RESUMO
EDEM3 encodes a protein that converts Man8GlcNAc2 isomer B to Man7-5GlcNAc2. It is involved in the endoplasmic reticulum-associated degradation pathway, responsible for the recognition of misfolded proteins that will be targeted and translocated to the cytosol and degraded by the proteasome. In this study, through a combination of exome sequencing and gene matching, we have identified seven independent families with 11 individuals with bi-allelic protein-truncating variants and one individual with a compound heterozygous missense variant in EDEM3. The affected individuals present with an inherited congenital disorder of glycosylation (CDG) consisting of neurodevelopmental delay and variable facial dysmorphisms. Experiments in human fibroblast cell lines, human plasma, and mouse plasma and brain tissue demonstrated decreased trimming of Man8GlcNAc2 isomer B to Man7GlcNAc2, consistent with loss of EDEM3 enzymatic activity. In human cells, Man5GlcNAc2 to Man4GlcNAc2 conversion is also diminished with an increase of Glc1Man5GlcNAc2. Furthermore, analysis of the unfolded protein response showed a reduced increase in EIF2AK3 (PERK) expression upon stimulation with tunicamycin as compared to controls, suggesting an impaired unfolded protein response. The aberrant plasma N-glycan profile provides a quick, clinically available test for validating variants of uncertain significance that may be identified by molecular genetic testing. We propose to call this deficiency EDEM3-CDG.
Assuntos
Proteínas de Ligação ao Cálcio/genética , Defeitos Congênitos da Glicosilação/genética , Retículo Endoplasmático/genética , alfa-Manosidase/genética , Adolescente , Alelos , Proteínas de Ligação ao Cálcio/deficiência , Linhagem Celular , Criança , Pré-Escolar , Defeitos Congênitos da Glicosilação/sangue , Deficiências do Desenvolvimento/genética , Feminino , Glicoproteínas/sangue , Glicosilação , Humanos , Lactente , Deficiência Intelectual/genética , Masculino , Mutação , Linhagem , Polissacarídeos/sangue , Deficiências na Proteostase/genética , alfa-Manosidase/deficiênciaRESUMO
PURPOSE: Hardikar syndrome (HS, MIM #301068) is a female-specific multiple congenital anomaly syndrome characterized by retinopathy, orofacial clefting, aortic coarctation, biliary dysgenesis, genitourinary malformations, and intestinal malrotation. We previously showed that heterozygous nonsense and frameshift variants in MED12 cause HS. The phenotypic spectrum of disease and the mechanism by which MED12 variants cause disease is unknown. We aim to expand the phenotypic and molecular landscape of HS and elucidate the mechanism by which MED12 variants cause disease. METHODS: We assembled and clinically and molecularly characterized a cohort of 11 previously-unreported individuals with HS. We additionally studied the effect of MED12 deficiency on ciliary biology and hedgehog and YAP signaling, pathways implicated in diseases with phenotypic overlap with HS. RESULTS: We report novel phenotypes associated with HS, including cardiomyopathy, arrhythmia, and vascular anomalies and expand the molecular landscape of HS to include splice site variants. We additionally demonstrate that MED12 deficiency causes decreased cell ciliation and impairs hedgehog and YAP signaling. CONCLUSION: Our data support updating HS standard-of-care to include regular cardiac imaging, arrhythmia screening, and vascular imaging. We further propose that dysregulation of ciliogenesis and YAP and hedgehog signaling contributes to the pathogenesis of HS.
