Circular RNA CircSLC22A23 Promotes Gastric Cancer Progression by Activating HNRNPU Expression.

BACKGROUND: Circular RNAs (CircRNAs) play essential roles in cancer occurrence as regulatory RNAs. However, circRNA-mediated regulation of gastric cancer (GC) remains poorly understood. AIM: The purpose of this study was to investigate the molecular mechanism of circSLC22A23 (hsa_circ_0075504) underlying GC occurrence. METHODS: CircSLC22A23 levels were first quantified by quantitative real-time reverse transcription-polymerase chain reaction in GC cell lines, 80 paired GC tissues and adjacent normal tissues, and 27 pairs of plasma samples from preoperative and postoperative patients with GC. Then circSLC22A23 was knocked-down with short hairpin RNA to analyze its oncogenic effects on the proliferation, migration, and invasion of GC cells. Finally, circRNA-binding proteins and their downstream target genes were identified by RNA pulldown, mass spectrometry, RNA immunoprecipitation, quantitative real-time reverse transcription-polymerase chain reaction, and Western blot assays. RESULTS: CircSLC22A23 was found to be highly expressed in GC cells, GC tissues, and plasma from GC patients. Knockdown of circSLC22A23 inhibited GC cell proliferation, migration and invasion. RNA pulldown and RNA immunoprecipitation assays verified the interaction between circSLC22A23 and heterogeneous nuclear ribonucleoprotein U (HNRNPU). Knockdown of circSLC22A23 decreased HNRNPU protein levels. Moreover, rescue assays showed that the tumor suppressive effect of circSLC22A23 knockdown was reversed by HNRNPU overexpression. Finally, epidermal growth factor receptor (EGFR) was found to be one of the downstream target genes of HNRNPU that was up regulated by circSLC22A23. CONCLUSION: CircSLC22A23 regulated the transcription of EGFR through activation of HNRNPU in GC cells, suggesting that circSLC22A23 may serve as a potential therapeutic target for the treatment of GC.

Alternative Splicing Factor Heterogeneous Nuclear Ribonucleoprotein U as a Promising Biomarker for Gastric Cancer Risk and Prognosis with Tumor-Promoting Properties.

Gastric cancer (GC) is a major global health concern with poor outcomes. Heterogeneous nuclear ribonucleoprotein U (HNRNPU) is a multifunctional protein that participates in pre-mRNA packaging, alternative splicing regulation, and chromatin remodeling. Its potential role in GC remains unclear. In this study, the expression characteristics of HNRNPU were analyzed by The Cancer Genome Atlas data, Gene Expression Omnibus data, and then further identified by real-time quantitative PCR and immunohistochemistry using tissue specimens. From superficial gastritis, atrophic gastritis, and hyperplasia to GC, the in situ expression of HNRNPU protein gradually increased, and the areas under the curve for diagnosis of GC and its precancerous lesions were 0.911 and 0.847, respectively. A nomogram integrating HNRNPU expression, lymph node metastasis, and other prognostic indicators exhibited an area under the curve of 0.785 for predicting survival risk. Knockdown of HNRNPU significantly inhibited GC cell proliferation, migration, and invasion and promoted apoptosis in vitro. In addition, RNA-sequencing analysis showed that HNRNPU could affect alternative splicing events in GC cells, with functional enrichment analysis revealing that HNRNPU may exert malignant biological function in GC progression through alternative splicing regulation. In summary, the increased expression of HNRNPU was significantly associated with the development of GC, with a good performance in diagnosing and predicting the prognostic risk of GC. Functionally, HNRNPU may play an oncogenic role in GC by regulating alternative splicing.

Deficiency of the Heterogeneous Nuclear Ribonucleoprotein U locus leads to delayed hindbrain neurogenesis.

