RESUMO
Dolichol is a lipid critical for N-glycosylation as a carrier for activated sugars and nascent oligosaccharides. It is commonly thought to be directly produced from polyprenol by the enzyme SRD5A3. Instead, we found that dolichol synthesis requires a three-step detour involving additional metabolites, where SRD5A3 catalyzes only the second reaction. The first and third steps are performed by DHRSX, whose gene resides on the pseudoautosomal regions of the X and Y chromosomes. Accordingly, we report a pseudoautosomal-recessive disease presenting as a congenital disorder of glycosylation in patients with missense variants in DHRSX (DHRSX-CDG). Of note, DHRSX has a unique dual substrate and cofactor specificity, allowing it to act as a NAD+-dependent dehydrogenase and as a NADPH-dependent reductase in two non-consecutive steps. Thus, our work reveals unexpected complexity in the terminal steps of dolichol biosynthesis. Furthermore, we provide insights into the mechanism by which dolichol metabolism defects contribute to disease.
Assuntos
Dolicóis , Dolicóis/metabolismo , Dolicóis/biossíntese , Humanos , Glicosilação , 3-Oxo-5-alfa-Esteroide 4-Desidrogenase/metabolismo , 3-Oxo-5-alfa-Esteroide 4-Desidrogenase/genética , Proteínas de Membrana/metabolismo , Proteínas de Membrana/genética , Defeitos Congênitos da Glicosilação/metabolismo , Defeitos Congênitos da Glicosilação/genética , Masculino , Mutação de Sentido Incorreto , FemininoRESUMO
The number one cause of human fetal death are defects in heart development. Because the human embryonic heart is inaccessible and the impacts of mutations, drugs, and environmental factors on the specialized functions of different heart compartments are not captured by in vitro models, determining the underlying causes is difficult. Here, we established a human cardioid platform that recapitulates the development of all major embryonic heart compartments, including right and left ventricles, atria, outflow tract, and atrioventricular canal. By leveraging 2D and 3D differentiation, we efficiently generated progenitor subsets with distinct first, anterior, and posterior second heart field identities. This advance enabled the reproducible generation of cardioids with compartment-specific in vivo-like gene expression profiles, morphologies, and functions. We used this platform to unravel the ontogeny of signal and contraction propagation between interacting heart chambers and dissect how mutations, teratogens, and drugs cause compartment-specific defects in the developing human heart.
Assuntos
Cardiopatias , Ventrículos do Coração , Coração , Humanos , Transcriptoma/genética , Linhagem Celular , Regulação da Expressão Gênica no Desenvolvimento , Cardiopatias/genética , Cardiopatias/metabolismoRESUMO
Giant congenital melanocytic nevi are NRAS-driven proliferations that may cover up to 80% of the body surface. Their most dangerous consequence is progression to melanoma. This risk often triggers preemptive extensive surgical excisions in childhood, producing severe lifelong challenges. We have presented preclinical models, including multiple genetically engineered mice and xenografted human lesions, which enabled testing locally applied pharmacologic agents to avoid surgery. The murine models permitted the identification of proliferative versus senescent nevus phases and treatments targeting both. These nevi recapitulated the histologic and molecular features of human giant congenital nevi, including the risk of melanoma transformation. Cutaneously delivered MEK, PI3K, and c-KIT inhibitors or proinflammatory squaric acid dibutylester (SADBE) achieved major regressions. SADBE triggered innate immunity that ablated detectable nevocytes, fully prevented melanoma, and regressed human giant nevus xenografts. These findings reveal nevus mechanistic vulnerabilities and suggest opportunities for topical interventions that may alter the therapeutic options for children with congenital giant nevi.