RESUMO
PURPOSE: Clinical next-generation sequencing is an effective approach for identifying pathogenic sequence variants that are medically actionable for participants and families but are not associated with the participant's primary diagnosis. These variants are called secondary findings (SFs). According to the literature, there is no report of the types and frequencies of SFs in a large pediatric cohort which includes substantial African-American participants. We sought to investigate the types (including American College of Medical Genetics and Genomics [ACMG] and non-ACMG recommended gene lists), frequencies, and rates of SFs, as well as the effects of SF disclosure on the participants and families of a large pediatric cohort at the Center for Applied Genomics at The Children's Hospital of Philadelphia (CHOP). METHODS: We systematically identified pathogenic (P) and likely pathogenic (LP) variants in established disease-causing genes, adhering to ACMG v3.2 secondary finding guidelines and beyond. For non-ACMG secondary findings, akin to incidental findings in clinical settings, we utilized a set of criteria focusing on pediatric onset, high penetrance, moderate to severe phenotypes, and the clinical actionability of the variants. This criteria-based approach was applied rather than using a fixed gene list to ensure that the variants identified are likely to impact participant health significantly. To identify and categorize these variants, we employed a clinical-grade variant classification standard per ACMG/AMP recommendations; additionally, we conducted a detailed literature search to ensure a comprehensive exploration of potential secondary findings relevant to pediatric participants. RESULTS: We report a distinctive distribution of 1,464 P/LP SF variants in 16,713 participants. There were 427 unique variants in ACMG genes and 265 in non-ACMG genes. The most frequently mutated genes among the ACMG and non-ACMG gene lists were TTR (41.6%) and CHEK2 (7.16%), respectively. Overall, variants of possible medical importance were found in 8.76% of participants in both ACMG (5.81%) and non-ACMG (2.95%) genes.
RESUMO
Pathogenic variants in USP9X, on X chromosome, have been implicated in syndromic intellectual disability (ID) in both males and females with distinct craniofacial features. We report a truncating variant, c.885_889delAAAAG, p.(Lys296Serfs*4), in the USP9X gene with incomplete penetrance in two nontwin female siblings with phenotypic resemblance to female-specific syndromic ID (MIM 300969, also known as MRX99F). To investigate the possible genetic etiology of the reduced penetrance, X-inactivation, RNA-Seq, and full quad exome analyses were attempted, but failed to identify a promising candidate modifier. While the penetrance of pathogenic variants in USP9X in female appears to be high (95%) and the variants frequently occur de novo, incomplete penetrance should be considered.
Assuntos
Deficiência Intelectual , Ubiquitina Tiolesterase , Exoma , Feminino , Humanos , Deficiência Intelectual/diagnóstico , Deficiência Intelectual/genética , Deficiência Intelectual/patologia , Masculino , Penetrância , RNA-Seq , Ubiquitina Tiolesterase/genética , Sequenciamento do ExomaRESUMO
Ichthyosis follicularis, atrichia, and photophobia syndrome (IFAP syndrome) is a rare, X-linked disorder caused by pathogenic variants in membrane-bound transcription factor protease, site 2 (MBTPS2). Pathogenic MBTPS2 variants also cause BRESHECK syndrome, characterized by the IFAP triad plus intellectual disability and multiple congenital anomalies. Here we present a patient with ichthyosis, sparse hair, pulmonic stenosis, kidney dysplasia, hypospadias, growth failure, thrombocytopenia, anemia, bone marrow fibrosis, and chronic diarrhea found by research-based exome sequencing to harbor a novel, maternally inherited MBTPS2 missense variant (c.766 G>A; (p.Val256Leu)). In vitro modeling supports variant pathogenicity, with impaired cell growth in cholesterol-depleted media, attenuated activation of the sterol regulatory element-binding protein pathway, and failure to activate the endoplasmic reticulum stress response pathway. Our case expands both the genetic and phenotypic spectrum of BRESHECK syndrome to include a novel MBTPS2 variant and cytopenias, bone marrow fibrosis, and chronic diarrhea.
