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1.
Am J Hum Genet ; 110(11): 1919-1937, 2023 11 02.
Artigo em Inglês | MEDLINE | ID: mdl-37827158

RESUMO

Misregulation of histone lysine methylation is associated with several human cancers and with human developmental disorders. DOT1L is an evolutionarily conserved gene encoding a lysine methyltransferase (KMT) that methylates histone 3 lysine-79 (H3K79) and was not previously associated with a Mendelian disease in OMIM. We have identified nine unrelated individuals with seven different de novo heterozygous missense variants in DOT1L through the Undiagnosed Disease Network (UDN), the SickKids Complex Care genomics project, and GeneMatcher. All probands had some degree of global developmental delay/intellectual disability, and most had one or more major congenital anomalies. To assess the pathogenicity of the DOT1L variants, functional studies were performed in Drosophila and human cells. The fruit fly DOT1L ortholog, grappa, is expressed in most cells including neurons in the central nervous system. The identified DOT1L variants behave as gain-of-function alleles in flies and lead to increased H3K79 methylation levels in flies and human cells. Our results show that human DOT1L and fly grappa are required for proper development and that de novo heterozygous variants in DOT1L are associated with a Mendelian disease.


Assuntos
Anormalidades Congênitas , Deficiências do Desenvolvimento , Histona-Lisina N-Metiltransferase , Humanos , Mutação com Ganho de Função , Histona-Lisina N-Metiltransferase/genética , Histonas/genética , Histonas/metabolismo , Lisina , Metilação , Metiltransferases/genética , Neoplasias/genética , Drosophila/genética , Proteínas de Drosophila/genética , Deficiências do Desenvolvimento/genética , Anormalidades Congênitas/genética
2.
Am J Hum Genet ; 110(5): 774-789, 2023 05 04.
Artigo em Inglês | MEDLINE | ID: mdl-37054711

RESUMO

The Integrator complex is a multi-subunit protein complex that regulates the processing of nascent RNAs transcribed by RNA polymerase II (RNAPII), including small nuclear RNAs, enhancer RNAs, telomeric RNAs, viral RNAs, and protein-coding mRNAs. Integrator subunit 11 (INTS11) is the catalytic subunit that cleaves nascent RNAs, but, to date, mutations in this subunit have not been linked to human disease. Here, we describe 15 individuals from 10 unrelated families with bi-allelic variants in INTS11 who present with global developmental and language delay, intellectual disability, impaired motor development, and brain atrophy. Consistent with human observations, we find that the fly ortholog of INTS11, dIntS11, is essential and expressed in the central nervous systems in a subset of neurons and most glia in larval and adult stages. Using Drosophila as a model, we investigated the effect of seven variants. We found that two (p.Arg17Leu and p.His414Tyr) fail to rescue the lethality of null mutants, indicating that they are strong loss-of-function variants. Furthermore, we found that five variants (p.Gly55Ser, p.Leu138Phe, p.Lys396Glu, p.Val517Met, and p.Ile553Glu) rescue lethality but cause a shortened lifespan and bang sensitivity and affect locomotor activity, indicating that they are partial loss-of-function variants. Altogether, our results provide compelling evidence that integrity of the Integrator RNA endonuclease is critical for brain development.


Assuntos
Proteínas de Drosophila , Doenças do Sistema Nervoso , Adulto , Animais , Humanos , Drosophila/genética , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Mutação/genética , RNA Mensageiro
3.
Genet Med ; 26(9): 101174, 2024 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-38847193

