Loss-of-Function of p21-Activated Kinase 2 Links BMP Signaling to Neural Tube Patterning Defects.

Closure of the neural tube represents a highly complex and coordinated process, the failure of which constitutes common birth defects. The serine/threonine kinase p21-activated kinase 2 (PAK2) is a critical regulator of cytoskeleton dynamics; however, its role in the neurulation and pathogenesis of neural tube defects (NTDs) remains unclear. Here, the results show that Pak2-/- mouse embryos fail to develop dorsolateral hinge points (DLHPs) and exhibit craniorachischisis, a severe phenotype of NTDs. Pak2 knockout activates BMP signaling that involves in vertebrate bone formation. Single-cell transcriptomes reveal abnormal differentiation trajectories and transcriptional events in Pak2-/- mouse embryos during neural tube development. Two nonsynonymous and one recurrent splice-site mutations in the PAK2 gene are identified in five human NTD fetuses, which exhibit attenuated PAK2 expression and upregulated BMP signaling in the brain. Mechanistically, PAK2 regulates Smad9 phosphorylation to inhibit BMP signaling and ultimately induce DLHP formation. Depletion of pak2a in zebrafish induces defects in the neural tube, which are partially rescued by the overexpression of wild-type, but not mutant PAK2. The findings demonstrate the conserved role of PAK2 in neurulation in multiple vertebrate species, highlighting the molecular pathogenesis of PAK2 mutations in NTDs.

Shared and divergent contribution of vitamin A and oxytocin to the aetiology of autism spectrum disorder.

Rare genetic variations contribute to the heterogeneity of autism spectrum disorder (ASD) and the responses to various interventions for ASD probands. However, the associated molecular underpinnings remain unclear. Herein, we estimated the association between rare genetic variations in 410 vitamin A (VA)-related genes (VARGs) and ASD aetiology using publicly available de novo mutations (DNMs), rare inherited variants, and copy number variations (CNVs) from about 50,000 ASD probands and 20,000 normal controls (discovery and validation cohorts). Additionally, given the functional relevance of VA and oxytocin, we systematically compared the similarities and differences between VA and oxytocin with respect to ASD aetiology and evaluated their potential for clinical applications. Functional DNMs and pathogenic CNVs in VARGs contributed to ASD pathogenesis in the discovery and validation cohorts. Additionally, 324 potential VA-related biomarkers were identified, 243 of which were shared with previously identified oxytocin-related biomarkers, while 81 were unique VA biomarkers. Moreover, multivariable logistic regression analysis revealed that both VA- and oxytocin-related biomarkers were able to predict ASD aetiology for individuals carrying functional DNM in corresponding biomarkers with an average precision of 0.94. As well as, convergent and divergent functions were also identified between VA- and oxytocin-related biomarkers. The findings of this study provide a basis for future studies aimed at understanding the pathophysiological mechanisms underlying ASD while also defining a set of potential molecular biomarkers for adjuvant diagnosis and intervention in ASD.

Npas3 deficiency impairs cortical astrogenesis and induces autistic-like behaviors.

Transcription factors with basic-helix-loop-helix (bHLH) motifs can control neural progenitor fate determination to neurons and oligodendrocytes. How bHLH transcription factors regulate astrogenesis remains largely unknown. Here, we report that NPAS3, a bHLH transcription factor, is a critical regulator of astrogenesis. Npas3 deficiency impairs cortical astrogenesis, correlating with abnormal brain development and autistic-like behaviors. Single-cell transcriptomes reveal that Npas3 knockout induces abnormal transition states in the differentiation trajectories from radial glia to astrocytes. Analysis of chromatin immunoprecipitation sequencing data in primary cortical astrocytes shows that NPAS3 binding targets are involved in functions of brain development and synapse organization. Co-culture assay further indicates that NPAS3-impaired astrogenesis induces synaptic deficits in wild-type neurons. Astrocyte-specific knockdown of NPAS3 in wild-type cortex causes synaptic and behavioral abnormalities associated with the core symptoms in autism. Together, our findings suggest that transcription factor NPAS3 regulates astrogenesis and its subsequent consequences for brain development and behavior.

Integrative analysis prioritised oxytocin-related biomarkers associated with the aetiology of autism spectrum disorder.

BACKGROUND: Autism spectrum disorder (ASD) is a neurodevelopmental disorder with high phenotypic and genetic heterogeneity. The common variants of specific oxytocin-related genes (OTRGs), particularly OXTR, are associated with the aetiology of ASD. The contribution of rare genetic variations in OTRGs to ASD aetiology remains unclear. METHODS: We catalogued publicly available de novo mutations (DNMs) [from 6,511 patients with ASD and 3,391 controls], rare inherited variants (RIVs) [from 1,786 patients with ASD and 1,786 controls], and both de novo copy number variations (dnCNVs) and inherited CNVs (ihCNVs) [from 15,581 patients with ASD and 6,017 controls] in 963 curated OTRGs to explore their contribution to ASD pathology, respectively. Finally, a combined model was designed to prioritise the contribution of each gene to ASD aetiology by integrating DNMs and CNVs. FINDINGS: The rare genetic variations of OTRGs were significantly associated with ASD aetiology, in the order of dnCNVs > ihCNVs > DNMs. Furthermore, 172 OTRGs and their connected 286 ASD core genes were prioritised to positively contribute to ASD aetiology, including top-ranked MAPK3. Probands carrying rare disruptive variations in these genes were estimated to account for 10∼11% of all ASD probands. INTERPRETATION: Our findings suggest that rare disruptive variations in 172 OTRGs and their connected 286 ASD core genes are associated with ASD aetiology and may be potential biomarkers predicting the effects of oxytocin treatment. FUNDING: Guangdong Key Project, National Natural Science Foundation of China, Key Laboratory of Clinical Laboratory Diagnosis and Translational Research of Zhejiang Province.