Assuntos
Deficiência Intelectual , Alopecia/genética , Encéfalo/anormalidades , Anormalidades Congênitas , Orelha/anormalidades , Displasia Ectodérmica , Estresse do Retículo Endoplasmático/genética , Doenças Genéticas Ligadas ao Cromossomo X , Doença de Hirschsprung , Humanos , Deficiência Intelectual/genética , Rim/anormalidades , Masculino , Metaloendopeptidases/genética , Peptídeo Hidrolases , Esteróis , Fatores de TranscriçãoRESUMO
Teebi hypertelorism syndrome (THS; OMIM 145420) is a rare craniofacial disorder characterized by hypertelorism, prominent forehead, short nose with broad or depressed nasal root. Some cases of THS have been attributed to SPECC1L variants. Homozygous variants in CDH11 truncating the transmembrane and intracellular domains have been implicated in Elsahy-Waters syndrome (EWS; OMIM 211380) with hypertelorism. We report THS due to CDH11 heterozygous missense variants on 19 subjects from 9 families. All affected residues in the extracellular region of Cadherin-11 (CHD11) are highly conserved across vertebrate species and classical cadherins. Six of the variants that cluster around the EC2-EC3 and EC3-EC4 linker regions are predicted to affect Ca2+ binding that is required for cadherin stability. Two of the additional variants [c.164G > C, p.(Trp55Ser) and c.418G > A, p.(Glu140Lys)] are also notable as they are predicted to directly affect trans-homodimer formation. Immunohistochemical study demonstrates that CDH11 is strongly expressed in human facial mesenchyme. Using multiple functional assays, we show that five variants from the EC1, EC2-EC3 linker, and EC3 regions significantly reduced the cell-substrate trans adhesion activity and one variant from EC3-EC4 linker results in changes in cell morphology, focal adhesion, and migration, suggesting dominant negative effect. Characteristic features in this cohort included depressed nasal root, cardiac and umbilical defects. These features distinguished this phenotype from that seen in SPECC1L-related hypertelorism syndrome and CDH11-related EWS. Our results demonstrate heterozygous variants in CDH11, which decrease cell-cell adhesion and increase cell migratory behavior, cause a form of THS, as termed CDH11-related THS.
Assuntos
Anormalidades Múltiplas/genética , Caderinas/genética , Adesão Celular/genética , Anormalidades Craniofaciais/genética , Deformidades Congênitas do Pé/genética , Variação Genética/genética , Deformidades Congênitas da Mão/genética , Hipertelorismo/genética , Sequência de Aminoácidos , Movimento Celular/genética , Feminino , Heterozigoto , Homozigoto , Humanos , Masculino , Linhagem , FenótipoRESUMO
OBJECTIVE: Juvenile idiopathic arthritis (JIA) is the most common type of arthritis among children, but a few studies have investigated the contribution of rare variants to JIA. In this study, we aimed to identify rare coding variants associated with JIA for the genome-wide landscape. METHODS: We established a rare variant calling and filtering pipeline and performed rare coding variant and gene-based association analyses on three RNA-seq datasets composed of 228 JIA patients in the Gene Expression Omnibus against different sets of controls, and further conducted replication in our whole-exome sequencing (WES) data of 56 JIA patients. Then we conducted differential gene expression analysis and assessed the impact of recurrent functional coding variants on gene expression and signalling pathway. RESULTS: By the RNA-seq data, we identified variants in two genes reported in literature as JIA causal variants, as well as additional 63 recurrent rare coding variants seen only in JIA patients. Among the 44 recurrent rare variants found in polyarticular patients, 10 were replicated by our WES of patients with the same JIA subtype. Several genes with recurrent functional rare coding variants have also common variants associated with autoimmune diseases. We observed immune pathways enriched for the genes with rare coding variants and differentially expressed genes. CONCLUSION: This study elucidated a novel landscape of recurrent rare coding variants in JIA patients and uncovered significant associations with JIA at the gene pathway level. The convergence of common variants and rare variants for autoimmune diseases is also highlighted in this study.
Assuntos
Artrite Juvenil/genética , Variação Genética/genética , Fenômenos do Sistema Imunitário/genética , Criança , Bases de Dados Genéticas , Feminino , Expressão Gênica , Estudo de Associação Genômica Ampla , Humanos , Masculino , RNA-Seq , Transdução de Sinais/genética , Sequenciamento do ExomaRESUMO
Central conducting lymphatic anomaly (CCLA) is one of the complex lymphatic anomalies characterized by dilated lymphatic channels, lymphatic channel dysmotility and distal obstruction affecting lymphatic drainage. We performed whole exome sequencing (WES) of DNA from a four-generation pedigree and examined the consequences of the variant by transfection of mammalian cells and morpholino and rescue studies in zebrafish. WES revealed a heterozygous mutation in EPHB4 (RefSeq NM_004444.4; c.2334 + 1G>C) and RNA-Seq demonstrated that the EPHB4 mutation destroys the normal donor site, which leads to the use of a cryptic splice donor that results in retention of the intervening 12-bp intron sequence. Transient co-expression of the wild-type and mutant EPHB4 proteins showed reduced phosphorylation of tyrosine, consistent with a loss-of-function effect. Zebrafish ephb4a morpholino resulted in vessel misbranching and deformities in the lymphatic vessel development, indicative of possible differentiation defects in lymphatic vessels, mimicking the lymphatic presentations of the patients. Immunoblot analysis using zebrafish lysates demonstrated over-activation of mTORC1 as a consequence of reduced EPHB4 signaling. Strikingly, drugs that inhibit mTOR signaling or RAS-MAPK signaling effectively rescued the misbranching phenotype in a comparable manner. Moreover, knock-in of EPHB4 mutation in HEK293T cells also induced mTORC1 activity. Our data demonstrate the pathogenicity of the identified EPHB4 mutation as a novel cause of CCLA and suggesting that ERK inhibitors may have therapeutic benefits in such patients with complex lymphatic anomalies.
