Requirement of Osr1 in second heart field (SHF) for proper outflow tract (OFT) alignment

Congenital heart disease (CHD) is current in 1/3 of all birth defects and has the prevalence of 1 in 1000 births.1 1/3 of all CHDs consist of cardiac outflow tract (OFT) defects.1 Despite its high prevalence, the genetic ontogeny of CHD has much to be studied. Knocking out Osr1, a gene encoding for a putative transcription factor containing four C2H2-type zinc finger motifs, is reported to cause heart defects including atrioventricular septal defects (AVSDs) in mice2-4 while its involvement in OFT development has yet to be studied. Osr1 is expressed in the second heart field (SHF), overlapping the expression of Gli1, an important modulator of sonic hedgehog (Shh) signaling pathway.4 Shh-signaling has been reported to contribute to the OFT development.5 Our preliminary study has shown that Osr1 deletion causes OFT defects including DORV and OA—suggesting its role in OFT alignment. We hypothesize that Osr1 is required in the SHF and Shh-signaling for proper OFT alignment. We will use a Cre-lox cell-specific KO technique to create embryos with Osr1 specific KO in SHF cells and Shh-receiving cells. We expect to observe high percentages of OFT misalignment including DORV and OA in embryos with Osr1 specific KO in SHF cells and Shh-receiving cells. This study will provide information which cell lineage is required for Osr1 in OFT development

Genetic risk and a primary role for cell-mediated immune mechanisms in multiple sclerosis.

Multiple sclerosis is a common disease of the central nervous system in which the interplay between inflammatory and neurodegenerative processes typically results in intermittent neurological disturbance followed by progressive accumulation of disability. Epidemiological studies have shown that genetic factors are primarily responsible for the substantially increased frequency of the disease seen in the relatives of affected individuals, and systematic attempts to identify linkage in multiplex families have confirmed that variation within the major histocompatibility complex (MHC) exerts the greatest individual effect on risk. Modestly powered genome-wide association studies (GWAS) have enabled more than 20 additional risk loci to be identified and have shown that multiple variants exerting modest individual effects have a key role in disease susceptibility. Most of the genetic architecture underlying susceptibility to the disease remains to be defined and is anticipated to require the analysis of sample sizes that are beyond the numbers currently available to individual research groups. In a collaborative GWAS involving 9,772 cases of European descent collected by 23 research groups working in 15 different countries, we have replicated almost all of the previously suggested associations and identified at least a further 29 novel susceptibility loci. Within the MHC we have refined the identity of the HLA-DRB1 risk alleles and confirmed that variation in the HLA-A gene underlies the independent protective effect attributable to the class I region. Immunologically relevant genes are significantly overrepresented among those mapping close to the identified loci and particularly implicate T-helper-cell differentiation in the pathogenesis of multiple sclerosis

Stroke genetics informs drug discovery and risk prediction across ancestries

Previous genome-wide association studies (GWASs) of stroke - the second leading cause of death worldwide - were conducted predominantly in populations of European ancestry(1,2). Here, in cross-ancestry GWAS meta-analyses of 110,182 patients who have had a stroke (five ancestries, 33% non-European) and 1,503,898 control individuals, we identify association signals for stroke and its subtypes at 89 (61 new) independent loci: 60 in primary inverse-variance-weighted analyses and 29 in secondary meta-regression and multitrait analyses. On the basis of internal cross-ancestry validation and an independent follow-up in 89,084 additional cases of stroke (30% non-European) and 1,013,843 control individuals, 87% of the primary stroke risk loci and 60% of the secondary stroke risk loci were replicated (P < 0.05). Effect sizes were highly correlated across ancestries. Cross-ancestry fine-mapping, in silico mutagenesis analysis(3), and transcriptome-wide and proteome-wide association analyses revealed putative causal genes (such as SH3PXD2A and FURIN) and variants (such as at GRK5 and NOS3). Using a three-pronged approach(4), we provide genetic evidence for putative drug effects, highlighting F11, KLKB1, PROC, GP1BA, LAMC2 and VCAM1 as possible targets, with drugs already under investigation for stroke for F11 and PROC. A polygenic score integrating cross-ancestry and ancestry-specific stroke GWASs with vascular-risk factor GWASs (integrative polygenic scores) strongly predicted ischaemic stroke in populations of European, East Asian and African ancestry(5). Stroke genetic risk scores were predictive of ischaemic stroke independent of clinical risk factors in 52,600 clinical-trial participants with cardiometabolic disease. Our results provide insights to inform biology, reveal potential drug targets and derive genetic risk prediction tools across ancestries.</p

