Identifying canonical and replicable multi-scale intrinsic connectivity networks in 100k+ resting-state fMRI datasets.

Despite the known benefits of data-driven approaches, the lack of approaches for identifying functional neuroimaging patterns that capture both individual variations and inter-subject correspondence limits the clinical utility of rsfMRI and its application to single-subject analyses. Here, using rsfMRI data from over 100k individuals across private and public datasets, we identify replicable multi-spatial-scale canonical intrinsic connectivity network (ICN) templates via the use of multi-model-order independent component analysis (ICA). We also study the feasibility of estimating subject-specific ICNs via spatially constrained ICA. The results show that the subject-level ICN estimations vary as a function of the ICN itself, the data length, and the spatial resolution. In general, large-scale ICNs require less data to achieve specific levels of (within- and between-subject) spatial similarity with their templates. Importantly, increasing data length can reduce an ICN's subject-level specificity, suggesting longer scans may not always be desirable. We also find a positive linear relationship between data length and spatial smoothness (possibly due to averaging over intrinsic dynamics), suggesting studies examining optimized data length should consider spatial smoothness. Finally, consistency in spatial similarity between ICNs estimated using the full data and subsets across different data lengths suggests lower within-subject spatial similarity in shorter data is not wholly defined by lower reliability in ICN estimates, but may be an indication of meaningful brain dynamics which average out as data length increases.

High-resolution copy number variation analysis of schizophrenia in Japan.

Recent schizophrenia (SCZ) studies have reported an increased burden of de novo copy number variants (CNVs) and identified specific high-risk CNVs, although with variable phenotype expressivity. However, the pathogenesis of SCZ has not been fully elucidated. Using array comparative genomic hybridization, we performed a high-resolution genome-wide CNV analysis on a mainly (92%) Japanese population (1699 SCZ cases and 824 controls) and identified 7066 rare CNVs, 70.0% of which were small (<100 kb). Clinically significant CNVs were significantly more frequent in cases than in controls (odds ratio=3.04, P=9.3 × 10-9, 9.0% of cases). We confirmed a significant association of X-chromosome aneuploidies with SCZ and identified 11 de novo CNVs (e.g., MBD5 deletion) in cases. In patients with clinically significant CNVs, 41.7% had a history of congenital/developmental phenotypes, and the rate of treatment resistance was significantly higher (odds ratio=2.79, P=0.0036). We found more severe clinical manifestations in patients with two clinically significant CNVs. Gene set analysis replicated previous findings (e.g., synapse, calcium signaling) and identified novel biological pathways including oxidative stress response, genomic integrity, kinase and small GTPase signaling. Furthermore, involvement of multiple SCZ candidate genes and biological pathways in the pathogenesis of SCZ was suggested in established SCZ-associated CNV loci. Our study shows the high genetic heterogeneity of SCZ and its clinical features and raises the possibility that genomic instability is involved in its pathogenesis, which may be related to the increased burden of de novo CNVs and variable expressivity of CNVs.

DISC1 regulates trafficking and processing of APP and Aß generation.

We report the novel regulation of proteolytic processing of amyloid precursor protein (APP) by DISC1, a major risk factor for psychiatric illnesses, such as depression and schizophrenia. RNAi knockdown of DISC1 in mature primary cortical neurons led to a significant increase in the levels of intracellular α-C-terminal fragment of APP (APP-CTFα) and the corresponding N-terminal-secreted ectodomain product sAPPα. DISC1 knockdown also elicited a significant decrease in the levels of amyloid beta (Aß)42 and Aß40. These aberrant proteolytic events were successfully rescued by co-expression of wild-type DISC1, but not by mutant DISC1 lacking the amino acids required for the interaction with APP, suggesting that APP-DISC1 protein interactions are crucial for the regulation of the C-terminal proteolysis. In a genetically engineered model in which a major full-length DISC1 isoform is depleted, consistent changes in APP processing were seen: an increase in APP-CTFα and decrease in Aß42 and Aß40 levels. Finally, we found that knockdown of DISC1 increased the expression of APP at the cell surface and decreased its internalization. The presented DISC1 mechanism of APP proteolytic processing and Aß peptide generation, which is central to Alzheimer's disease pathology, suggests a novel interface between neurological and psychiatric conditions.