Transcriptomic signatures associated with autoimmune thyroiditis in papillary thyroid carcinoma and cancer immunotherapy-induced thyroid dysfunction.

Up to 20% of patients treated with anti-PD-1/PD-L1 inhibitors suffered from thyroid dysfunctions, yet the mediators associated with their occurrence remain unclear. The increasing coincidence of papillary thyroid carcinoma (PTC) with Hashimoto thyroiditis (HT) and the high vulnerability of thyroid to immunotherapy motivated us to discover the similarities and their underlying transcriptomic basis. Clinical characteristics analysis of 468 PTC patients from two independent cohorts and meta-analysis of 22,155 PTC patients unveiled a strong negative association between HT and recurrence in PTC patients. Transcriptome analysis of both cohorts showed PTC patients with HT were enriched in macrophages, CD8+ and CD4+ cytotoxic T cells, which was further validated by single-cell transcriptome analysis of 17,438 cells from PTC patients, and CD8+ T cells were correlated with disease-free survival of PTC patients. In both cohorts and single-cell dataset, elevated expression of PD-1-related genes was observed in the HT group, and CD3D appeared to be a target for enhancing the activation of CD8+ T cells. Correlation analysis of 3,318 thyroid adverse events from 39,123 patients across 24 tumor types and molecular signatures demonstrated similar signatures associated with autoimmune thyroiditis in PTC and thyroid immune-related adverse events (irAEs), and several multi-omics signatures, including signatures of CD8A and CD8+ T cells, showed positive associations with the odds ratio of thyroid irAEs. Our results unveil shared molecular signatures underlying thyroid dysfunction between patients receiving immunotherapies and PTC patients suffering from HT, which may shed light on managing the adverse events during cancer immunotherapy.

Putative complement control protein CSMD3 dysfunction impairs synaptogenesis and induces neurodevelopmental disorders.

Recent studies have reported that complement-related proteins modulate brain development through regulating synapse processes in the cortex. CSMD3 belongs to a group of putative complement control proteins. However, its role in the central nervous system and synaptogenesis remains largely unknown. Here we report that CSMD3 deleterious mutations occur frequently in patients with neurodevelopmental disorders (NDDs). Csmd3 is predominantly expressed in cortical neurons of the developing cortex. In mice, Csmd3 disruption induced retarded development and NDD-related behaviors. Csmd3 deficiency impaired synaptogenesis and neurogenesis, allowing fewer neurons reaching the cortical plate. Csmd3 deficiency also induced perturbed functional networks in the developing cortex, involving a number of downregulated synapse-associated genes that influence early synaptic organization and upregulated genes related to immune activity. Our study provides mechanistic insights into the endogenous regulation of complement-related proteins in synaptic development and supports the pathological role of CSMD3 in NDDs.

The p21-activated kinases in neural cytoskeletal remodeling and related neurological disorders.

The serine/threonine p21-activated kinases (PAKs), as main effectors of the Rho GTPases Cdc42 and Rac, represent a group of important molecular switches linking the complex cytoskeletal networks to broad neural activity. PAKs show wide expression in the brain, but they differ in specific cell types, brain regions, and developmental stages. PAKs play an essential and differential role in controlling neural cytoskeletal remodeling and are related to the development and fate of neurons as well as the structural and functional plasticity of dendritic spines. PAK-mediated actin signaling and interacting functional networks represent a common pathway frequently affected in multiple neurodevelopmental and neurodegenerative disorders. Considering specific small-molecule agonists and inhibitors for PAKs have been developed in cancer treatment, comprehensive knowledge about the role of PAKs in neural cytoskeletal remodeling will promote our understanding of the complex mechanisms underlying neurological diseases, which may also represent potential therapeutic targets of these diseases.

Targeted sequencing and integrative analysis of 3,195 Chinese patients with neurodevelopmental disorders prioritized 26 novel candidate genes.

Neurodevelopmental disorders (NDDs) are a set of complex disorders characterized by diverse and co-occurring clinical symptoms. The genetic contribution in patients with NDDs remains largely unknown. Here, we sequence 519 NDD-related genes in 3,195 Chinese probands with neurodevelopmental phenotypes and identify 2,522 putative functional mutations consisting of 137 de novo mutations (DNMs) in 86 genes and 2,385 rare inherited mutations (RIMs) with 22 X-linked hemizygotes in 13 genes, 2 homozygous mutations in 2 genes and 23 compound heterozygous mutations in 10 genes. Furthermore, the DNMs of 16,807 probands with NDDs are retrieved from public datasets and combine in an integrated analysis with the mutation data of our Chinese NDD probands by taking 3,582 in-house controls of Chinese origin as background. We prioritize 26 novel candidate genes. Notably, six of these genes - ITSN1, UBR3, CADM1, RYR3, FLNA, and PLXNA3 - preferably contribute to autism spectrum disorders (ASDs), as demonstrated by high co-expression and/or interaction with ASD genes confirmed via rescue experiments in a mouse model. Importantly, these genes are differentially expressed in the ASD cortex in a significant manner and involved in ASD-associated networks. Together, our study expands the genetic spectrum of Chinese NDDs, further facilitating both basic and translational research.