RESUMO

PURPOSE: We identified 2 individuals with de novo variants in SREBF2 that disrupt a conserved site 1 protease (S1P) cleavage motif required for processing SREBP2 into its mature transcription factor. These individuals exhibit complex phenotypic manifestations that partially overlap with sterol regulatory element binding proteins (SREBP) pathway-related disease phenotypes, but SREBF2-related disease has not been previously reported. Thus, we set out to assess the effects of SREBF2 variants on SREBP pathway activation. METHODS: We undertook ultrastructure and gene expression analyses using fibroblasts from an affected individual and utilized a fly model of lipid droplet (LD) formation to investigate the consequences of SREBF2 variants on SREBP pathway function. RESULTS: We observed reduced LD formation, endoplasmic reticulum expansion, accumulation of aberrant lysosomes, and deficits in SREBP2 target gene expression in fibroblasts from an affected individual, indicating that the SREBF2 variant inhibits SREBP pathway activation. Using our fly model, we discovered that SREBF2 variants fail to induce LD production and act in a dominant-negative manner, which can be rescued by overexpression of S1P. CONCLUSION: Taken together, these data reveal a mechanism by which SREBF2 pathogenic variants that disrupt the S1P cleavage motif cause disease via dominant-negative antagonism of S1P, limiting the cleavage of S1P targets, including SREBP1 and SREBP2.


Assuntos
Fibroblastos , Mutação de Sentido Incorreto , Proteína de Ligação a Elemento Regulador de Esterol 2 , Humanos , Proteína de Ligação a Elemento Regulador de Esterol 2/genética , Proteína de Ligação a Elemento Regulador de Esterol 2/metabolismo , Animais , Fibroblastos/metabolismo , Mutação de Sentido Incorreto/genética , Masculino , Feminino , Gotículas Lipídicas/metabolismo , Fenótipo , Retículo Endoplasmático/metabolismo , Retículo Endoplasmático/genética , Serina Endopeptidases , Pró-Proteína Convertases
4.
Am J Med Genet A ; : e63854, 2024 Aug 21.
Artigo em Inglês | MEDLINE | ID: mdl-39166407

RESUMO

We report three unrelated individuals with atypical clinical findings for cardio-facio-cutaneous (CFC) syndrome, all of whom have the same novel, heterozygous de novo p.H119Y (c.355 C>T) transition variant in MAP2K1, identified by exome sequencing. MAP2K1 encodes MEK1, dual specificity mitogen-activated protein kinase kinase 1, and is one of four genes in the canonical RAS/MAPK signal transduction pathway associated with CFC syndrome. The p.H119Y variant is a non-conservative amino acid substitution that is predicted to impact the tertiary protein structure, and it occurs at a position in the protein kinase domain of MAP2K1 that is highly conserved across species. The clinical findings in these three individuals include facial features that are nonclassical for CFC syndrome, extremely poor weight gain, absence of congenital cardiac defects or cardiomyopathy, normal cognition or only mild intellectual disabilities, normal hair, mild skin abnormalities, and consistent behavioral features of anxiety, photophobia, and sensory hypersensitivities. These individuals expand the phenotypic spectrum of MAP2K1-related RASopathy.

5.
Am J Hum Genet ; 107(6): 1062-1077, 2020 12 03.
Artigo em Inglês | MEDLINE | ID: mdl-33217309

RESUMO

Dysfunction of the endolysosomal system is often associated with neurodegenerative disease because postmitotic neurons are particularly reliant on the elimination of intracellular aggregates. Adequate function of endosomes and lysosomes requires finely tuned luminal ion homeostasis and transmembrane ion fluxes. Endolysosomal CLC Cl-/H+ exchangers function as electric shunts for proton pumping and in luminal Cl- accumulation. We now report three unrelated children with severe neurodegenerative disease, who carry the same de novo c.1658A>G (p.Tyr553Cys) mutation in CLCN6, encoding the late endosomal Cl-/H+-exchanger ClC-6. Whereas Clcn6-/- mice have only mild neuronal lysosomal storage abnormalities, the affected individuals displayed severe developmental delay with pronounced generalized hypotonia, respiratory insufficiency, and variable neurodegeneration and diffusion restriction in cerebral peduncles, midbrain, and/or brainstem in MRI scans. The p.Tyr553Cys amino acid substitution strongly slowed ClC-6 gating and increased current amplitudes, particularly at the acidic pH of late endosomes. Transfection of ClC-6Tyr553Cys, but not ClC-6WT, generated giant LAMP1-positive vacuoles that were poorly acidified. Their generation strictly required ClC-6 ion transport, as shown by transport-deficient double mutants, and depended on Cl-/H+ exchange, as revealed by combination with the uncoupling p.Glu200Ala substitution. Transfection of either ClC-6Tyr553Cys/Glu200Ala or ClC-6Glu200Ala generated slightly enlarged vesicles, suggesting that p.Glu200Ala, previously associated with infantile spasms and microcephaly, is also pathogenic. Bafilomycin treatment abrogated vacuole generation, indicating that H+-driven Cl- accumulation osmotically drives vesicle enlargement. Our work establishes mutations in CLCN6 associated with neurological diseases, whose spectrum of clinical features depends on the differential impact of the allele on ClC-6 function.