Assuntos
Sequenciamento do Exoma , Anormalidades Linfáticas/genética , Vasos Linfáticos/metabolismo , Receptor EphB4/genética , Animais , Modelos Animais de Doenças , Células HEK293 , Heterozigoto , Humanos , Anormalidades Linfáticas/metabolismo , Anormalidades Linfáticas/patologia , Vasos Linfáticos/patologia , Alvo Mecanístico do Complexo 1 de Rapamicina/genética , Linhagem , Fosforilação , Receptores Proteína Tirosina Quinases/genética , Transdução de Sinais , Peixe-Zebra/genéticaRESUMO
BACKGROUND: Absence of the anterior (ACL) or posterior cruciate ligament (PCL) are rare congenital malformations that result in knee joint instability, with a prevalence of 1.7 per 100,000 live births and can be associated with other lower-limb abnormalities such as ACL agnesia and absence of the menisci of the knee. While a few cases of absence of ACL/PCL are reported in the literature, a number of large familial case series of related conditions such as ACL agnesia suggest a potential underlying monogenic etiology. We performed whole exome sequencing of a family with two individuals affected by ACL/PCL. RESULTS: We identified copy number variation (CNV) deletion impacting the exon sequences of CEP57L1, present in the affected mother and her affected daughter based on the exome sequencing data. The deletion was validated using quantitative PCR (qPCR), and the gene was confirmed to be expressed in ACL ligament tissue. Interestingly, we detected reduced expression of CEP57L1 in Epstein-Barr virus (EBV) cells from the two patients in comparison with healthy controls. Evaluation of 3D protein structure showed that the helix-binding sites of the protein remain intact with the deletion, but other functional binding sites related to microtubule attachment are missing. The specificity of the CNV deletion was confirmed by showing that it was absent in ~700 exome sequencing samples as well as in the database of genomic variations (DGV), a database containing large numbers of annotated CNVs from previous scientific reports. CONCLUSIONS: We identified a novel CNV deletion that was inherited through an autosomal dominant transmission from an affected mother to her affected daughter, both of whom suffered from the absence of the anterior and posterior cruciate ligaments of the knees.
Assuntos
Anormalidades Congênitas/genética , Variações do Número de Cópias de DNA/genética , Articulação do Joelho/patologia , Proteínas Associadas aos Microtúbulos/genética , Proteínas Nucleares/genética , Adolescente , Adulto , Ligamento Cruzado Anterior/crescimento & desenvolvimento , Ligamento Cruzado Anterior/patologia , Anormalidades Congênitas/patologia , Exoma , Feminino , Humanos , Instabilidade Articular/genética , Instabilidade Articular/patologia , Articulação do Joelho/crescimento & desenvolvimento , Masculino , Ligamento Cruzado Posterior/crescimento & desenvolvimento , Ligamento Cruzado Posterior/patologia , Análise de Sequência de DNA , Deleção de SequênciaRESUMO
BACKGROUND: Systemic sclerosis (SSc) is a rheumatologic disease with a multifactorial etiology. Genome-wide association studies imply a polygenic, complex mode of inheritance with contributions from variation at the human leukocyte antigen locus and non-coding variation at a locus on chromosome 6p21, among other modestly impactful loci. Here we describe an 8-year-old female proband presenting with diffuse cutaneous SSc/scleroderma and a family history of SSc in a grandfather and maternal aunt. METHODS: We employed whole exome sequencing (WES) of three members of this family. We examined rare missense, nonsense, splice-altering, and coding indels matching an autosomal dominant inheritance model. We selected one missense variant for Sanger sequencing confirmation based on its predicted impact on gene function and location in a known SSc genetic locus. RESULTS: Bioinformatic analysis found eight candidate variants meeting our criteria. We identified a very rare missense variant in the regulatory NODP domain of NOTCH4 located at the 6p21 locus, c.4245G > A:p.Met1415Ile, segregating with the phenotype. This allele has a frequency of 1.83 × 10-5 by the data of the Exome Aggregation Consortium. CONCLUSION: This family suggests a novel mechanism of SSc pathogenesis in which a rare and penetrant coding variation can substantially elevate disease risk in contrast to the more modest non-coding variation typically found at this locus. These results suggest that modulation of the NOTCH4 gene might be responsible for the association signal at chromosome 6p21 in SSc.