Stroke genetics informs drug discovery and risk prediction across ancestries

Previous genome-wide association studies (GWASs) of stroke — the second leading cause of death worldwide — were conducted predominantly in populations of European ancestry1,2. Here, in cross-ancestry GWAS meta-analyses of 110,182 patients who have had a stroke (five ancestries, 33% non-European) and 1,503,898 control individuals, we identify association signals for stroke and its subtypes at 89 (61 new) independent loci: 60 in primary inverse-variance-weighted analyses and 29 in secondary meta-regression and multitrait analyses. On the basis of internal cross-ancestry validation and an independent follow-up in 89,084 additional cases of stroke (30% non-European) and 1,013,843 control individuals, 87% of the primary stroke risk loci and 60% of the secondary stroke risk loci were replicated (P < 0.05). Effect sizes were highly correlated across ancestries. Cross-ancestry fine-mapping, in silico mutagenesis analysis3, and transcriptome-wide and proteome-wide association analyses revealed putative causal genes (such as SH3PXD2A and FURIN) and variants (such as at GRK5 and NOS3). Using a three-pronged approach4, we provide genetic evidence for putative drug effects, highlighting F11, KLKB1, PROC, GP1BA, LAMC2 and VCAM1 as possible targets, with drugs already under investigation for stroke for F11 and PROC. A polygenic score integrating cross-ancestry and ancestry-specific stroke GWASs with vascular-risk factor GWASs (integrative polygenic scores) strongly predicted ischaemic stroke in populations of European, East Asian and African ancestry5. Stroke genetic risk scores were predictive of ischaemic stroke independent of clinical risk factors in 52,600 clinical-trial participants with cardiometabolic disease. Our results provide insights to inform biology, reveal potential drug targets and derive genetic risk prediction tools across ancestries

A gene pathway analysis highlights the role of cellular adhesion molecules in multiple sclerosis susceptibility

Genome-wide association studies (GWASs) perform per-SNP association tests to identify variants involved in disease or trait susceptibility. However, such an approach is not powerful enough to unravel genes that are not individually contributing to the disease/trait, but that may have a role in interaction with other genes as a group. Pathway analysis is an alternative way to highlight such group of genes. Using SNP association P-values from eight multiple sclerosis (MS) GWAS data sets, we performed a candidate pathway analysis for MS susceptibility by considering genes interacting in the cell adhesion molecule (CAMs) biological pathway using Cytoscape software. This network is a strong candidate, as it is involved in the crossing of the blood-brain barrier by the T cells, an early event in MS pathophysiology, and is used as an efficient therapeutic target. We drew up a list of 76 genes belonging to the CAM network. We highlighted 64 networks enriched with CAM genes with low P-values. Filtering by a percentage of CAM genes up to 50% and rejecting enriched signals mainly driven by transcription factors, we highlighted five networks associated with MS susceptibility. One of them, constituted of ITGAL, ICAM1 and ICAM3 genes, could be of interest to develop novel therapeutic targets

Multiple sclerosis genomic map implicates peripheral immune cells and microglia in susceptibility