Genetic variants affecting Heterogeneous Nuclear Ribonucleoprotein U (HNRNPU) have been identified in several neurodevelopmental disorders (NDDs). HNRNPU is widely expressed in the human brain and shows the highest postnatal expression in the cerebellum. Recent studies have investigated the role of HNRNPU in cerebral cortical development, but the effects of HNRNPU deficiency on cerebellar development remain unknown. Here, we describe the molecular and cellular outcomes of HNRNPU locus deficiency during in vitro neural differentiation of patient-derived and isogenic neuroepithelial stem cells with a hindbrain profile. We demonstrate that HNRNPU deficiency leads to chromatin remodeling of A/B compartments, and transcriptional rewiring, partly by impacting exon inclusion during mRNA processing. Genomic regions affected by the chromatin restructuring and host genes of exon usage differences show a strong enrichment for genes implicated in epilepsies, intellectual disability, and autism. Lastly, we show that at the cellular level HNRNPU downregulation leads to an increased fraction of neural progenitors in the maturing neuronal population. We conclude that the HNRNPU locus is involved in delayed commitment of neural progenitors to differentiate in cell types with hindbrain profile.

Selective DNA-binding of SP120 (rat ortholog of human hnRNP U) is mediated by arginine-glycine rich domain and modulated by RNA.

A human protein heterogeneous ribonucleoprotein U (hnRNP U) also known as Scaffold attachment factor A (SAF-A) and its orthologous rat protein SP120 are abundant and multifunctional nuclear protein that directly binds to both DNA and RNA. The C-terminal region of hnRNP U enriched with arginine and glycine is essential for the interaction with RNA and the N-terminal region of SAF-A termed SAP domain has been ascribed to the DNA binding. We have reported that rat hnRNP U specifically and cooperatively binds to AT-rich DNA called nuclear scaffold/matrix-associated region (S/MAR) although its detailed mechanism remained unclear. In the present study analysis of hnRNP U deletion mutants revealed for the first time that a C-terminal domain enriched with Arg-Gly (defined here as 'RG domain') is predominantly important for the S/MAR-selective DNA binding activities. RG domain alone directly bound to S/MAR and coexistence with the SAP domain exerted a synergistic effect. The binding was inhibited by netropsin, a minor groove binder with preference to AT pairs that are enriched in S/MAR, suggesting that RG domain interacts with minor groove of S/MAR DNA. Interestingly, excess amounts of RNA attenuated the RG domain-dependent S/MAR-binding of hnRNP U. Taken together, hnRNP U may be the key element for the RNA-regulated recognition of S/MAR DNA and thus contributing to the dynamic structural changes of chromatin compartments.

HNRNPU's multi-tasking is essential for proper cortical development.

Heterogeneous nuclear ribonucleoprotein U (HNRNPU) is a nuclear protein that plays a crucial role in various biological functions, such as RNA splicing and chromatin organization. HNRNPU/scaffold attachment factor A (SAF-A) activities are essential for regulating gene expression, DNA replication, genome integrity, and mitotic fidelity. These functions are critical to ensure the robustness of developmental processes, particularly those involved in shaping the human brain. As a result, HNRNPU is associated with various neurodevelopmental disorders (HNRNPU-related neurodevelopmental disorder, HNRNPU-NDD) characterized by developmental delay and intellectual disability. Our research demonstrates that the loss of HNRNPU function results in the death of both neural progenitor cells and post-mitotic neurons, with a higher sensitivity observed in the former. We reported that HNRNPU truncation leads to the dysregulation of gene expression and alternative splicing of genes that converge on several signaling pathways, some of which are likely to be involved in the pathology of HNRNPU-related NDD.

Malat1 deficiency prevents neonatal heart regeneration by inducing cardiomyocyte binucleation.