Assuntos
Exoma/genética , Genes Dominantes/genética , Mutação de Sentido Incorreto , Receptor Notch4/genética , Escleroderma Sistêmico/genética , Alelos , Criança , Cromossomos Humanos Par 6/genética , Biologia Computacional , Feminino , Predisposição Genética para Doença , Avós , Heterozigoto , Humanos , Masculino , Linhagem , Penetrância , Domínios Proteicos/genética , Análise de Sequência de DNAAssuntos
Negro ou Afro-Americano/genética , Dermatite Atópica/genética , População Branca/genética , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas de Ligação a DNA/genética , Loci Gênicos , Humanos , Proteínas de Membrana/genética , Polimorfismo de Nucleotídeo Único , RNA Longo não Codificante/genética , Tioléster Hidrolases/genética , Fatores de Transcrição/genéticaRESUMO
Background: The relationship between ambient air pollution (AAP) exposure and asthma exacerbations is well-established. However, mitigation efforts have yielded mixed results, potentially due to genetic variability in the response to AAP. We hypothesize that common single nucleotide polymorphisms (SNPs) are linked to AAP sensitivity and test this through a Genome Wide Association Study (GWAS). Methods: We selected a cohort of pediatric asthma patients frequently exposed to AAP. Patients experiencing exacerbations immediately following AAP spikes were deemed sensitive. A GWAS compared sensitive versus non-sensitive patients. Findings were validated using data from the All of Us program. Results: Our study included 6,023 pediatric asthma patients. Due to the association between AAP exposure and race, GWAS analysis was feasible only in the African ancestry cohort. Seven risk loci reached genome-wide significance, including four non-intergenic variants. Two variants were validated: rs111970601 associated with sensitivity to CO (odds ratio [OR], 6.58; PL=L1.63L×L10-8; 95% CI, 3.42-12.66) and rs9836522 to PM2.5 sensitivity (OR 0.75; PL=L3,87 ×L10-9; 95% CI, 0.62-0.91). Interpretation: While genetic variants have been previously linked to asthma incidence and AAP exposure, this study is the first to link specific SNPs with AAP-related asthma exacerbations. The identified variants implicate genes with a known role in asthma and established links to AAP. Future research should explore how clinical interventions interact with genetic risk to mitigate the effects of AAP, particularly to enhance health equity for vulnerable populations. What is already known on this topic: The relationship between ambient air pollution (AAP) exposure and asthma exacerbations is well-established. However, efforts to mitigate the impact of AAP on children with asthma have yielded mixed results, potentially due to genetic variability in response to AAP. What this study adds: Using publicly available AAP data, we identify which children with asthma experience exacerbations immediately following spikes in AAP. We then conduct a Genome Wide Association Study (GWAS) comparing these patients with those who have no temporal association between AAP spikes and asthma exacerbations, identifying several Single Nucleotide Polymorphisms (SNPs) significantly associated with AAP sensitivity. How this study might affect research practice or policy: While genetic variants have previously been linked to asthma incidence and AAP exposure, this study is the first to link specific SNPs with AAP-related asthma exacerbations. This creates a framework for identifying children especially at risk when exposed to AAP. These children should be targeted with policy interventions to reduce exposure and may require specific treatments to mitigate the effects of ongoing AAP exposure in the interim.