Assuntos
Canais de Cloreto/genética , Mutação com Ganho de Função , Doenças Neurodegenerativas/genética , Alelos , Animais , Células CHO , Criança , Cricetulus , Eletrofisiologia , Endossomos/metabolismo , Feminino , Células HeLa , Heterozigoto , Homeostase , Humanos , Concentração de Íons de Hidrogênio , Lactente , Transporte de Íons , Íons , Proteínas de Membrana Lisossomal/metabolismo , Lisossomos/metabolismo , Macrolídeos/farmacologia , Imageamento por Ressonância Magnética , Masculino , Camundongos , Camundongos Knockout , Microscopia de Vídeo , Transfecção
6.
Pediatr Nephrol ; 38(2): 605-609, 2023 02.
Artigo em Inglês | MEDLINE | ID: mdl-35695966

RESUMO

BACKGROUND: Bardet-Biedl syndrome (BBS) is a rare, autosomal recessive ciliopathy characterized by early onset retinal dystrophy, renal anomalies, postaxial polydactyly, and cognitive impairment with considerable phenotypic heterogeneity. BBS results from biallelic pathogenic variants in over 20 genes that encode key proteins required for the assembly or primary ciliary functions of the BBSome, a heterooctameric protein complex critical for homeostasis of primary cilia. While variants in BBS1 are most frequently identified in affected individuals, the renal and pulmonary phenotypes associated with BBS1 variants are reportedly less severe than those seen in affected individuals with pathogenic variants in the other BBS-associated genes. CASE-DIAGNOSIS: We report an infant with severe renal dysplasia and lethal pulmonary hypoplasia who was homozygous for the most common BBS1 pathogenic variant (c.1169 T > G; p.M390R) and also carried a predicted pathogenic variant in TTC21B (c.1846C > T; p.R616C), a genetic modifier of disease severity of ciliopathies associated with renal dysplasia and pulmonary hypoplasia. CONCLUSIONS: This report expands the phenotypic spectrum of BBS with the first infant with lethal neonatal respiratory failure associated with biallelic, pathogenic variants in BBS1 and a monoallelic, predicted pathogenic variant in TTC21B. BBS should be considered among the ciliopathies in the differential diagnosis of neonates with renal dysplasia and severe respiratory failure.


Assuntos
Síndrome de Bardet-Biedl , Insuficiência Respiratória , Humanos , Proteínas Associadas aos Microtúbulos/genética , Proteínas Associadas aos Microtúbulos/metabolismo , Mutação , Fenótipo
7.
Am J Hum Genet ; 103(6): 968-975, 2018 12 06.
Artigo em Inglês | MEDLINE | ID: mdl-30414627

RESUMO

Wiedemann-Rautenstrauch syndrome (WRS), also known as neonatal progeroid syndrome, is a rare disorder of unknown etiology. It has been proposed to be autosomal-recessive and is characterized by variable clinical features, such as intrauterine growth restriction and poor postnatal weight gain, characteristic facial features (triangular appearance to the face, convex nasal profile or pinched nose, and small mouth), widened fontanelles, pseudohydrocephalus, prominent scalp veins, lipodystrophy, and teeth abnormalities. A previous report described a single WRS patient with bi-allelic truncating and splicing variants in POLR3A. Here we present seven additional infants, children, and adults with WRS and bi-allelic truncating and/or splicing variants in POLR3A. POLR3A, the largest subunit of RNA polymerase III, is a DNA-directed RNA polymerase that transcribes many small noncoding RNAs that regulate transcription, RNA processing, and translation. Bi-allelic missense variants in POLR3A have been associated with phenotypes distinct from WRS: hypogonadotropic hypogonadism and hypomyelinating leukodystrophy with or without oligodontia. Our findings confirm the association of bi-allelic POLR3A variants with WRS, expand the clinical phenotype of WRS, and suggest specific POLR3A genotypes associated with WRS and hypomyelinating leukodystrophy.