Assuntos
Melanoma , Nevo Pigmentado , Neoplasias Cutâneas , Animais , Xenoenxertos , Humanos , Melanoma/tratamento farmacológico , Melanoma/patologia , Camundongos , Transplante de Neoplasias , Nevo Pigmentado/congênito , Nevo Pigmentado/tratamento farmacológico , Nevo Pigmentado/patologia , Neoplasias Cutâneas/tratamento farmacológico , Neoplasias Cutâneas/patologia , Neoplasias Cutâneas/prevenção & controleRESUMO
Congenital heart disease (CHD) is present in 1% of live births, yet identification of causal mutations remains challenging. We hypothesized that genetic determinants for CHDs may lie in the protein interactomes of transcription factors whose mutations cause CHDs. Defining the interactomes of two transcription factors haplo-insufficient in CHD, GATA4 and TBX5, within human cardiac progenitors, and integrating the results with nearly 9,000 exomes from proband-parent trios revealed an enrichment of de novo missense variants associated with CHD within the interactomes. Scoring variants of interactome members based on residue, gene, and proband features identified likely CHD-causing genes, including the epigenetic reader GLYR1. GLYR1 and GATA4 widely co-occupied and co-activated cardiac developmental genes, and the identified GLYR1 missense variant disrupted interaction with GATA4, impairing in vitro and in vivo function in mice. This integrative proteomic and genetic approach provides a framework for prioritizing and interrogating genetic variants in heart disease.
Assuntos
Fator de Transcrição GATA4/metabolismo , Cardiopatias Congênitas , Proteínas Nucleares/metabolismo , Oxirredutases/metabolismo , Fatores de Transcrição , Animais , Cardiopatias Congênitas/genética , Camundongos , Mutação , Proteômica , Proteínas com Domínio T/genética , Fatores de Transcrição/genéticaRESUMO
Traumatic brain injury (TBI) is the largest non-genetic, non-aging related risk factor for Alzheimer's disease (AD). We report here that TBI induces tau acetylation (ac-tau) at sites acetylated also in human AD brain. This is mediated by S-nitrosylated-GAPDH, which simultaneously inactivates Sirtuin1 deacetylase and activates p300/CBP acetyltransferase, increasing neuronal ac-tau. Subsequent tau mislocalization causes neurodegeneration and neurobehavioral impairment, and ac-tau accumulates in the blood. Blocking GAPDH S-nitrosylation, inhibiting p300/CBP, or stimulating Sirtuin1 all protect mice from neurodegeneration, neurobehavioral impairment, and blood and brain accumulation of ac-tau after TBI. Ac-tau is thus a therapeutic target and potential blood biomarker of TBI that may represent pathologic convergence between TBI and AD. Increased ac-tau in human AD brain is further augmented in AD patients with history of TBI, and patients receiving the p300/CBP inhibitors salsalate or diflunisal exhibit decreased incidence of AD and clinically diagnosed TBI.
Assuntos
Doença de Alzheimer/etiologia , Doença de Alzheimer/prevenção & controle , Lesões Encefálicas Traumáticas/complicações , Neuroproteção , Proteínas tau/metabolismo , Acetilação , Doença de Alzheimer/metabolismo , Animais , Anti-Inflamatórios não Esteroides/uso terapêutico , Biomarcadores/sangue , Biomarcadores/metabolismo , Lesões Encefálicas Traumáticas/metabolismo , Linhagem Celular , Diflunisal/uso terapêutico , Feminino , Gliceraldeído-3-Fosfato Desidrogenase (Fosforiladora) , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Neurônios/metabolismo , Salicilatos/uso terapêutico , Sirtuína 1/metabolismo , Fatores de Transcrição de p300-CBP/antagonistas & inibidores , Fatores de Transcrição de p300-CBP/metabolismo , Proteínas tau/sangueRESUMO
Throughout development and aging, human cells accumulate mutations resulting in genomic mosaicism and genetic diversity at the cellular level. Mosaic mutations present in the gonads can affect both the individual and the offspring and subsequent generations. Here, we explore patterns and temporal stability of clonal mosaic mutations in male gonads by sequencing ejaculated sperm. Through 300× whole-genome sequencing of blood and sperm from healthy men, we find each ejaculate carries on average 33.3 ± 12.1 (mean ± SD) clonal mosaic variants, nearly all of which are detected in serial sampling, with the majority absent from sampled somal tissues. Their temporal stability and mutational signature suggest origins during embryonic development from a largely immutable stem cell niche. Clonal mosaicism likely contributes a transmissible, predicted pathogenic exonic variant for 1 in 15 men, representing a life-long threat of transmission for these individuals and a significant burden on human population health.