DISC1-ATF4 transcriptional repression complex: dual regulation of the cAMP-PDE4 cascade by DISC1.

Disrupted-In-Schizophrenia 1 (DISC1), a risk factor for major mental illnesses, has been studied extensively in the context of neurodevelopment. However, the role of DISC1 in neuronal signaling, particularly in conjunction with intracellular cascades that occur in response to dopamine, a neurotransmitter implicated in numerous psychiatric disorders, remains elusive. Previous data suggest that DISC1 interacts with numerous proteins that impact neuronal function, including activating transcription factor 4 (ATF4). In this study, we identify a novel DISC1 and ATF4 binding region in the genomic locus of phosphodiesterase 4D (PDE4D), a gene implicated in psychiatric disorders. We found that the loss of function of either DISC1 or ATF4 increases PDE4D9 transcription, and that the association of DISC1 with the PDE4D9 locus requires ATF4. We also show that PDE4D9 is increased by D1-type dopamine receptor dopaminergic stimulation. We demonstrate that the mechanism for this increase is due to DISC1 dissociation from the PDE4D locus in mouse brain. We further characterize the interaction of DISC1 with ATF4 to show that it is regulated via protein kinase A-mediated phosphorylation of DISC1 serine-58. Our results suggest that the release of DISC1-mediated transcriptional repression of PDE4D9 acts as feedback inhibition to regulate dopaminergic signaling. Furthermore, as DISC1 loss-of-function leads to a specific increase in PDE4D9, PDE4D9 itself may represent an attractive target for therapeutic approaches in psychiatric disorders.

The psychiatric disease risk factors DISC1 and TNIK interact to regulate synapse composition and function.

Disrupted in schizophrenia 1 (DISC1), a genetic risk factor for multiple serious psychiatric diseases including schizophrenia, bipolar disorder and autism, is a key regulator of multiple neuronal functions linked to both normal development and disease processes. As these diseases are thought to share a common deficit in synaptic function and architecture, we have analyzed the role of DISC1 using an approach that focuses on understanding the protein-protein interactions of DISC1 specifically at synapses. We identify the Traf2 and Nck-interacting kinase (TNIK), an emerging risk factor itself for disease, as a key synaptic partner for DISC1, and provide evidence that the DISC1-TNIK interaction regulates synaptic composition and activity by stabilizing the levels of key postsynaptic density proteins. Understanding the novel DISC1-TNIK interaction is likely to provide insights into the etiology and underlying synaptic deficits found in major psychiatric diseases.

Improving the depth resolution of STEM-ADF sectioning by 3D deconvolution.

Although the possibility of locating single atom in three dimensions using the scanning transmission electron microscope (STEM) has been discussed with the advent of aberration correction technology, it is still a big challenge. In this report we have developed deconvolution routines based on maximum entropy method (MEM) and Richardson-Lucy algorithm (RLA), which are applicable to the STEM-annular dark-field (ADF) though-focus images to improve the depth resolution. The new three-dimensional (3D) deconvolution routines require a limited defocus-range of STEM-ADF images that covers a whole sample and some vacuum regions. Since the STEM-ADF probe is infinitely elongated along the optical axis, a 3D convolution is performed with a two-dimensional (2D) convolution over xy-plane using the 2D fast Fourier transform in reciprocal space, and a one-dimensional convolution along the z-direction in real space. Using our new deconvolution routines, we have processed simulated focal series of STEM-ADF images for single Ce dopants embedded in wurtzite-type AlN. Applying the MEM, the Ce peaks are clearly localized along the depth, and the peak width is reduced down to almost one half. We also applied the new deconvolution routines to experimental focal series of STEM-ADF images of a monolayer graphene. The RLA gives smooth and high-P/B ratio scattering distribution, and the graphene layer can be easily detected. Using our deconvolution algorithms, we can determine the depth locations of the heavy dopants and the graphene layer within the precision of 0.1 and 0.2 nm, respectively. Thus, the deconvolution must be extremely useful for the optical sectioning with 3D STEM-ADF images.