Assuntos
Retardo do Crescimento Fetal/genética , Variação Genética/genética , Perda de Heterozigosidade/genética , Progéria/genética , RNA Polimerase III/genética , Adolescente , Adulto , Alelos , Pré-Escolar , Feminino , Genótipo , Humanos , Fenótipo , Adulto Jovem
8.
Genet Med ; 23(6): 1075-1085, 2021 06.
Artigo em Inglês | MEDLINE | ID: mdl-33580225

RESUMO

PURPOSE: Genomic sequencing has become an increasingly powerful and relevant tool to be leveraged for the discovery of genetic aberrations underlying rare, Mendelian conditions. Although the computational tools incorporated into diagnostic workflows for this task are continually evolving and improving, we nevertheless sought to investigate commonalities across sequencing processing workflows to reveal consensus and standard practice tools and highlight exploratory analyses where technical and theoretical method improvements would be most impactful. METHODS: We collected details regarding the computational approaches used by a genetic testing laboratory and 11 clinical research sites in the United States participating in the Undiagnosed Diseases Network via meetings with bioinformaticians, online survey forms, and analyses of internal protocols. RESULTS: We found that tools for processing genomic sequencing data can be grouped into four distinct categories. Whereas well-established practices exist for initial variant calling and quality control steps, there is substantial divergence across sites in later stages for variant prioritization and multimodal data integration, demonstrating a diversity of approaches for solving the most mysterious undiagnosed cases. CONCLUSION: The largest differences across diagnostic workflows suggest that advances in structural variant detection, noncoding variant interpretation, and integration of additional biomedical data may be especially promising for solving chronically undiagnosed cases.


Assuntos
Genômica , Doenças não Diagnosticadas , Biologia Computacional , Testes Genéticos , Genoma , Humanos , Software , Fluxo de Trabalho
9.
J Hum Genet ; 66(7): 717-724, 2021 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-33517344

RESUMO

HCFC1, a global transcriptional regulator, has been shown to associate with MMACHC expression. Pathogenic variants in HCFC1 cause X-linked combined methylmalonic acidemia and hyperhomocysteinemia, CblX type (MIM# 309541). Recent studies showed that certain variants in HCFC1 are associated with X-linked intellectual disability with mild or absent metabolic abnormalities. Here, we report five subjects (three males, two females) from the same family with a novel predicted loss of function HCFC1 variant. All five patients exhibit developmental delay or intellectual disability/learning difficulty and some dysmorphic features; findings were milder in the female as compared to male subjects. Biochemical studies in all patients did not show methylmalonic acidemia or hyperhomocysteinemia but revealed elevated vitamin B12 levels. Trio exome sequencing of the proband and his parents revealed a maternally inherited novel variant in HCFC1 designated as c.1781_1803 + 3del26insCA (NM_005334). Targeted testing confirmed the presence of the same variant in two half-siblings and maternal great uncle. In silico analysis showed that the variant is expected to reduce the quality of the splice donor site in intron 10 and causes abnormal splicing. Sequencing of proband's cDNA revealed exon 10 skipping. Further molecular studies in the two manifesting females revealed moderate and high skewing of X inactivation. Our results support previous observation that HCFC1 variants located outside the Kelch domain exhibit dissociation of the clinical and biochemical phenotype and cause milder or no metabolic changes. We also show that this novel variant can be associated with a phenotype in females, although with milder severity, but further studies are needed to understand the role of skewed X inactivation among females in this rare disorder. Our work expands the genotypes and phenotypes associated with HCFC1-related disorder.