Assuntos
Crescimento e Desenvolvimento , Mosaicismo , Espermatozoides/metabolismo , Adolescente , Envelhecimento/sangue , Alelos , Células Clonais , Estudos de Coortes , Humanos , Masculino , Modelos Biológicos , Mutação/genética , Fatores de Risco , Fatores de Tempo , Adulto JovemRESUMO
Organoids capable of forming tissue-like structures have transformed our ability to model human development and disease. With the notable exception of the human heart, lineage-specific self-organizing organoids have been reported for all major organs. Here, we established self-organizing cardioids from human pluripotent stem cells that intrinsically specify, pattern, and morph into chamber-like structures containing a cavity. Cardioid complexity can be controlled by signaling that instructs the separation of cardiomyocyte and endothelial layers and by directing epicardial spreading, inward migration, and differentiation. We find that cavity morphogenesis is governed by a mesodermal WNT-BMP signaling axis and requires its target HAND1, a transcription factor linked to developmental heart chamber defects. Upon cryoinjury, cardioids initiated a cell-type-dependent accumulation of extracellular matrix, an early hallmark of both regeneration and heart disease. Thus, human cardioids represent a powerful platform to mechanistically dissect self-organization, congenital heart defects and serve as a foundation for future translational research.
Assuntos
Coração/embriologia , Organogênese , Organoides/embriologia , Ativinas/metabolismo , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Proteínas Morfogenéticas Ósseas/metabolismo , Cálcio/metabolismo , Linhagem Celular , Linhagem da Célula , Galinhas , Células Endoteliais/citologia , Proteínas da Matriz Extracelular/metabolismo , Feminino , Fibroblastos/citologia , Proteína Homeobox Nkx-2.5/metabolismo , Humanos , Masculino , Mesoderma/embriologia , Modelos Biológicos , Miocárdio/metabolismo , Células-Tronco Pluripotentes/citologia , Células-Tronco Pluripotentes/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo , Proteínas Wnt/metabolismoRESUMO
Development of the human intestine is not well understood. Here, we link single-cell RNA sequencing and spatial transcriptomics to characterize intestinal morphogenesis through time. We identify 101 cell states including epithelial and mesenchymal progenitor populations and programs linked to key morphogenetic milestones. We describe principles of crypt-villus axis formation; neural, vascular, mesenchymal morphogenesis, and immune population of the developing gut. We identify the differentiation hierarchies of developing fibroblast and myofibroblast subtypes and describe diverse functions for these including as vascular niche cells. We pinpoint the origins of Peyer's patches and gut-associated lymphoid tissue (GALT) and describe location-specific immune programs. We use our resource to present an unbiased analysis of morphogen gradients that direct sequential waves of cellular differentiation and define cells and locations linked to rare developmental intestinal disorders. We compile a publicly available online resource, spatio-temporal analysis resource of fetal intestinal development (STAR-FINDer), to facilitate further work.
