescence microscope. We demonstrate that E-cadherin is expressed throughout the late 8 cell to fully expanded blastocysts. We observed that E-cadherin is present only in the cell-cell contact sites and not in the apical sites of the membrane of late 8 cell stage embryo. In morula, E-cadherin localized on the surface of blastomeres and cell-cell junctions. In blastocyst stage, embryos showed membrane staining in cell-cell contact regions in trophectoderm cells while localized on the surface of inner cell mass. Our results indicate that E-cadherin mediated cell adhesion has major roles on compaction, trophectoderm epithelial differentation and implantation.

A study combining whole-exome sequencing and structural neuroimaging analysis for major depressive disorder

BackgroundGenetic variations associated with major depressive disorder (MDD) may affect the structural aspects of neural networks mediated by the molecular pathways involved in neuronal survival and synaptic plasticity. However, few studies have applied a novel approach such as whole-exome sequencing (WES) analysis to investigate the genetic contribution to the neurostructural changes in MDD. MethodsIn the first part of the study, we investigated rare variants of selected genes from previous WES studies using a WES analysis in 184 patients with MDD and 82 healthy controls. In the second part of the study, we explored the association between the common genetic variants from the WES analysis and cortical thickness in 91 patients with MDD and 75 healthy controls. The gray-matter thickness of each cortical region was measured using FreeSurfer. ResultsWe identified recurrent non-silent variants in 24 MDD-related genes including FASN, MYH13, UNC13D, LILRA1, CACNA1B, TRIO, HOMER3, and BCAR3, and observed eleven recurrently altered copy number alternations where a gain on 15q11.2 and losses on 7q34 and 15q11.1-q11.2 in MDD genomes. We also found that rs11592462 in CDH23, a calcium-dependent cell-adhesion molecule encoding gene, was significantly associated with thinning in the right anterior cingulate cortex. LimitationThe small sample size may lead our findings to be underpowered regarding rare variants. Conclusion: The present study identified that non-synonymous rare variants were significantly associated with risk of MDD and found that genetic contributions to the development of MDD may be mediated by alterations in cortical thickness of emotion-processing neural circuits.

Structure-Based Virtual Screening Allows the Identification of Efficient Modulators of E-Cadherin-Mediated Cell-Cell Adhesion.

Cadherins are a large family of transmembrane calcium-dependent cell adhesion proteins that orchestrate adherens junction formation and are crucially involved in tissue morphogenesis. Due to their important role in cancer development and metastasis, cadherins can be considered attractive targets for drug discovery. A recent crystal structure of the complex of a cadherin extracellular portion and a small molecule inhibitor allowed the identification of a druggable interface, thus providing a viable strategy for the design of cadherin dimerization modulators. Here, we report on a structure-based virtual screening approach that led to the identification of efficient and selective modulators of E-cadherin-mediated cell–cell adhesion. Of all the putative inhibitors that were identified and experimentally tested by cell adhesion assays using human pancreatic tumor BxPC-3 cells expressing both E-cadherin and P-cadherin, two compounds turned out to be effective in inhibiting stable cell–cell adhesion at micromolar concentrations. Moreover, at the same concentrations, one of them also showed anti-invasive properties in cell invasion assays. These results will allow further development of novel and selective cadherin-mediated cell–cell adhesion modulators for the treatment of a variety of cadherin-expressing solid tumors and for improving the efficiency of drug delivery across biological barriers.