Assuntos
Erros Inatos do Metabolismo dos Aminoácidos/genética , Predisposição Genética para Doença , Fator C1 de Célula Hospedeira/genética , Inativação do Cromossomo X/genética , Adulto , Erros Inatos do Metabolismo dos Aminoácidos/patologia , Criança , Pré-Escolar , Éxons/genética , Feminino , Regulação da Expressão Gênica/genética , Estudos de Associação Genética , Doenças Genéticas Ligadas ao Cromossomo X/genética , Doenças Genéticas Ligadas ao Cromossomo X/patologia , Humanos , Lactente , Deficiência Intelectual/complicações , Deficiência Intelectual/genética , Deficiência Intelectual/patologia , Íntrons/genética , Masculino , Herança Materna/genética , Pessoa de Meia-Idade , Mutação/genética , Linhagem , Fenótipo , Sítios de Splice de RNA/genética , Splicing de RNA/genética , Vitamina B 12/genética , Sequenciamento do Exoma
10.
Am J Med Genet A ; 185(2): 544-548, 2021 02.
Artigo em Inglês | MEDLINE | ID: mdl-33184947

RESUMO

Chromodomain helicase DNA-binding protein 7 (CHD7) pathogenic variants are identified in more than 90% of infants and children with CHARGE (Coloboma of the iris, retina, and/or optic disk; congenital Heart defects, choanal Atresia, Retardation of growth and development, Genital hypoplasia, and characteristic outer and inner Ear anomalies and deafness) syndrome. Approximately, 10% of cases have no known genetic cause identified. We report a male child with clinical features of CHARGE syndrome and nondiagnostic genetic testing that included chromosomal microarray, CHD7 sequencing and deletion/duplication analysis, SEMA3E sequencing, and trio exome and whole-genome sequencing (WGS). We used a comprehensive clinical assessment, genome-wide methylation analysis (GMA), reanalysis of WGS data, and CHD7 RNA studies to discover a novel variant that causes CHD7 haploinsufficiency. The 7-year-old Hispanic male proband has typical phenotypic features of CHARGE syndrome. GMA revealed a CHD7-associated epigenetic signature. Reanalysis of the WGS data with focused bioinformatic analysis of CHD7 detected a novel, de novo 15 base pair deletion in Intron 4 of CHD7 (c.2239-20_2239-6delGTCTTGGGTTTTTGT [NM_017780.3]). Using proband RNA, we confirmed that this novel deletion causes CHD7 haploinsufficiency by disrupting the canonical 3' splice site and introducing a premature stop codon. Integrated genomic, epigenomic, and transcriptome analyses discovered a novel CHD7 variant that causes CHARGE syndrome.


Assuntos
Síndrome CHARGE/genética , Atresia das Cóanas/genética , Coloboma/genética , DNA Helicases/genética , Proteínas de Ligação a DNA/genética , Cardiopatias Congênitas/genética , Síndrome CHARGE/complicações , Síndrome CHARGE/patologia , Criança , Pré-Escolar , Atresia das Cóanas/complicações , Atresia das Cóanas/patologia , Coloboma/complicações , Coloboma/patologia , Cardiopatias Congênitas/complicações , Cardiopatias Congênitas/patologia , Humanos , Lactente , Íntrons/genética , Masculino , Mutação/genética , Fenótipo , Sequenciamento do Exoma
11.
Am J Med Genet A ; 185(7): 2190-2197, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-33931933

RESUMO

Spinal muscular atrophy with congenital bone fractures 2 (SMABF2), a type of arthrogryposis multiplex congenita (AMC), is characterized by congenital joint contractures, prenatal fractures of long bones, and respiratory distress and results from biallelic variants in ASCC1. Here, we describe an infant with severe, diffuse hypotonia, congenital contractures, and pulmonary hypoplasia characteristic of SMABF2, with the unique features of cleft palate, small spleen, transverse liver, and pulmonary thromboemboli with chondroid appearance. This infant also had impaired coagulation with diffuse petechiae and ecchymoses which has only been reported in one other infant with AMC. Using trio whole genome sequencing, our proband was identified to have biallelic variants in ASCC1. Using deep next generation sequencing of parental cDNA, we characterized alteration of splicing encoded by the novel, maternally inherited ASCC1 variant (c.297-8 T > G) which provides a mechanism for functional pathogenicity. The paternally inherited ASCC1 variant is a rare nonsense variant (c.466C > T; p.Arg156*) that has been previously identified in one other infant with AMC. This report extends the phenotypic characteristics of ASCC1-associated AMC (SMABF2) and describes a novel intronic variant that partially disrupts RNA splicing.