The adult mammalian heart has limited regenerative capacity, while the neonatal heart fully regenerates during the first week of life. Postnatal regeneration is mainly driven by proliferation of preexisting cardiomyocytes and supported by proregenerative macrophages and angiogenesis. Although the process of regeneration has been well studied in the neonatal mouse, the molecular mechanisms that define the switch between regenerative and nonregenerative cardiomyocytes are not well understood. Here, using in vivo and in vitro approaches, we identified the lncRNA Malat1 as a key player in postnatal cardiac regeneration. Malat1 deletion prevented heart regeneration in mice after myocardial infarction on postnatal day 3 associated with a decline in cardiomyocyte proliferation and reparative angiogenesis. Interestingly, Malat1 deficiency increased cardiomyocyte binucleation even in the absence of cardiac injury. Cardiomyocyte-specific deletion of Malat1 was sufficient to block regeneration, supporting a critical role of Malat1 in regulating cardiomyocyte proliferation and binucleation, a landmark of mature nonregenerative cardiomyocytes. In vitro, Malat1 deficiency induced binucleation and the expression of a maturation gene program. Finally, the loss of hnRNP U, an interaction partner of Malat1, induced similar features in vitro, suggesting that Malat1 regulates cardiomyocyte proliferation and binucleation by hnRNP U to control the regenerative window in the heart.

HNRNPU promotes the progression of triple-negative breast cancer via RNA transcription and alternative splicing mechanisms.

Triple-negative breast cancer (TNBC) is a great detriment to women's health due to the lack of effective therapeutic targets. In this study, we employed an integrated genetic screen to identify a pivotal oncogenic factor, heterogeneous nuclear ribonucleoprotein U (HNRNPU), which is required for the progression of TNBC. We elucidated the pro-oncogenic role of HNRNPU, which can induce the proliferation and migration of TNBC cells via its association with DEAD box helicase 5 (DDX5) protein. Elevated levels of the HNRNPU-DDX5 complex prohibited the intron retention of minichromosome maintenance protein 10 (MCM10) pre-mRNA, decreased nonsense-mediated mRNA decay, and activated Wnt/ß-catenin signalling; on the other hand, HNRNPU-DDX5 is located in the transcriptional start sites (TSS) of LIM domain only protein 4 (LMO4) and its upregulation promoted the transcription of LMO4, consequently activating PI3K-Akt-mTOR signalling. Our data highlight the synergetic effects of HNRNPU in RNA transcription and splicing in regulating cancer progression and suggest that HNRNPU may act as a potential molecular target in the treatment of TNBC.

Modulation of pre-mRNA structure by hnRNP proteins regulates alternative splicing of MALT1.

Alternative splicing plays key roles for cell type-specific regulation of protein function. It is controlled by cis-regulatory RNA elements that are recognized by RNA binding proteins (RBPs). The MALT1 paracaspase is a key factor of signaling pathways that mediate innate and adaptive immune responses. Alternative splicing of MALT1 is critical for controlling optimal T cell activation. We demonstrate that MALT1 splicing depends on RNA structural elements that sequester the splice sites of the alternatively spliced exon7. The RBPs hnRNP U and hnRNP L bind competitively to stem-loop RNA structures that involve the 5' and 3' splice sites flanking exon7. While hnRNP U stabilizes RNA stem-loop conformations that maintain exon7 skipping, hnRNP L disrupts these RNA elements to facilitate recruitment of the essential splicing factor U2AF2, thereby promoting exon7 inclusion. Our data represent a paradigm for the control of splice site selection by differential RBP binding and modulation of pre-mRNA structure.

Heterogeneous nuclear ribonucleoprotein U (HNRNPU) safeguards the developing mouse cortex.

HNRNPU encodes the heterogeneous nuclear ribonucleoprotein U, which participates in RNA splicing and chromatin organization. Microdeletions in the 1q44 locus encompassing HNRNPU and other genes and point mutations in HNRNPU cause brain disorders, including early-onset seizures and severe intellectual disability. We aimed to understand HNRNPU's roles in the developing brain. Our work revealed that HNRNPU loss of function leads to rapid cell death of both postmitotic neurons and neural progenitors, with an apparent higher sensitivity of the latter. Further, expression and alternative splicing of multiple genes involved in cell survival, cell motility, and synapse formation are affected following Hnrnpu's conditional truncation. Finally, we identified pharmaceutical and genetic agents that can partially reverse the loss of cortical structures in Hnrnpu mutated embryonic brains, ameliorate radial neuronal migration defects and rescue cultured neural progenitors' cell death.