INTRODUCTION: Multiple sclerosis (MS) is an inflammatory and degenerative disease of the central nervous system (CNS) that often presents in young adults. Over the past decade, certain elements of the genetic architecture of susceptibility have gradually emerged, but most of the genetic risk for MS remained unknown. RATIONALE: Earlier versions of the MS genetic map had highlighted the role of the adaptive arm of the immune system, implicating multiple different T cell subsets. We expanded our knowledge of MS susceptibility by performing a genetic association study in MS that leveraged genotype data from 47,429 MS cases and 68,374 control subjects. We enhanced this analysis with an in-depth and comprehensive evaluation of the functional impact of the susceptibility variants that we uncovered. RESULTS: We identified 233 statistically independent associations with MS susceptibility that are genome-wide significant. The major histocompatibility complex (MHC) contains 32 of these associations, and one, the first MS locus on a sex chromosome, is found in chromosome X. The remaining 200 associations are found in the autosomal non-MHC genome. Our genome-wide partitioning approach and large-scale replication effort allowed the evaluation of other variants that did not meet our strict threshold of significance, such as 416 variants that had evidence of statistical replication but did not reach the level of genome-wide statistical significance. Many of these loci are likely to be true susceptibility loci. The genome-wide and suggestive effects jointly explain ~48% of the estimated heritability for MS. Using atlases of gene expression patterns and epigenomic features, we documented that enrichment for MS susceptibility loci was apparent in many different immune cell types and tissues, whereas there was an absence of enrichment in tissue-level brain profiles. We extended the annotation analyses by analyzing new data generated from human induced pluripotent stem cell–derived neurons as well as from purified primary human astrocytes and microglia, observing that enrichment for MS genes is seen in human microglia, the resident immune cells of the brain, but not in astrocytes or neurons. Further, we have characterized the functional consequences of many MS susceptibility variants by identifying those that influence the expression of nearby genes in immune cells or brain. Last, we applied an ensemble of methods to prioritize 551 putative MS susceptibility genes that may be the target of the MS variants that meet a threshold of genome-wide significance. This extensive list of MS susceptibility genes expands our knowledge more than twofold and highlights processes relating to the development, maturation, and terminal differentiation of B, T, natural killer, and myeloid cells that may contribute to the onset of MS. These analyses focus our attention on a number of different cells in which the function of MS variants should be further investigated. Using reference protein-protein interaction maps, these MS genes can also be assembled into 13 communities of genes encoding proteins that interact with one another; this higher-order architecture begins to assemble groups of susceptibility variants whose functional consequences may converge on certain protein complexes that can be prioritized for further evaluation as targets for MS prevention strategies. CONCLUSION: We report a detailed genetic and genomic map of MS susceptibility, one that explains almost half of this disease’s heritability. We highlight the importance of several cells of the peripheral and brain resident immune systems—implicating both the adaptive and innate arms—in the translation of MS genetic risk into an auto-immune inflammatory process that targets the CNS and triggers a neurodegenerative cascade. In particular, the myeloid component highlights a possible role for microglia that requires further investigation, and the B cell component connects to the narrative of effective B cell–directed therapies in MS. These insights set the stage for a new generation of functional studies to uncover the sequence of molecular events that lead to disease onset. This perspective on the trajectory of disease onset will lay the foundation for developing primary prevention strategies that mitigate the risk of developing MS

Partial Loss of USP9X Function Leads to a Male Neurodevelopmental and Behavioral Disorder Converging on Transforming Growth Factor β Signaling

The X-chromosome gene USP9X encodes a deubiquitylating enzyme that has been associated with neurodevelopmental disorders primarily in female subjects. USP9X escapes X inactivation, and in female subjects de novo heterozygous copy number loss or truncating mutations cause haploinsufficiency culminating in a recognizable syndrome with intellectual disability and signature brain and congenital abnormalities. In contrast, the involvement of USP9X in male neurodevelopmental disorders remains tentative. We used clinically recommended guidelines to collect and interrogate the pathogenicity of 44 USP9X variants associated with neurodevelopmental disorders in males. Functional studies in patient-derived cell lines and mice were used to determine mechanisms of pathology. Twelve missense variants showed strong evidence of pathogenicity. We define a characteristic phenotype of the central nervous system (white matter disturbances, thin corpus callosum, and widened ventricles); global delay with significant alteration of speech, language, and behavior; hypotonia; joint hypermobility; visual system defects; and other common congenital and dysmorphic features. Comparison of in silico and phenotypical features align additional variants of unknown significance with likely pathogenicity. In support of partial loss-of-function mechanisms, using patient-derived cell lines, we show loss of only specific USP9X substrates that regulate neurodevelopmental signaling pathways and a united defect in transforming growth factor β signaling. In addition, we find correlates of the male phenotype in Usp9x brain-specific knockout mice, and further resolve loss of hippocampal-dependent learning and memory. Our data demonstrate the involvement of USP9X variants in a distinctive neurodevelopmental and behavioral syndrome in male subjects and identify plausible mechanisms of pathogenesis centered on disrupted transforming growth factor β signaling and hippocampal function