Assuntos
Artrogripose/genética , Proteínas de Transporte/genética , Atrofia Muscular Espinal/genética , Artrogripose/diagnóstico por imagem , Artrogripose/fisiopatologia , Códon sem Sentido/genética , Feminino , Humanos , Recém-Nascido , Atrofia Muscular Espinal/diagnóstico por imagem , Atrofia Muscular Espinal/fisiopatologia , Sequenciamento Completo do Genoma
12.
Hum Mutat ; 41(7): 1298-1307, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32196812

RESUMO

ABCA3 transports phospholipids across lamellar body membranes in pulmonary alveolar type II cells and is required for surfactant assembly. Rare, biallelic, pathogenic ABCA3 variants result in lethal neonatal respiratory distress syndrome and childhood interstitial lung disease. Qualitative functional characterization of ABCA3 missense variants suggests two pathogenic classes: disrupted intracellular trafficking (type I mutant) or impaired ATPase-mediated phospholipid transport into the lamellar bodies (type II mutant). We qualitatively compared wild-type (WT-ABCA3) with four uncharacterized ABCA3 variants (c.418A>C;p.Asn140His, c.3609_3611delCTT;p.Phe1203del, c.3784A>G;p.Ser1262Gly, and c.4195G>A;p.Val1399Met) in A549 cells using protein processing, colocalization with intracellular organelles, lamellar body ultrastructure, and ATPase activity. We quantitatively measured lamellar body-like vesicle diameter and intracellular ABCA3 trafficking using fluorescence-based colocalization. Three ABCA3 variants (p.Asn140His, p.Ser1262Gly, and p.Val1399Met) were processed and trafficked normally and demonstrated well-organized lamellar body-like vesicles, but had reduced ATPase activity consistent with type II mutants. P.Phe1203del was processed normally, had reduced ATPase activity, and well-organized lamellar body-like vesicles, but quantitatively colocalized with both endoplasmic reticulum and lysosomal markers, an intermediate phenotype suggesting disruption of both intracellular trafficking and phospholipid transport. All ABCA3 mutants demonstrated mean vesicle diameters smaller than WT-ABCA3. Qualitative and quantitative functional characterization of ABCA3 variants informs mechanisms of pathogenicity.


Assuntos
Transportadores de Cassetes de Ligação de ATP/genética , Células A549 , Vesículas Citoplasmáticas , Humanos , Doenças Pulmonares Intersticiais/genética , Mutação de Sentido Incorreto , Alvéolos Pulmonares , Surfactantes Pulmonares
13.
Am J Respir Cell Mol Biol ; 63(4): 436-443, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32692933

RESUMO

Rare or private, biallelic variants in the ABCA3 (ATP-binding cassette transporter A3) gene are the most common monogenic cause of lethal neonatal respiratory failure and childhood interstitial lung disease. Functional characterization of fewer than 10% of over 200 disease-associated ABCA3 variants (majority missense) suggests either disruption of ABCA3 protein trafficking (type I) or of ATPase-mediated phospholipid transport (type II). Therapies remain limited and nonspecific. A scalable platform is required for functional characterization of ABCA3 variants and discovery of pharmacologic correctors. To address this need, we first silenced the endogenous ABCA3 locus in A549 cells with CRISPR/Cas9 genome editing. Next, to generate a parent cell line (A549/ABCA3-/-) with a single recombination target site for genomic integration and stable expression of individual ABCA3 missense variant cDNAs, we used lentiviral-mediated integration of a LoxFAS cassette, FACS, and dilutional cloning. To assess the fidelity of this cell-based model, we compared functional characterization (ABCA3 protein processing, ABCA3 immunofluorescence colocalization with intracellular markers, ultrastructural vesicle phenotype) of two individual ABCA3 mutants (type I mutant, p.L101P; type II mutant, p.E292V) in A549/ABCA3-/- cells and in both A549 cells and primary, human alveolar type II cells that transiently express each cDNA after adenoviral-mediated transduction. We also confirmed pharmacologic rescue of ABCA3 variant-encoded mistrafficking and vesicle diameter in A549/ABCA3-/- cells that express p.G1421R (type I mutant). A549/ABCA3-/- cells provide a scalable, genetically versatile, physiologically relevant functional genomics platform for discovery of variant-specific mechanisms that disrupt ABCA3 function and for screening of potential ABCA3 pharmacologic correctors.