Heterogeneous nuclear ribonucleoprotein U-actin complex derived from extracellular vesicles facilitates proliferation and migration of human coronary artery endothelial cells by promoting RNA polymerase II transcription.

Coronary artery disease (CAD) represents a fatal public threat. The involvement of extracellular vesicles (EVs) in CAD has been documented. This study explored the regulation of embryonic stem cells (ESCs)-derived EVs-hnRNPU-actin complex in human coronary artery endothelial cell (HCAEC) growth. Firstly, in vitro HCAEC hypoxia models were established. EVs were extracted from ESCs by ultracentrifugation. HCAECs were treated with EVs and si-VEGF for 24 h under hypoxia, followed by assessment of cell proliferation, apoptosis, migration, and tube formation. Uptake of EVs by HCAECs was testified. Additionally, hnRNPU, VEGF, and RNA Pol II levels were determined using Western blotting and CHIP assays. Interaction between hnRNPU and actin was evaluated by Co-immunoprecipitation assay. HCAEC viability and proliferation were lowered, apoptosis was enhanced, wound fusion was decreased, and the number of tubular capillary structures was reduced under hypoxia, whereas ESC-EVs treatment counteracted these effects. Moreover, EVs transferred hnRNPU into HCAECs. EVs-hnRNPU-actin complex increased RNA Pol II level on the VEGF gene promoter and promoted VEGF expression in HCAECs. Inhibition of hnRNPU or VEGF both annulled the promotion of EVs on HCAEC growth. Collectively, ESC-EVs-hnRNPU-actin increased RNA Pol II phosphorylation and VEGF expression, thus promoting HCAEC growth.

Expanding the phenotype of HNRNPU-related neurodevelopmental disorder with emphasis on seizure phenotype and review of literature.

Pathogenic variants in heterogeneous nuclear ribonucleoprotein U (HNRNPU) results in a novel neurodevelopmental disorder recently delineated. Here, we report on 17 previously unpublished patients carrying HNRNPU pathogenic variants. All patients were found to harbor de novo loss-of-function variants except for one individual where the inheritance could not be determined, as a parent was unavailable for testing. All patients had seizures which started in early childhood, global developmental delay, intellectual disability, and dysmorphic features. In addition, hypotonia, behavioral abnormalities (such as autistic features, aggression, anxiety, and obsessive-compulsive behaviors), and cardiac (septal defects) and/or brain abnormalities (ventriculomegaly and corpus callosum thinning/agenesis) were frequently observed. We have noted four recurrent variants in the literature (c.1089G>A p.(Trp363*), c.706_707del p.(Glu236Thrfs*6), c.847_857del p.(Phe283Serfs*5), and c.1681dels p.(Gln561Serfs*45)).

hnRNPub inhibits LPS-induced NF-κB pathway by targeting TRAF6 for K48-linked ubiquitination in miiuy croaker (Miichthys miiuy).

As an important adaptor protein in innate immunity, TRAF6 is not only responsible for the transduction of signal pathways, but its E3 ligase activity to transfer ubiquitination has also been widely studied. Under LPS stimulation, TRAF6 transfers the K63-linked ubiquitination chain to TAK1, which in turn activates the transcription factor NF-κB and cell signaling factors downstream of the signaling pathway. However, how TRAF6 expression is regulated remains largely unknown, especially in teleost. In this study, we identified hnRNPub as a suppressor of TRAF6 expression. The mRNA level of hnRNPub significantly increased under LPS stimulation, and hnRNPub inhibited NF-κB signaling pathway by targeting TRAF6. Knockdown of hnRNPub potentiated inflammatory cytokines, such as TNFα,IL-1ß,IL-8. Mechanistically, hnRNPub inhibited NF-κB signaling pathway through mediating K48-linked ubiquitination and proteasomal degradation of TRAF6. Thus, our findings reveal that hnRNPub limits LPS-induced innate activation by promoting K48-linked polyubiquitination and proteasomal degradation of TRAF6.