RESUMO
Atopic dermatitis (AD) is a highly heritable and common inflammatory skin condition affecting children and adults worldwide. Multi-ancestry approaches to AD genetic association studies are poised to boost power to detect genetic signal and identify ancestry-specific loci contributing to AD risk. Here, we present a multi-ancestry GWAS meta-analysis of twelve AD cohorts from five ancestral populations totaling 56,146 cases and 602,280 controls. We report 101 genomic loci associated with AD, including 15 loci that have not been previously associated with AD or eczema. Fine-mapping, QTL colocalization, and cell-type enrichment analyses identified genes and cell types implicated in AD pathophysiology. Functional analyses in keratinocytes provide evidence for genes that could play a role in AD through epidermal barrier function. Our study provides new insights into the etiology of AD by harnessing multiple genetic and functional approaches to unveil the mechanisms by which AD-associated variants impact genes and cell types.
RESUMO
Pre-mRNA splicing is a highly coordinated process. While its dysregulation has been linked to neurological deficits, our understanding of the underlying molecular and cellular mechanisms remains limited. We implicated pathogenic variants in U2AF2 and PRPF19, encoding spliceosome subunits in neurodevelopmental disorders (NDDs), by identifying 46 unrelated individuals with 23 de novo U2AF2 missense variants (including 7 recurrent variants in 30 individuals) and 6 individuals with de novo PRPF19 variants. Eight U2AF2 variants dysregulated splicing of a model substrate. Neuritogenesis was reduced in human neurons differentiated from human pluripotent stem cells carrying two U2AF2 hyper-recurrent variants. Neural loss of function (LoF) of the Drosophila orthologs U2af50 and Prp19 led to lethality, abnormal mushroom body (MB) patterning, and social deficits, which were differentially rescued by wild-type and mutant U2AF2 or PRPF19. Transcriptome profiling revealed splicing substrates or effectors (including Rbfox1, a third splicing factor), which rescued MB defects in U2af50-deficient flies. Upon reanalysis of negative clinical exomes followed by data sharing, we further identified 6 patients with NDD who carried RBFOX1 missense variants which, by in vitro testing, showed LoF. Our study implicates 3 splicing factors as NDD-causative genes and establishes a genetic network with hierarchy underlying human brain development and function.
Assuntos
Transtornos do Neurodesenvolvimento , Spliceossomos , Humanos , Spliceossomos/genética , Redes Reguladoras de Genes , Transtornos do Neurodesenvolvimento/genética , Mutação de Sentido Incorreto , Splicing de RNA , Fatores de Processamento de RNA/genética , Proteínas Nucleares/genética , Enzimas Reparadoras do DNA/genéticaRESUMO
Cytotoxic lymphocytes kill target cells through polarized release of the content of lytic granules at the immunological synapse. In human NK cells, signals for granule polarization and for degranulation can be uncoupled: Binding of ß(2) integrin LFA-1 to ICAM is sufficient to induce polarization but not degranulation, whereas CD16 binding to IgG triggers unpolarized degranulation. In this study, we investigated the basis for this difference. IL-2-expanded human NK cells were stimulated by incubation with plate-bound ligands of LFA-1 (ICAM-1) and CD16 (human IgG). Surprisingly, LFA-1 elicited signals similar to those induced by CD16, including tyrosine phosphorylation of the TCR ζ-chain, tyrosine kinase Syk, and phospholipase C-γ. Whereas CD16 activated Ca(2+) mobilization and LAT phosphorylation, LFA-1 did not, but induced strong Pyk2 and paxillin phosphorylation. LFA-1-dependent granule polarization was blocked by inhibition of Syk, phospholipase C-γ, and protein kinase C, as well as by paxillin knockdown. Therefore, common signals triggered by CD16 and LFA-1 bifurcate to provide independent control of Ca(2+)-dependent degranulation and paxillin-dependent granule polarization.