Assuntos
Transportadores de Cassetes de Ligação de ATP/genética , Genoma/genética , Mutação de Sentido Incorreto/genética , Células A549 , Adenosina Trifosfatases/genética , Células Epiteliais Alveolares/metabolismo , Sistemas CRISPR-Cas/genética , Linhagem Celular Tumoral , DNA Complementar/genética , Imunofluorescência/métodos , Edição de Genes/métodos , Genômica/métodos , Humanos , Pulmão/metabolismo , Doenças Pulmonares Intersticiais/genética
14.
Am J Med Genet A ; 182(5): 1053-1065, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32083401

RESUMO

Pathogenic variants in KMT2D, which encodes lysine specific methyltransferase 2D, cause autosomal dominant Kabuki syndrome, associated with distinctive dysmorphic features including arched eyebrows, long palpebral fissures with eversion of the lower lid, large protuberant ears, and fetal finger pads. Most disease-causing variants identified to date are putative loss-of-function alleles, although 15-20% of cases are attributed to missense variants. We describe here four patients (including one previously published patient) with de novo KMT2D missense variants and with shared but unusual clinical findings not typically seen in Kabuki syndrome, including athelia (absent nipples), choanal atresia, hypoparathyroidism, delayed or absent pubertal development, and extreme short stature. These individuals also lack the typical dysmorphic facial features found in Kabuki syndrome. Two of the four patients had severe interstitial lung disease. All of these variants cluster within a 40-amino-acid region of the protein that is located just N-terminal of an annotated coiled coil domain. These findings significantly expand the phenotypic spectrum of features associated with variants in KMT2D beyond those seen in Kabuki syndrome and suggest a possible new underlying disease mechanism for these patients.


Assuntos
Anormalidades Múltiplas/genética , Mama/anormalidades , Anormalidades Congênitas/genética , Proteínas de Ligação a DNA/genética , Face/anormalidades , Predisposição Genética para Doença , Doenças Hematológicas/genética , Proteínas de Neoplasias/genética , Doenças Vestibulares/genética , Anormalidades Múltiplas/diagnóstico por imagem , Anormalidades Múltiplas/patologia , Adolescente , Adulto , Mama/diagnóstico por imagem , Mama/fisiopatologia , Doenças Mamárias , Criança , Anormalidades Congênitas/diagnóstico por imagem , Anormalidades Congênitas/fisiopatologia , Face/diagnóstico por imagem , Face/patologia , Feminino , Doenças Hematológicas/diagnóstico por imagem , Doenças Hematológicas/patologia , Humanos , Mutação com Perda de Função/genética , Masculino , Mutação/genética , Fenótipo , Doenças Vestibulares/diagnóstico por imagem , Doenças Vestibulares/patologia , Sequenciamento do Exoma , Adulto Jovem
16.
J Pediatr ; 194: 158-164.e1, 2018 03.
Artigo em Inglês | MEDLINE | ID: mdl-29198536