The role of SAF-A/hnRNP U in regulating chromatin structure.

Scaffold attachment factor A (SAF-A) or hnRNP U is a nuclear RNA-binding protein with a well-documented role in processing newly transcribed RNA. Recent studies also indicate that SAF-A can oligomerise in an ATP-dependent manner and interact with RNA to form a dynamic nuclear mesh. This mesh is thought to regulate nuclear and chromatin architecture, yet a mechanistic understanding is lacking. Here, we review developments in the field to understand how the SAF-A/RNA mesh affects chromatin organisation in interphase and mitosis. As SAF-A has an intrinsically disordered domain we discuss how the chromatin mesh is related to nuclear phase-separated condensates, which in other situations have been shown to regulate transcription and cell functions. Finally, we infer possible links between diseases emerging from SAF-A mutations and its role in chromatin organisation and regulation.

HNRNPU promotes the progression of hepatocellular carcinoma by enhancing CDK2 transcription.

The nuclear matrix-associated protein Heterogeneous Nuclear Ribonucleoprotein U (HNRNPU), also known as SAF-A, is known to maintain active chromatin structure in mouse hepatocytes. However, the functional roles and molecular mechanisms of HNRNPU in the development of hepatocellular carcinoma (HCC) remain largely unknown. Herein, we found that HNRNPU was upregulated in HCC, and the proliferation of HCC cells was inhibited in vitro and in vivo upon HNRNPU knockdown. Moreover, the upregulation of HNRNPU was correlated with poor prognosis in HCC. Mechanistically, HNRNPU bound to the CDK2 gene locus, a key factor in cell cycle regulation, where it was enriched with H3K27 acetylation (H3K27ac), H3K9 acetylation (H3K9ac), and H3K4 mono-methylation (H3K4me1). Furthermore, HNRNPU knockdown reduced the levels of H3K27ac and H3K9ac at the binding site, where the levels of H3K27 tri-methylation (H3K27me3) were increased, eventually leading to the downregulation of CDK2. Collectively, our results provide a new mechanism whereby HNRNPU promotes HCC development by enhancing the transcription of CDK2.

hnRNPU in Sertoli cells cooperates with WT1 and is essential for testicular development by modulating transcriptional factors Sox8/9.

Background: Sertoli cells are essential regulators of testicular fate in the differentiating gonad; however, its role and underlying molecular mechanism of regulating testicular development in prepubertal testes are poorly understood. Although several critical regulatory factors of Sertoli cell development and function have been identified, identifying extrinsic factors that regulate gonocyte proliferation and migration processes during neonatal testis development remains largely unknown. Methods: We used the Sertoli cell-specific conditional knockout strategy (Cre/Loxp) in mice and molecular biological analyses (Luciferase assay, ChIP-qPCR, RNA-Seq, etc.) in vitro and in vivo to study the physiological roles of hnRNPU in Sertoli cells on regulating testicular development in prepubertal testes. Results: We identified a co-transcription factor, hnRNPU, which is highly expressed in mouse and human Sertoli cells and required for neonatal Sertoli cell and pre-pubertal testicular development. Conditional knockout of hnRNPU in murine Sertoli cells leads to severe testicular atrophy and male sterility, characterized by rapid depletion of both Sertoli cells and germ cells and failure of spermatogonia proliferation and migration during pre-pubertal testicular development. At molecular levels, we found that hnRNPU interacts with two Sertoli cell markers WT1 and SOX9, and enhances the expression of two transcriptional factors, Sox8 and Sox9, in Sertoli cells by directly binding to their promoter regions. Further RNA-Seq and bioinformatics analyses revealed the transcriptome-wide of key genes essential for Sertoli cell and germ cell fate control, such as biological adhesion, proliferation and migration, were deregulated in Sertoli cell-specific hnRNPU mutant testes. Conclusion: Our findings demonstrate an essential role of hnRNPU in Sertoli cells for prepubertal testicular development and testis microenvironment maintenance and define a new insight for our understanding of male infertility therapy.