Nuclear DISC1 regulates CRE-mediated gene transcription and sleep homeostasis in the fruit fly.

Disrupted-in-schizophrenia-1 (DISC1) is one of major susceptibility factors for a wide range of mental illnesses, including schizophrenia, bipolar disorder, major depression and autism spectrum conditions. DISC1 is located in several subcellular domains, such as the centrosome and the nucleus, and interacts with various proteins, including NudE-like (NUDEL/NDEL1) and activating transcription factor 4 (ATF4)/CREB2. Nevertheless, a role for DISC1 in vivo remains to be elucidated. Therefore, we have generated a Drosophila model for examining normal functions of DISC1 in living organisms. DISC1 transgenic flies with preferential accumulation of exogenous human DISC1 in the nucleus display disturbance in sleep homeostasis, which has been reportedly associated with CREB signaling/CRE-mediated gene transcription. Thus, in mammalian cells, we characterized nuclear DISC1, and identified a subset of nuclear DISC1 that colocalizes with the promyelocytic leukemia (PML) bodies, a nuclear compartment for gene transcription. Furthermore, we identified three functional cis-elements that regulate the nuclear localization of DISC1. We also report that DISC1 interacts with ATF4/CREB2 and a corepressor N-CoR, modulating CRE-mediated gene transcription.

Tobacco MAP kinase: a possible mediator in wound signal transduction pathways.

A complementary DNA encoding a mitogen-activated protein (MAP) kinase homolog has been isolated from tobacco plants. Transcripts of the corresponding gene were not observed in healthy tobacco leaves but began to accumulate 1 minute after mechanical wounding. In tobacco plants transformed with the cloned complementary DNA, trans inactivation of the endogenous homologous gene occurred, and both production of wound-induced jasmonic acid and accumulation of wound-inducible gene transcripts were inhibited. In contrast, the levels of salicylic acid and transcripts for pathogen-inducible, acidic pathogenesis-related proteins were increased upon wounding. These results indicate that this MAP kinase is part of the initial response of higher plants to mechanical wounding.

Rare genetic variants in CX3CR1 and their contribution to the increased risk of schizophrenia and autism spectrum disorders.

CX3CR1, a G protein-coupled receptor solely expressed by microglia in the brain, has been repeatedly reported to be associated with neurodevelopmental disorders including schizophrenia (SCZ) and autism spectrum disorders (ASD) in transcriptomic and animal studies but not in genetic studies. To address the impacts of variants in CX3CR1 on neurodevelopmental disorders, we conducted coding exon-targeted resequencing of CX3CR1 in 370 Japanese SCZ and 192 ASD patients using next-generation sequencing technology, followed by a genetic association study in a sample comprising 7054 unrelated individuals (2653 SCZ, 574 ASD and 3827 controls). We then performed in silico three-dimensional (3D) structural modeling and in vivo disruption of Akt phosphorylation to determine the impact of the detected variant on CX3CR1-dependent signal transduction. We detected a statistically significant association between the variant Ala55Thr in CX3CR1 with SCZ and ASD phenotypes (odds ratio=8.3, P=0.020). A 3D structural model indicated that Ala55Thr could destabilize the conformation of the CX3CR1 helix 8 and affect its interaction with a heterotrimeric G protein. In vitro functional analysis showed that the CX3CR1-Ala55Thr mutation inhibited cell signaling induced by fractalkine, the ligand for CX3CR1. The combined data suggested that the variant Ala55Thr in CX3CR1 might result in the disruption of CX3CR1 signaling. Our results strengthen the association between microglia-specific genes and neurodevelopmental disorders.