Assuntos
Intestinos/citologia , Intestinos/crescimento & desenvolvimento , Análise de Célula Única , Células Endoteliais/citologia , Sistema Nervoso Entérico/citologia , Feto/embriologia , Fibroblastos/citologia , Humanos , Imunidade , Enteropatias/congênito , Enteropatias/patologia , Mucosa Intestinal/crescimento & desenvolvimento , Intestinos/irrigação sanguínea , Ligantes , Mesoderma/citologia , Neovascularização Fisiológica , Pericitos/citologia , Células-Tronco/citologia , Fatores de Tempo , Fatores de Transcrição/metabolismoRESUMO
Protein N-glycosylation is a widespread post-translational modification. The first committed step in this process is catalysed by dolichyl-phosphate N-acetylglucosamine-phosphotransferase DPAGT1 (GPT/E.C. 2.7.8.15). Missense DPAGT1 variants cause congenital myasthenic syndrome and disorders of glycosylation. In addition, naturally-occurring bactericidal nucleoside analogues such as tunicamycin are toxic to eukaryotes due to DPAGT1 inhibition, preventing their clinical use. Our structures of DPAGT1 with the substrate UDP-GlcNAc and tunicamycin reveal substrate binding modes, suggest a mechanism of catalysis, provide an understanding of how mutations modulate activity (thus causing disease) and allow design of non-toxic "lipid-altered" tunicamycins. The structure-tuned activity of these analogues against several bacterial targets allowed the design of potent antibiotics for Mycobacterium tuberculosis, enabling treatment in vitro, in cellulo and in vivo, providing a promising new class of antimicrobial drug.
Assuntos
Antibióticos Antituberculose/farmacologia , Defeitos Congênitos da Glicosilação/metabolismo , Inibidores Enzimáticos/farmacologia , N-Acetilglucosaminiltransferases/química , Animais , Antibióticos Antituberculose/química , Sítios de Ligação , Defeitos Congênitos da Glicosilação/genética , Inibidores Enzimáticos/química , Feminino , Células HEK293 , Células Hep G2 , Humanos , Metabolismo dos Lipídeos , Camundongos , Simulação de Acoplamento Molecular , Mutação , N-Acetilglucosaminiltransferases/antagonistas & inibidores , N-Acetilglucosaminiltransferases/genética , N-Acetilglucosaminiltransferases/metabolismo , Ligação Proteica , Células Sf9 , Spodoptera , Tunicamicina/química , Tunicamicina/farmacologia , Uridina Difosfato Ácido Glucurônico/química , Uridina Difosfato Ácido Glucurônico/metabolismoRESUMO
Ribonucleoprotein enzymes require dynamic conformations of their RNA constituents for regulated catalysis. Human telomerase employs a non-coding RNA (hTR) with a bipartite arrangement of domains-a template-containing core and a distal three-way junction (CR4/5) that stimulates catalysis through unknown means. Here, we show that telomerase activity unexpectedly depends upon the holoenzyme protein TCAB1, which in turn controls conformation of CR4/5. Cells lacking TCAB1 exhibit a marked reduction in telomerase catalysis without affecting enzyme assembly. Instead, TCAB1 inactivation causes unfolding of CR4/5 helices that are required for catalysis and for association with the telomerase reverse-transcriptase (TERT). CR4/5 mutations derived from patients with telomere biology disorders provoke defects in catalysis and TERT binding similar to TCAB1 inactivation. These findings reveal a conformational "activity switch" in human telomerase RNA controlling catalysis and TERT engagement. The identification of two discrete catalytic states for telomerase suggests an intramolecular means for controlling telomerase in cancers and progenitor cells.
Assuntos
RNA não Traduzido/química , Telomerase/metabolismo , Biocatálise , Linhagem Celular , Células HeLa , Humanos , Chaperonas Moleculares , Proteínas Nucleares/deficiência , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Conformação de Ácido Nucleico , Ligação Proteica , Interferência de RNA , RNA Interferente Pequeno/metabolismo , RNA não Traduzido/metabolismo , Telomerase/antagonistas & inibidores , Telomerase/química , Telomerase/genética , Telômero/metabolismoRESUMO
Mutation of highly conserved residues in transcription factors may affect protein-protein or protein-DNA interactions, leading to gene network dysregulation and human disease. Human mutations in GATA4, a cardiogenic transcription factor, cause cardiac septal defects and cardiomyopathy. Here, iPS-derived cardiomyocytes from subjects with a heterozygous GATA4-G296S missense mutation showed impaired contractility, calcium handling, and metabolic activity. In human cardiomyocytes, GATA4 broadly co-occupied cardiac enhancers with TBX5, another transcription factor that causes septal defects when mutated. The GATA4-G296S mutation disrupted TBX5 recruitment, particularly to cardiac super-enhancers, concomitant with dysregulation of genes related to the phenotypic abnormalities, including cardiac septation. Conversely, the GATA4-G296S mutation led to failure of GATA4 and TBX5-mediated repression at non-cardiac genes and enhanced open chromatin states at endothelial/endocardial promoters. These results reveal how disease-causing missense mutations can disrupt transcriptional cooperativity, leading to aberrant chromatin states and cellular dysfunction, including those related to morphogenetic defects.