MicroRNA-132-3p alleviates neuron apoptosis and impairments of learning and memory abilities in Alzheimer's disease by downregulation of HNRNPU stabilized BACE1.

Alzheimer's disease (AD) is a progressive neuro-degenerative disease characterized by dementia. MicroRNAs (miRNAs) are involved in many diseases, including AD. MiR-132-3p has been identified to be downregulated in AD. In this study, we explored the effects of miR-132-3p on neuron apoptosis and impairments of learning and memory abilities. Aß1-42-stimulated SH-SY5Y cells were used as in vitro models of AD. An AD-like homocysteine (Hcy) rat model was established to evaluate the effects of miR-132-3p on AD pathogenesis in vivo. RIP, RNA pull down and luciferase reporter assays were conducted to investigate the relationship between miR-132-3p and its downstream target genes. The viability and apoptosis of SH-SY5Y cells were measured by CCK-8 and TUNEL assays. The rat spatial learning and memory abilities were accessed using Morris water maze test. Results indicated that miR-132-3p was downregulated in SH-SY5Y cells after Aß1-42 treatment and promoted cell apoptosis. Mechanistically, miR-132-3p targeted heterogeneous nuclear ribonucleoprotein U (HNRNPU). HNRNPU acted as an RNA binding protein (RBP) to regulate the mRNA stability of ß-site amyloid precursor protein cleaving enzyme 1 (BACE1). Overexpression of HNRNPU or BACE1 reversed the effects of miR-132-3p overexpression on the viability and apoptosis of Aß1-42-treated SH-SY5Y cells. In vivo experiments revealed the downregulation of miR-132-3p in the hippocampus of Hcy-treated rats. MiR-132-3p suppressed levels of apoptotic genes in hippocampus and reduced impairments of learning and memory abilities in Hcy-treated rats. In conclusion, miR-132-3p reduces apoptosis of SH-SY5Y cells and alleviates impairments of learning and memory abilities in AD rats by modulating the HNRNPU/BACE1 axis.

HNRNPU-AS1 Regulates Cell Proliferation and Apoptosis via the MicroRNA 205-5p/AXIN2 Axis and Wnt/ß-Catenin Signaling Pathway in Cervical Cancer.

Long noncoding RNAs (lncRNAs) have key functions in modulating cervical cancer (CC) genesis and progression. This work focused on exploring lncRNA HNRNPU-AS1's function in CC and the underlying mechanism. HNRNPU-AS1, AXIN2, and microRNA 205-5p (miR-205-5p) levels in CC cases were measured through reverse transcription-quantitative PCR. The relationship between miR-205-5p and AXIN2 or HNRNPU-AS1 was validated through a dual-luciferase assay. Cell proliferation was examined by CCK-8 and cell apoptosis by colony formation and flow cytometry analysis. HNRNPU-AS1 expression loss could be observed in CC patients and cell lines, which predicted the dismal prognosis of CC cases. Moreover, it was identified that the miR-205-5p level was upregulated, which acted as an inhibitory target of HNRNPU-AS1 and AXIN2. HNRNPU-AS1 inhibited cell proliferation and promoted apoptosis. As revealed by Kaplan-Meier curve, CC cases showing low HNRNPU-AS1, high miR-205-5p, and low AXIN2 levels had the poorest prognosis. AXIN2 reversed the CC cell proliferation-promoting, apoptosis-inhibiting, and Wnt/ß-catenin signaling-activating behavior mediated by miR-205-5p or HNRNPU-AS1 knockout. In conclusion, the overexpression of lncRNA HNRNPU-AS1 suppressed CC progression by inhibiting the Wnt/ß-catenin pathway through the miR-205-5p/AXIN2 axis.