The cadherin-catenin complex is necessary for cell adhesion and embryogenesis in Nematostella vectensis

The cadherin-catenin complex is a conserved, calcium-dependent cell-cell adhesion module that is necessary for normal development and the maintenance of tissue integrity in bilaterian animals. Despite longstanding evidence of a deep ancestry of calcium-dependent cell adhesion in animals, the requirement of the cadherin-catenin complex to coordinate cell-cell adhesion has not been tested directly in a non-bilaterian organism. Here, we provide the first analysis of classical cadherins and catenins in the Starlet Sea Anemone, Nematostella vectensis. Gene expression, protein localization, siRNA-mediated knockdown of α-catenin, and calcium-dependent cell aggregation assays provide evidence that a bonafide cadherin-catenin complex is present in the early embryo, and that α-catenin is required for normal embryonic development and the formation of cell-cell adhesions between cells dissociated from whole embryos. Together these results support the hypothesis that the cadherin-catenin complex was likely a complete and functional cell-cell adhesion module in the last common cnidarian-bilaterian ancestor. Summary statementEmbryonic manipulations and ex vivo adhesion assays in the sea anemone, Nematostella vectensis, indicate that the necessity of the cadherin-catenin complex for mediating cell-cell adhesion is deeply conserved in animal evolution.

SA27 - EXOME SEQUENCING OF MULTIPLY AFFECTED BIPOLAR DISORDER FAMILIES AND FOLLOW-UP RESEQUENCING IMPLICATE RARE VARIANTS IN NEURONAL GENES CONTRIBUTING TO DISEASE ETIOLOGY

Background Bipolar Disorder (BD) is a complex neuropsychiatric disorder characterized by recurrent episodes of mania and depression. It is a severe disorder of mood with a lifetime prevalence of about 1% and a high heritability of about 70%. Models of illness are most consistent with a polygenic contribution of common and rare variants to disease susceptibility. As the cumulative impact of common alleles with small effect may only explain around 25–38% of the phenotypic variance for BD, rare variants of high penetrance have been suggested to contribute to BD susceptibility. One way to evaluate this hypothesis is to investigate large pedigrees densely affected with BD, in which the existence of a genetic variant of strong effect inherited from a common ancestor may be more likely. Methods In the present study, we investigated 226 individuals of 68 large multiply affected families of European origin using Whole Exome Sequencing (WES). In each family, two to five affected individuals with BD or recurrent major depression were selected for sequencing. WES was performed on the Illumina HiSeq. 2500 platform and the Varbank pipeline of the Cologne Center for Genomics was used for data analysis. All identified variants shared within each family were filtered for a minor allele frequency Results We identified a total of 1214 rare, segregating and functional variants implicating 1122 different genes, of which 903 were brain expressed. Subsequently, we applied the Residual Variation Intolerance Scores (Petrovski et al., 2013) and identified 294 genes that were ranked among the 20% most “intolerant” genes in the genome. Gene enrichment analysis of these genes showed a significant enrichment for 18 pathways (p We prioritized genes that were (i) found in at least two unrelated families in the present study, (ii) previously reported in next generation sequencing or GWAS studies of BD, or (iii) predominantly driving the significant pathways in our gene enrichment analysis. The different approaches of prioritization yielded 42 promising candidate genes including CDH22 which encodes a calcium-dependent cell adhesion protein that plays an important role in the morphogenesis of neural cells during brain development. Discussion Our preliminary results strongly suggest that rare and highly-penetrant variants in neuronal and cell adhesion genes contribute to BD etiology. The 42 prioritized candidate genes are currently being followed up by resequencing in cohorts of about 2500 independent BD cases and 2500 controls of European ancestry. For resequencing, we use the single molecule molecular inversion probes (smMIPs) technology that enables multiplex targeted resequencing in large cohorts. The smMIPs sequences have been designed with the empirically trained design algorithm MIPgen (Boyle et al., 2014) and sequencing is currently performed on the Illumina HiSeq. 2500 platform. The resequencing results will provide further insights into the particular genes and pathways which are involved in BD development.