Assuntos
Fator de Transcrição GATA4/genética , Cardiopatias Congênitas/genética , Cardiopatias Congênitas/patologia , Cromatina , Elementos Facilitadores Genéticos , Feminino , Coração/crescimento & desenvolvimento , Humanos , Células-Tronco Pluripotentes Induzidas , Masculino , Mutação de Sentido Incorreto , Miócitos Cardíacos/metabolismo , Miócitos Cardíacos/patologia , Fosfatidilinositol 3-Quinases/metabolismo , Transdução de Sinais , Proteínas com Domínio T/genéticaRESUMO
The RNA-binding protein TRIM71/LIN-41 is a phylogenetically conserved developmental regulator that functions in mammalian stem cell reprogramming, brain development, and cancer. TRIM71 recognizes target mRNAs through hairpin motifs and silences them through molecular mechanisms that await identification. Here, we uncover that TRIM71 represses its targets through RNA-supported interaction with TNRC6/GW182, a core component of the miRNA-induced silencing complex (miRISC). We demonstrate that AGO2, TRIM71, and UPF1 each recruit TNRC6 to specific sets of transcripts to silence them. As cellular TNRC6 levels are limiting, competition occurs among the silencing pathways, such that the loss of AGO proteins or of AGO binding to TNRC6 enhances the activities of the other pathways. We conclude that a miRNA-like silencing activity is shared among different mRNA silencing pathways and that the use of TNRC6 as a central hub provides a means to integrate their activities.
Assuntos
Proteínas Argonautas , MicroRNAs , Animais , Proteínas Argonautas/genética , Proteínas Argonautas/metabolismo , MicroRNAs/genética , MicroRNAs/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ligação Proteica , Células-Tronco/metabolismo , Mamíferos/metabolismoRESUMO
Congenital genetic disorders affecting limb morphology in humans and other mammals are particularly well described, due to both their rather high frequencies of occurrence and the ease of their detection when expressed as severe forms. In most cases, their molecular and cellular etiology remained unknown long after their initial description, often for several decades, and sometimes close to a century. Over the past 20 yr, however, experimental and conceptual advances in our understanding of gene regulation, in particular over large genomic distances, have allowed these cold cases to be reopened and, eventually, for some of them to be solved. These investigations led not only to the isolation of the culprit genes and mechanisms, but also to the understanding of the often complex regulatory processes that are disturbed in such mutant genetic configurations. Here, we present several cases in which dormant regulatory mutations have been retrieved from the archives, starting from a historical perspective up to their molecular explanations. While some cases remain open, waiting for new tools and/or concepts to bring their investigations to an end, the solutions to others have contributed to our understanding of particular features often found in the regulation of developmental genes and hence can be used as benchmarks to address the impact of noncoding variants in the future.
Assuntos
Genoma , Mamíferos , Animais , Humanos , MutaçãoRESUMO
Meiosis, a key process in the creation of haploid gametes, is a complex cellular division incorporating unique timing and intricate chromosome dynamics. Abnormalities in this elaborate dance can lead to the production of aneuploid gametes, i.e., eggs containing an incorrect number of chromosomes, many of which cannot generate a viable pregnancy. For many decades, research has been attempting to address why this process is notoriously error prone in humans compared to many other organisms. Rapidly developing technologies, access to new clinical material, and a mounting public infertility crisis have kept the field both active and quickly evolving. In this review, we discuss the history of aneuploidy in humans with a focus on its origins in maternal meiosis. We also gather current working mechanistic hypotheses, as well as up-and-coming areas of interest that point to future scientific avenues and their potential clinical applications.