A novel rare variant R292H in RTN4R affects growth cone formation and possibly contributes to schizophrenia susceptibility.

Reticulon 4 receptor (RTN4R) plays an essential role in regulating axonal regeneration and plasticity in the central nervous system through the activation of rho kinase, and is located within chromosome 22q11.2, a region that is known to be a hotspot for schizophrenia (SCZ) and autism spectrum disorder (ASD). Recently, rare variants such as copy-number variants and single-nucleotide variants have been a focus of research because of their large effect size associated with increased susceptibility to SCZ and ASD and the possibility of elucidating the pathophysiology of mental disorder through functional analysis of the discovered rare variants. To discover rare variants with large effect size and to evaluate their role in the etiopathophysiology of SCZ and ASD, we sequenced the RTN4R coding exons with a sample comprising 370 SCZ and 192 ASD patients, and association analysis using a large number of unrelated individuals (1716 SCZ, 382 ASD and 4009 controls). Through this mutation screening, we discovered four rare (minor allele frequency <1%) missense mutations (R68H, D259N, R292H and V363M) of RTN4R. Among these discovered rare mutations, R292H was found to be significantly associated with SCZ (P=0.048). Furthermore, in vitro functional assays showed that the R292H mutation affected the formation of growth cones. This study strengthens the evidence for association between rare variants within RTN4R and SCZ, and may shed light on the molecular mechanisms underlying the neurodevelopmental disorder.

Abnormal neutrophil scattergram obtained using Pentra MS CRP in the patients with myelodysplastic syndrome showing dysgranulopoiesis.

INTRODUCTION: Pentra MS CRP is an automated hematology analyzer capable of cytochemistry using Chlorazol black E, a lipid-staining agent, for white blood cell (WBC) differentials. Pentra MS CRP displays a WBC scattergram according to the cell volume obtained using flow impedance and light absorbance reflecting the Chlorazol black E (CBE)-positive lipid content. METHOD: Neutrophil scattergrams obtained using Pentra MS CRP were compared between 5 patients with myelodysplastic syndrome (MDS) and normal controls. Sudan black B (SBB)-staining patterns of peripheral blood neutrophils were subdivided into four types (types I, II, III, and VI) based on their staining intensity and scored by counting 200 cells. Such SBB scores were also compared between the two groups. RESULTS: Neutrophil scattergrams deviated downward in the MDS group, suggesting the decreased CBE positivity that seemed reflect the reduction of the lipid content in dysplastic neutrophils. SBB scores determined in this study were also decreased in the MDS group when compared with those in normal controls. CONCLUSION: Pentra MS CRP might rapidly generate useful information on dysplastic neutrophils in patients with MDS based on its cytochemistry for WBC differentials during routine laboratory hematology.

Molecular signatures associated with cognitive deficits in schizophrenia: a study of biopsied olfactory neural epithelium.

Cognitive impairment is a key feature of schizophrenia (SZ) and determines functional outcome. Nonetheless, molecular signatures in neuronal tissues that associate with deficits are not well understood. We conducted nasal biopsy to obtain olfactory epithelium from patients with SZ and control subjects. The neural layers from the biopsied epithelium were enriched by laser-captured microdissection. We then performed an unbiased microarray expression study and implemented a systematic neuropsychological assessment on the same participants. The differentially regulated genes in SZ were further filtered based on correlation with neuropsychological traits. This strategy identified the SMAD 5 gene, and real-time quantitative PCR analysis also supports downregulation of the SMAD pathway in SZ. The SMAD pathway has been important in multiple tissues, including the role for neurodevelopment and bone formation. Here the involvement of the pathway in adult brain function is suggested. This exploratory study establishes a strategy to better identify neuronal molecular signatures that are potentially associated with mental illness and cognitive deficits. We propose that the SMAD pathway may be a novel target in addressing cognitive deficit of SZ in future studies.