Differential Methylation of H3K79 Reveals DOT1L Target Genes and Function in the Cerebellum In Vivo

The disruptor of telomeric silencing 1-like (DOT1L) mediates methylation of histone H3 at position lysine 79 (H3K79). Conditional knockout of Dot1l in mouse cerebellar granule cells (Dot1l-cKOAtoh1) led to a smaller external granular layer with fewer precursors of granule neurons. Dot1l-cKOAtoh1 mice had impaired proliferation and differentiation of granular progenitors, which resulted in a smaller cerebellum. Mutant mice showed mild ataxia in motor behavior tests. In contrast, Purkinje cell-specific conditional knockout mice showed no obvious phenotype. Genome-wide transcription analysis of Dot1l-cKOAtoh1 cerebella using microarrays revealed changes in genes that function in cell cycle, cell migration, axon guidance, and metabolism. To identify direct DOT1L target genes, we used genome-wide profiling of H3K79me2 and transcriptional analysis. Analysis of differentially methylated regions (DR) and differentially expressed genes (DE) revealed in total 12 putative DOT1L target genes in Dot1l-cKOAtoh1 affecting signaling (Tnfaip8l3, B3galt5), transcription (Otx1), cell migration and axon guidance (Sema4a, Sema5a, Robo1), cholesterol and lipid metabolism (Lss, Cyp51), cell cycle (Cdkn1a), calcium-dependent cell-adhesion or exocytosis (Pcdh17, Cadps2), and unknown function (Fam174b). Dysregulated expression of these target genes might be implicated in the ataxia phenotype observed in Dot1l-cKOAtoh1.

Early-life stress impairs developmental programming in Cadherin 13 (CDH13)-deficient mice

ObjectiveCadherin-13 (CDH13), a member of the calcium-dependent cell adhesion molecule family, has been linked to neurodevelopmental disorders, including autism spectrum (ASD) and attention-deficit/hyperactivity (ADHD) disorders, but also to depression. In the adult brain, CDH13 expression is restricted e.g. to the presynaptic compartment of inhibitory GABAergic synapses in the hippocampus and Cdh13 knockout mice show an increased inhibitory drive onto hippocampal CA1 pyramidal neurons, leading to a shift in excitatory/inhibitory balance. CDH13 is also moderating migration of serotonergic neurons in the dorsal raphe nucleus, establishing projections preferentially to the thalamus and cerebellum during brain development. Furthermore, CDH13 is upregulated by chronic stress as well as in depression, suggesting a role in early-life adaptation to stressful experience. Here, we therefore investigated the interaction between Cdh13 variation and neonatal maternal separation (MS) in mice. MethodsMale and female wild-type (Cdh13+/+), heterozygous (Cdh13+/−) and homozygous (Cdh13−/−) knockout mice exposed to MS, or daily handling as control, were subjected to a battery of behavioural tests to assess motor activity, learning and memory as well as anxiety-like behaviour. A transcriptome analysis of the hippocampus was performed in an independent cohort of mice which was exposed to MS or handling, but remained naïve for behavioural testing. ResultsMS lead to increased anxiety-like behaviour in Cdh13−/− mice compared to the other two MS groups. Cdh13−/− mice showed a context-dependent effect on stress- and anxiety-related behaviour, impaired extinction learning following contextual fear conditioning and decreased impulsivity, as well as a mild decrease in errors in the Barnes maze and reduced risk-taking in the light-dark transition test after MS. We also show sex differences, with increased locomotor activity in female Cdh13−/− mice, but unaltered impulsivity and activity in male Cdh13−/− mice. Transcriptome analysis revealed several pathways associated with cell surface/adhesion molecules to be altered following Cdh13 deficiency, together with an influence on endoplasmic reticulum function. ConclusionMS resulted in increased stress resilience, increased exploration and an overall anxiolytic behavioural phenotype in male Cdh13+/+ and Cdh13+/− mice. Cdh13 deficiency, however, obliterated most of the effects caused by early-life stress, with Cdh13−/− mice exhibiting delayed habituation, no reduction of anxiety-like behaviour and decreased fear extinction. Our behavioural findings indicate a role of CDH13 in the programming of and adaptation to early-life stress. Finally, our transcriptomic data support the view of CDH13 as a neuroprotective factor as well as a mediator in cell-cell interactions, with an impact on synaptic plasticity.