Assuntos
Aneuploidia , Células Germinativas , Feminino , Gravidez , Humanos , Meiose/genética , HaploidiaRESUMO
Congenital heart defects (CHDs) are among the most common birth defects, but their etiology has long been mysterious. In recent decades, the development of a variety of experimental models has led to a greater understanding of the molecular basis of CHDs. In this review, we contrast mouse models of CHD, which maintain the anatomical arrangement of the heart, and human cellular models of CHD, which are more likely to capture human-specific biology but lack anatomical structure. We also discuss the recent development of cardiac organoids, which are a promising step toward more anatomically informative human models of CHD.
Assuntos
Cardiopatias Congênitas , Organoides , Animais , Modelos Animais de Doenças , Coração , Cardiopatias Congênitas/genética , Humanos , CamundongosRESUMO
The respiratory endoderm develops from a small cluster of cells located on the ventral anterior foregut. This population of progenitors generates the myriad epithelial lineages required for proper lung function in adults through a complex and delicately balanced series of developmental events controlled by many critical signaling and transcription factor pathways. In the past decade, understanding of this process has grown enormously, helped in part by cell lineage fate analysis and deep sequencing of the transcriptomes of various progenitors and differentiated cell types. This review explores how these new techniques, coupled with more traditional approaches, have provided a detailed picture of development of the epithelial lineages in the lung and insight into how aberrant development can lead to lung disease.
Assuntos
Endoderma/fisiologia , Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Pulmão/fisiologia , Morfogênese/fisiologia , Animais , Linhagem da Célula/fisiologia , Humanos , Organogênese/fisiologiaRESUMO
Expansion mutations in polyalanine stretches are associated with a growing number of diseases sharing a high degree of genotypic and phenotypic commonality. These similarities prompted us to query the normal function of physiological polyalanine stretches and to investigate whether a common molecular mechanism is involved in these diseases. Here, we show that UBA6, an E1 ubiquitin-activating enzyme, recognizes a polyalanine stretch within its cognate E2 ubiquitin-conjugating enzyme USE1. Aberrations in this polyalanine stretch reduce ubiquitin transfer to USE1 and, subsequently, polyubiquitination and degradation of its target, the ubiquitin ligase E6AP. Furthermore, we identify competition for the UBA6-USE1 interaction by various proteins with polyalanine expansion mutations in the disease state. The deleterious interactions of expanded polyalanine tract proteins with UBA6 in mouse primary neurons alter the levels and ubiquitination-dependent degradation of E6AP, which in turn affects the levels of the synaptic protein Arc. These effects are also observed in induced pluripotent stem cell-derived autonomic neurons from patients with polyalanine expansion mutations, where UBA6 overexpression increases neuronal resilience to cell death. Our results suggest a shared mechanism for such mutations that may contribute to the congenital malformations seen in polyalanine tract diseases.
Assuntos
Peptídeos , Enzimas Ativadoras de Ubiquitina , Ubiquitina , Humanos , Animais , Camundongos , Ubiquitinação , Ubiquitina/genética , Ubiquitina/metabolismo , Enzimas Ativadoras de Ubiquitina/genética , Enzimas Ativadoras de Ubiquitina/metabolismo , MutaçãoRESUMO
Infection directly influences adult hematopoietic stem cell (HSC) function and differentiation, but the fetal hematopoietic response to infection during pregnancy is not well-studied. Here, we investigated the fetal hematopoietic response to maternal infection with Toxoplasma gondii (T. gondii), an intracellular parasite that elicits Type II IFNγ-mediated maternal immunity. While it is known that maternal infection without direct pathogen transmission can affect fetal immune development, the effects of maternal IFNγ on developing HSCs and the signals that mediate these interactions have not been investigated. Our investigation reveals that the fetal HSCs respond to T. gondii infection with virulence-dependent changes in proliferation, self-renewal potential, and lineage output. Furthermore, maternal IFNγ crosses the fetal-maternal interface, where it is perceived by fetal HSCs. By comparing the effects of maternal IFNγ injection with maternal T. gondii infection, we reveal that the effects of IFNγ treatment mimic some aspects of the fetal HSC response to infection. Moreover, our findings illuminate that the fetal HSC response to prenatal infection is distinct from the adult HSC response to IFNγ-induced inflammation. Altogether, our data disentangle the role of infection-induced inflammatory cytokines in driving the expansion of downstream hematopoietic progenitors.
