Detailed phenotyping of Tbr1-2A-CreER knock-in mice demonstrates significant impacts on TBR1 protein levels and axon development.

Cre recombinase knock-in mouse lines have served as invaluable genetic tools for understanding key developmental processes altered in autism. However, insertion of exogenous DNA into the genome can have unintended effects on local gene regulation or protein function that must be carefully considered. Here, we analyze a recently generated Tbr1-2A-CreER knock-in mouse line, where a 2A-CreER cassette was inserted in-frame before the stop codon of the transcription factor gene Tbr1.

Detailed phenotyping of knock-in mice demonstrates significant impacts on TBR1 protein levels and axon development.

Spatiotemporal control of Cre-mediated recombination has been an invaluable tool for understanding key developmental processes. For example, knock-in of into cell type marker gene loci drives expression under endogenous promoter and enhancer sequences, greatly facilitating the study of diverse neuronal subtypes in the cerebral cortex. However, insertion of exogenous DNA into the genome can have unintended effects on local gene regulation or protein function that must be carefully considered.

Novel recruitment approaches and operational results for a statewide population Cohort for cancer research: The Healthy Oregon Project.

Background: Cancer health research relies on large-scale cohorts to derive generalizable results for different populations. While traditional epidemiological cohorts often use costly random sampling or self-motivated, preselected groups, a shift toward health system-based cohorts has emerged. However, such cohorts depend on participants remaining within a single system.

Aerosol delivery of SARS-CoV-2 human monoclonal antibodies in macaques limits viral replication and lung pathology.

Passively administered monoclonal antibodies (mAbs) given before or after viral infection can prevent or blunt disease. Here, we examine the efficacy of aerosol mAb delivery to prevent infection and disease in rhesus macaques inoculated with the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Delta variant via intranasal and intratracheal routes. SARS-CoV-2 human mAbs or a human mAb directed to respiratory syncytial virus (RSV) are nebulized and delivered using positive airflow via facemask to sedated macaques pre- and post-infection.

Population screening shows risk of inherited cancer and familial hypercholesterolemia in Oregon.

The Healthy Oregon Project (HOP) is a statewide effort that aims to build a large research repository and influence the health of Oregonians through providing no-cost genetic screening to participants for a next-generation sequencing 32-gene panel comprising genes related to inherited cancers and familial hypercholesterolemia. This type of unbiased population screening can detect at-risk individuals who may otherwise be missed by conventional medical approaches. However, challenges exist for this type of high-throughput testing in an academic setting, including developing a low-cost high-efficiency test and scaling up the clinical laboratory for processing large numbers of samples.

Integrating de novo and inherited variants in 42,607 autism cases identifies mutations in new moderate-risk genes.

To capture the full spectrum of genetic risk for autism, we performed a two-stage analysis of rare de novo and inherited coding variants in 42,607 autism cases, including 35,130 new cases recruited online by SPARK. We identified 60 genes with exome-wide significance (P < 2.5 × 10), including five new risk genes (NAV3, ITSN1, MARK2, SCAF1 and HNRNPUL2).

Shared and Distinct Functional Effects of Patient-Specific Mutations on Cortical Development.

T-Box Brain Transcription Factor 1 (TBR1) plays essential roles in brain development, mediating neuronal migration, fate specification, and axon tract formation. While heterozygous loss-of-function and missense mutations are associated with neurodevelopmental conditions, the effects of these heterogeneous mutations on brain development have yet to be fully explored. We characterized multiple mouse lines carrying mutations differing by type and exonic location, including the previously generated exon 2-3 knock-out (KO) line, and we analyzed male and female mice at neonatal and adult stages.

Cas12a-Capture: A Novel, Low-Cost, and Scalable Method for Targeted Sequencing.

Targeted sequencing remains a valuable technique for clinical and research applications. However, many existing technologies suffer from pervasive guanine-cytosine (GC) sequence content bias, high input DNA requirements, and high cost for custom panels. We have developed Cas12a-Capture, a low-cost and highly scalable method for targeted sequencing.

Validation of Autism Diagnosis and Clinical Data in the SPARK Cohort.

The SPARK cohort was established to facilitate recruitment in studies of large numbers of participants with autism spectrum disorder (ASD). Online registration requires participants to have received a lifetime professional diagnosis by health or school providers although diagnoses are not independently verified. This study was set to examine the validity of self- and caregiver-reported autism diagnoses.

High-content single-cell combinatorial indexing.

Single-cell combinatorial indexing (sci) with transposase-based library construction increases the throughput of single-cell genomics assays but produces sparse coverage in terms of usable reads per cell. We develop symmetrical strand sci ('s3'), a uracil-based adapter switching approach that improves the rate of conversion of source DNA into viable sequencing library fragments following tagmentation. We apply this chemistry to assay chromatin accessibility (s3-assay for transposase-accessible chromatin, s3-ATAC) in human cortical and mouse whole-brain tissues, with mouse datasets demonstrating a six- to 13-fold improvement in usable reads per cell compared with other available methods.

An Integrated Deep-Mutational-Scanning Approach Provides Clinical Insights on PTEN Genotype-Phenotype Relationships.

Germline variation in PTEN results in variable clinical presentations, including benign and malignant neoplasia and neurodevelopmental disorders. Despite decades of research, it remains unclear how the PTEN genotype is related to clinical outcomes. In this study, we combined two recent deep mutational scanning (DMS) datasets probing the effects of single amino acid variation on enzyme activity and steady-state cellular abundance with a large, well-curated clinical cohort of PTEN-variant carriers.

Autism and attention-deficit/hyperactivity disorders and symptoms in children with neurofibromatosis type 1.