De novo frameshift variants of HNRNPU in patients with early infantile epileptic encephalopathy: Two case reports and literature review.

Variants in HNRNPU have been reported in patients with epileptic encephalopathy, early infantile 54 (OMIM 602,869). We hereby describe two children from different families with autosomal dominance early-onset epileptic encephalopathy and summarize the genotype and phenotype of reported individuals. Whole-exome sequencing analysis was applied to the patients. De novo frameshift variants in the HNRNPU, c.143_149del7 (p.G48Afs*11) and c.1282delC(p.G429Afs*53) were identified. This is the first time to report Chinese patients with early infantile epileptic encephalopathy caused by HNRNPU variants, and so far, these variants have not been reported in population gene database. This study expands our knowledge of HNRNPU variants and emphasizes the importance of early gene diagnosis.

Identification of therapeutic targets and prognostic biomarkers from the hnRNP family in invasive breast carcinoma.

Heterogeneous nuclear ribonucleoproteins (hnRNPs) are RNA-binding proteins that are reported to play a crucial role in the pathogenic process of multiple malignancies. However, their expression patterns, clinical application significance and prognostic values in invasive breast carcinoma (BRCA) remain unknown. In this study, we investigated hnRNP family members in BRCA using accumulated data from Oncomine 4.5, UALCAN Web portal and other available databases. We explored the expression and prognostic value level of hnRNPs in BRCA. We further analyzed their association with the clinicopathological features of BRCA patients. Subsequently, we calculated the alteration frequency of hnRNPs, constructed the interaction network of hnRNPs, and examined the potential coexpression genes of hnRNPs, revealing that HNRNPU and SYNCRIP are the core molecular genes requiring further investigation for BRCA. We validated the immunohistochemistry (IHC) pattern to simulate clinical applications based on pathology. Cell function experiments conducted in vitro indicated that HNRNPU can promote epithelial-mesenchymal transition, functionally stimulating the invasion capacity and inhibiting the viability of invasive BRCA cells. In summary, our systematic analysis demonstrated that HNRNPU was the key molecule that played a fundamental role in BRCA metastasis, which may facilitate the development of new diagnostic and prognostic markers for the analysis of BRCA progression.

Large-scale targeted sequencing identifies risk genes for neurodevelopmental disorders.

Most genes associated with neurodevelopmental disorders (NDDs) were identified with an excess of de novo mutations (DNMs) but the significance in case-control mutation burden analysis is unestablished. Here, we sequence 63 genes in 16,294 NDD cases and an additional 62 genes in 6,211 NDD cases. By combining these with published data, we assess a total of 125 genes in over 16,000 NDD cases and compare the mutation burden to nonpsychiatric controls from ExAC. We identify 48 genes (25 newly reported) showing significant burden of ultra-rare (MAF < 0.01%) gene-disruptive mutations (FDR 5%), six of which reach family-wise error rate (FWER) significance (p < 1.25E-06). Among these 125 targeted genes, we also reevaluate DNM excess in 17,426 NDD trios with 6,499 new autism trios. We identify 90 genes enriched for DNMs (FDR 5%; e.g., GABRG2 and UIMC1); of which, 61 reach FWER significance (p < 3.64E-07; e.g., CASZ1). In addition to doubling the number of patients for many NDD risk genes, we present phenotype-genotype correlations for seven risk genes (CTCF, HNRNPU, KCNQ3, ZBTB18, TCF12, SPEN, and LEO1) based on this large-scale targeted sequencing effort.