Assuntos
Antígenos CD18/fisiologia , Polaridade Celular/imunologia , Grânulos Citoplasmáticos/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Células Matadoras Naturais/imunologia , Paxilina/fisiologia , Fosfolipase C gama/metabolismo , Proteínas Tirosina Quinases/metabolismo , Receptores de Antígenos de Linfócitos T/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/fisiologia , Animais , Sinalização do Cálcio/imunologia , Degranulação Celular/imunologia , Linhagem Celular , Células Cultivadas , Grânulos Citoplasmáticos/imunologia , Drosophila melanogaster , Proteínas Ligadas por GPI/fisiologia , Humanos , Molécula 1 de Adesão Intercelular/metabolismo , Molécula 1 de Adesão Intercelular/fisiologia , Células Matadoras Naturais/citologia , Células Matadoras Naturais/metabolismo , Ativação Linfocitária/imunologia , Proteínas de Membrana/fisiologia , Perforina/metabolismo , Fosforilação/imunologia , Ligação Proteica/imunologia , Receptores de IgG/fisiologia , Quinase Syk , Tirosina/metabolismoRESUMO
BACKGROUND: Neurodevelopmental disorders (NDDs), such as attention deficit hyperactivity disorder (ADHD) and autism spectrum disorder (ASD), are examples of complex and partially overlapping phenotypes that often lack definitive corroborating genetic information. ADHD and ASD have complex genetic associations implicated by rare recurrent copy number variations (CNVs). Both of these NDDs have been shown to share similar biological etiologies as well as genetic pleiotropy. METHODS: Platforms aimed at investigating genetic-based associations, such as high-density microarray technologies, have been groundbreaking techniques in the field of complex diseases, aimed at elucidating the underlying disease biology. Previous studies have uncovered CNVs associated with genes within shared candidate genomic networks, including glutamate receptor genes, across multiple different NDDs. To examine shared biological pathways across two of the most common NDDs, we investigated CNVs across 15,689 individuals with ADHD (n = 7920), ASD (n = 4318), or both (n = 3,416), as well as 19,993 controls. Cases and controls were matched by genotype array (i.e., Illumina array versions). Three case-control association studies each calculated and compared the observed vs. expected frequency of CNVs across individual genes, loci, pathways, and gene networks. Quality control measures of confidence in CNV-calling, prior to association analyses, included visual inspection of genotype and hybridization intensity. RESULTS: Here, we report results from CNV analysis in search for individual genes, loci, pathways, and gene networks. To extend our previous observations implicating a key role of the metabotropic glutamate receptor (mGluR) network in both ADHD and autism, we exhaustively queried patients with ASD and/or ADHD for CNVs associated with the 273 genomic regions of interest within the mGluR gene network (genes with one or two degrees protein-protein interaction with mGluR 1-8 genes). Among CNVs in mGluR network genes, we uncovered CNTN4 deletions enriched in NDD cases (P = 3.22E - 26, OR = 2.49). Additionally, we uncovered PRLHR deletions in 40 ADHD cases and 12 controls (P = 5.26E - 13, OR = 8.45) as well as clinically diagnostic relevant 22q11.2 duplications and 16p11.2 duplications in 23 ADHD + ASD cases and 9 controls (P = 4.08E - 13, OR = 15.05) and 22q11.2 duplications in 34 ADHD + ASD cases and 51 controls (P = 9.21E - 9, OR = 3.93); those control samples were not with previous 22qDS diagnosis in their EHR records. CONCLUSION: Together, these results suggest that disruption in neuronal cell-adhesion pathways confers significant risk to NDDs and showcase that rare recurrent CNVs in CNTN4, 22q11.2, and 16p11.2 are overrepresented in NDDs that constitute patients predominantly suffering from ADHD and ASD. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT02286817 First Posted: 10 November 14, ClinicalTrials.gov Identifier: NCT02777931 first posted: 19 May 2016, ClinicalTrials.gov Identifier: NCT03006367 first posted: 30 December 2016, ClinicalTrials.gov Identifier: NCT02895906 first posted: 12 September 2016.