Assuntos
Toxoplasma , Toxoplasmose , Gravidez , Feminino , Humanos , Células-Tronco Hematopoéticas , Diferenciação Celular , Toxoplasmose/metabolismo , InflamaçãoRESUMO
Clinical exome and genome sequencing have revolutionized the understanding of human disease genetics. Yet many genes remain functionally uncharacterized, complicating the establishment of causal disease links for genetic variants. While several scoring methods have been devised to prioritize these candidate genes, these methods fall short of capturing the expression heterogeneity across cell subpopulations within tissues. Here, we introduce single-cell tissue-specific gene prioritization using machine learning (STIGMA), an approach that leverages single-cell RNA-seq (scRNA-seq) data to prioritize candidate genes associated with rare congenital diseases. STIGMA prioritizes genes by learning the temporal dynamics of gene expression across cell types during healthy organogenesis. To assess the efficacy of our framework, we applied STIGMA to mouse limb and human fetal heart scRNA-seq datasets. In a cohort of individuals with congenital limb malformation, STIGMA prioritized 469 variants in 345 genes, with UBA2 as a notable example. For congenital heart defects, we detected 34 genes harboring nonsynonymous de novo variants (nsDNVs) in two or more individuals from a set of 7,958 individuals, including the ortholog of Prdm1, which is associated with hypoplastic left ventricle and hypoplastic aortic arch. Overall, our findings demonstrate that STIGMA effectively prioritizes tissue-specific candidate genes by utilizing single-cell transcriptome data. The ability to capture the heterogeneity of gene expression across cell populations makes STIGMA a powerful tool for the discovery of disease-associated genes and facilitates the identification of causal variants underlying human genetic disorders.
Assuntos
Cardiopatias Congênitas , Transcriptoma , Humanos , Animais , Camundongos , Exoma/genética , Cardiopatias Congênitas/genética , Sequenciamento do Exoma , Aprendizado de Máquina , Análise de Célula Única/métodos , Enzimas Ativadoras de Ubiquitina/genéticaRESUMO
Pathogenic variants in multiple genes on the X chromosome have been implicated in syndromic and non-syndromic intellectual disability disorders. ZFX on Xp22.11 encodes a transcription factor that has been linked to diverse processes including oncogenesis and development, but germline variants have not been characterized in association with disease. Here, we present clinical and molecular characterization of 18 individuals with germline ZFX variants. Exome or genome sequencing revealed 11 variants in 18 subjects (14 males and 4 females) from 16 unrelated families. Four missense variants were identified in 11 subjects, with seven truncation variants in the remaining individuals. Clinical findings included developmental delay/intellectual disability, behavioral abnormalities, hypotonia, and congenital anomalies. Overlapping and recurrent facial features were identified in all subjects, including thickening and medial broadening of eyebrows, variations in the shape of the face, external eye abnormalities, smooth and/or long philtrum, and ear abnormalities. Hyperparathyroidism was found in four families with missense variants, and enrichment of different tumor types was observed. In molecular studies, DNA-binding domain variants elicited differential expression of a small set of target genes relative to wild-type ZFX in cultured cells, suggesting a gain or loss of transcriptional activity. Additionally, a zebrafish model of ZFX loss displayed an altered behavioral phenotype, providing additional evidence for the functional significance of ZFX. Our clinical and experimental data support that variants in ZFX are associated with an X-linked intellectual disability syndrome characterized by a recurrent facial gestalt, neurocognitive and behavioral abnormalities, and an increased risk for congenital anomalies and hyperparathyroidism.