Aim: To evaluate if autism symptoms and diagnoses are more common in children with neurofibromatosis type 1 (NF1) than in typically developing children, to which levels, and to determine if co-occurring attention-deficit/hyperactivity disorder (ADHD) symptomatology accounts for this increase.

Method: We searched hospital electronic medical records (EMR) for International Classification of Diseases, 10th Revision NF1 and co-occurring diagnoses codes. We recruited a subsample of 45 children (mean age 9y 2mo; SD 2y 7mo; range 5-12y; 22 males, 23 females) and collected parental reports of autism symptomatology, adaptive behavior, and behavioral problems that were compared to those of 360 age- and sex-matched controls from the Simons Simplex Collection (SSC) with autism spectrum disorder (ASD; SSC-ASD) or typically developing (SSC-TD).

Beliefs in vaccine as causes of autism among SPARK cohort caregivers.

Background: Fear of autism has led to a decline in childhood-immunization uptake and to a resurgence of preventable infectious diseases. Identifying characteristics of parents who believe in a causal role of vaccines for autism spectrum disorder (ASD) in their child may help targeting educational activities and improve adherence to the immunization schedule.

Objectives: To compare caregivers of children with ASD who agree or disagree that vaccines play an etiological role in autism for 1) socio-demographics characteristics and 2) developmental and clinical profiles of their children.

Clinical Phenotypes of Carriers of Mutations in CHD8 or Its Conserved Target Genes.

Background: Variants disruptive to CHD8 (which codes for the protein CHD8 [chromodomain-helicase-DNA-binding protein 8]) are among the most common mutations revealed by exome sequencing in autism spectrum disorder (ASD). Recent work has indicated that CHD8 plays a role in the regulation of other ASD-risk genes. However, it is unclear whether a possible shared genetic ontology extends to the phenotype.

Exome sequencing of 457 autism families recruited online provides evidence for autism risk genes.

Autism spectrum disorder (ASD) is a genetically heterogeneous condition, caused by a combination of rare de novo and inherited variants as well as common variants in at least several hundred genes. However, significantly larger sample sizes are needed to identify the complete set of genetic risk factors. We conducted a pilot study for SPARK (SPARKForAutism.

A Saturation Mutagenesis Approach to Understanding PTEN Lipid Phosphatase Activity and Genotype-Phenotype Relationships.

Phosphatase and tensin homolog (PTEN) is a tumor suppressor frequently mutated in diverse cancers. Germline PTEN mutations are also associated with a range of clinical outcomes, including PTEN hamartoma tumor syndrome (PHTS) and autism spectrum disorder (ASD). To empower new insights into PTEN function and clinically relevant genotype-phenotype relationships, we systematically evaluated the effect of PTEN mutations on lipid phosphatase activity in vivo.

Highly scalable generation of DNA methylation profiles in single cells.

We present a highly scalable assay for whole-genome methylation profiling of single cells. We use our approach, single-cell combinatorial indexing for methylation analysis (sci-MET), to produce 3,282 single-cell bisulfite sequencing libraries and achieve read alignment rates of 68 ± 8%. We apply sci-MET to discriminate the cellular identity of a mixture of three human cell lines and to identify excitatory and inhibitory neuronal populations from mouse cortical tissue.

Exonic Mosaic Mutations Contribute Risk for Autism Spectrum Disorder.

Genetic risk factors for autism spectrum disorder (ASD) have yet to be fully elucidated. Postzygotic mosaic mutations (PMMs) have been implicated in several neurodevelopmental disorders and overgrowth syndromes. By leveraging whole-exome sequencing data on a large family-based ASD cohort, the Simons Simplex Collection, we systematically evaluated the potential role of PMMs in autism risk.

Current Perspectives in Autism Spectrum Disorder: From Genes to Therapy.

Autism spectrum disorder (ASD) is a constellation of neurodevelopmental presentations with high heritability and both phenotypic and genetic heterogeneity. To date, mutations in hundreds of genes have been associated to varying degrees with increased ASD risk. A better understanding of the functions of these genes and whether they fit together in functional groups or impact similar neuronal circuits is needed to develop rational treatment strategies.

Mutations and Modeling of the Chromatin Remodeler CHD8 Define an Emerging Autism Etiology.

Autism Spectrum Disorder (ASD) is a common neurodevelopmental disorder with a strong but complex genetic component. Recent family based exome-sequencing strategies have identified recurrent de novo mutations at specific genes, providing strong evidence for ASD risk, but also highlighting the extreme genetic heterogeneity of the disorder. However, disruptions in these genes converge on key molecular pathways early in development.

Large-scale targeted sequencing comparison highlights extreme genetic heterogeneity in nephronophthisis-related ciliopathies.

Background: The term nephronophthisis-related ciliopathies (NPHP-RC) describes a group of rare autosomal-recessive cystic kidney diseases, characterised by broad genetic and clinical heterogeneity. NPHP-RC is frequently associated with extrarenal manifestations and accounts for the majority of genetically caused chronic kidney disease (CKD) during childhood and adolescence. Generation of a molecular diagnosis has been impaired by this broad genetic heterogeneity.

Joubert syndrome: a model for untangling recessive disorders with extreme genetic heterogeneity.

Background: Joubert syndrome (JS) is a recessive neurodevelopmental disorder characterised by hypotonia, ataxia, cognitive impairment, abnormal eye movements, respiratory control disturbances and a distinctive mid-hindbrain malformation. JS demonstrates substantial phenotypic variability and genetic heterogeneity. This study provides a comprehensive view of the current genetic basis, phenotypic range and gene-phenotype associations in JS.