Assuntos
Transtorno do Espectro Autista , Receptores de Glutamato Metabotrópico , Humanos , Transtorno do Espectro Autista/genética , Variações do Número de Cópias de DNA/genética , Predisposição Genética para Doença , Estudo de Associação Genômica Ampla , Receptores de Glutamato Metabotrópico/genéticaRESUMO
Central conducting lymphatic anomaly (CCLA) due to congenital maldevelopment of the lymphatics can result in debilitating and life-threatening disease with limited treatment options. We identified 4 individuals with CCLA, lymphedema, and microcystic lymphatic malformation due to pathogenic, mosaic variants in KRAS. To determine the functional impact of these variants and identify a targeted therapy for these individuals, we used primary human dermal lymphatic endothelial cells (HDLECs) and zebrafish larvae to model the lymphatic dysplasia. Expression of the p.Gly12Asp and p.Gly13Asp variants in HDLECs in a 2dimensional (2D) model and 3D organoid model led to increased ERK phosphorylation, demonstrating these variants activate the RAS/MAPK pathway. Expression of activating KRAS variants in the venous and lymphatic endothelium in zebrafish resulted in lymphatic dysplasia and edema similar to the individuals in the study. Treatment with MEK inhibition significantly reduced the phenotypes in both the organoid and the zebrafish model systems. In conclusion, we present the molecular characterization of the observed lymphatic anomalies due to pathogenic, somatic, activating KRAS variants in humans. Our preclinical studies suggest that MEK inhibition should be studied in future clinical trials for CCLA due to activating KRAS pathogenic variants.
Assuntos
Proteínas Proto-Oncogênicas p21(ras) , Peixe-Zebra , Animais , Humanos , Proteínas Proto-Oncogênicas p21(ras)/genética , Proteínas Proto-Oncogênicas p21(ras)/metabolismo , Células Endoteliais/metabolismo , Fosforilação , Quinases de Proteína Quinase Ativadas por Mitógeno/metabolismo , Proteínas de Peixe-Zebra/genética , Proteínas de Peixe-Zebra/metabolismoRESUMO
Vascular anomalies are malformations or tumors of the blood or lymphatic vasculature and can be life-threatening. Although molecularly targeted therapies can be life-saving, identification of the molecular etiology is often impeded by lack of accessibility to affected tissue samples, mosaicism or insufficient sequencing depth. In a cohort of 356 participants with vascular anomalies, including 104 with primary complex lymphatic anomalies (pCLAs), DNA from CD31+ cells isolated from lymphatic fluid or cell-free DNA from lymphatic fluid or plasma underwent ultra-deep sequencing thereby uncovering pathogenic somatic variants down to a variant allele fraction of 0.15%. A molecular diagnosis, including previously undescribed genetic causes, was obtained in 41% of participants with pCLAs and 72% of participants with other vascular malformations, leading to a new medical therapy for 63% (43/69) of participants and resulting in improvement in 63% (35/55) of participants on therapy. Taken together, these data support the development of liquid biopsy-based diagnostic techniques to identify previously undescribed genotype-phenotype associations and guide medical therapy in individuals with vascular anomalies.
Assuntos
Anormalidades Linfáticas , Malformações Vasculares , Humanos , Mutação , Testes Genéticos/métodos , Malformações Vasculares/diagnóstico , Malformações Vasculares/genética , Malformações Vasculares/terapia , Alelos , Anormalidades Linfáticas/genética , GenômicaRESUMO
OBJECTIVE: Juvenile idiopathic arthritis (JIA) is the most common chronic immune-mediated joint disease among children and encompasses a heterogeneous group of immune-mediated joint disorders classified into 7 subtypes according to clinical presentation. However, phenotype overlap and biologic evidence suggest a shared mechanistic basis between subtypes. This study was undertaken to systematically investigate shared genetic underpinnings of JIA subtypes. METHODS: We performed a heterogeneity-sensitive genome-wide association study encompassing a total of 1,245 JIA cases (classified into 7 subtypes) and 9,250 controls, followed by fine-mapping of candidate causal variants at each genome-wide significant locus, functional annotation, and pathway and network analysis. We further identified candidate drug targets and drug repurposing opportunities by in silico analyses. RESULTS: In addition to the major histocompatibility complex locus, we identified 15 genome-wide significant loci shared between at least 2 JIA subtypes, including 10 novel loci. Functional annotation indicated that candidate genes at these loci were expressed in diverse immune cell types. CONCLUSION: This study identified novel genetic loci shared by JIA subtypes. Our findings identified candidate mechanisms underlying JIA subtypes and candidate targets with drug repurposing opportunities for JIA treatment.