RESUMO

OBJECTIVE: To describe disease course, histopathology, and outcomes for infants with atypical presentations of alveolar capillary dysplasia with misalignment of the pulmonary veins (ACDMPV) who underwent bilateral lung transplantation. STUDY DESIGN: We reviewed clinical history, diagnostic studies, explant histology, genetic sequence results, and post-transplant course for 6 infants with atypical ACDMPV who underwent bilateral lung transplantation at St. Louis Children's Hospital. We compared their histology with infants with classic ACDMPV and compared their outcomes with infants transplanted for other indications. RESULTS: In contrast with neonates with classic ACDPMV who present with severe hypoxemia and refractory pulmonary hypertension within hours of birth, none of the infants with atypical ACDMPV presented with progressive neonatal respiratory failure. Three infants had mild neonatal respiratory distress and received nasal cannula oxygen. Three other infants had no respiratory symptoms at birth and presented with hypoxemia and pulmonary hypertension at 2-3 months of age. Bilateral lung transplantation was performed at 4-20 months of age. Unlike in classic ACDMPV, histopathologic findings were not distributed uniformly and were not diffuse. Three subjects had apparent nonmosaic genetic defects involving FOXF1. Two infants had extrapulmonary anomalies (posterior urethral valves, inguinal hernia). Three transplanted children are alive at 5-16 years of age, similar to outcomes for infants transplanted for other indications. Lung explants from infants with atypical ACDMPV demonstrated diagnostic but nonuniform histopathologic findings. CONCLUSIONS: The 1- and 5-year survival rates for infants with atypical ACDMPV are similar to infants transplanted for other indications. Given the clinical and histopathologic spectra, ACDMPV should be considered in infants with hypoxemia and pulmonary hypertension, even beyond the newborn period.


Assuntos
Transplante de Pulmão/métodos , Síndrome da Persistência do Padrão de Circulação Fetal/diagnóstico , Alvéolos Pulmonares/anormalidades , Feminino , Fatores de Transcrição Forkhead/genética , Humanos , Lactente , Recém-Nascido , Pulmão/patologia , Masculino , Mutação , Síndrome da Persistência do Padrão de Circulação Fetal/complicações , Síndrome da Persistência do Padrão de Circulação Fetal/cirurgia , Alvéolos Pulmonares/cirurgia , Veias Pulmonares/anormalidades , Taxa de Sobrevida
17.
Pediatr Res ; 84(3): 435-441, 2018 09.
Artigo em Inglês | MEDLINE | ID: mdl-29967526

RESUMO

BACKGROUND: Biallelic deleterious variants in RTTN, which encodes rotatin, are associated with primary microcephaly, polymicrogyria, seizures, intellectual disability, and primordial dwarfism in human infants. METHODS AND RESULTS: We performed exome sequencing of an infant with primary microcephaly, pontocerebellar hypoplasia, and intractable seizures and his healthy, unrelated parents. We cultured the infant's fibroblasts to determine primary ciliary phenotype. RESULTS: We identified biallelic variants in RTTN in the affected infant: a novel missense variant and a rare, intronic variant that results in aberrant transcript splicing. Cultured fibroblasts from the infant demonstrated reduced length and number of primary cilia. CONCLUSION: Biallelic variants in RTTN cause primary microcephaly in infants. Functional characterization of primary cilia length and number can be used to determine pathogenicity of RTTN variants.


Assuntos
Encéfalo/anormalidades , Proteínas de Transporte/genética , Doenças Cerebelares/genética , Microcefalia/genética , Convulsões/genética , Alelos , Encéfalo/diagnóstico por imagem , Proteínas de Ciclo Celular , Cílios , Exoma , Evolução Fatal , Fibroblastos/metabolismo , Deleção de Genes , Variação Genética , Humanos , Lactente , Imageamento por Ressonância Magnética , Masculino , Mutação de Sentido Incorreto , Fenótipo , Insuficiência Respiratória
18.
Hum Mutat ; 38(11): 1477-1484, 2017 11.
Artigo em Inglês | MEDLINE | ID: mdl-28726266

RESUMO

Biallelic GLDN mutations have recently been identified among infants with lethal congenital contracture syndrome 11 (LCCS11). GLDN encodes gliomedin, a protein required for the formation of the nodes of Ranvier and development of the human peripheral nervous system. We report six infants and children from four unrelated families with biallelic GLDN mutations, four of whom survived beyond the neonatal period into infancy, childhood, and late adolescence with intensive care and chronic respiratory and nutritional support. Our findings expand the genotypic and phenotypic spectrum of LCCS11 and demonstrate that the condition may not necessarily be lethal in the neonatal period.


Assuntos
Artrogripose/diagnóstico , Artrogripose/genética , Genes Letais , Proteínas de Membrana/genética , Mutação , Proteínas do Tecido Nervoso/genética , Fenótipo , Artrogripose/mortalidade , Biópsia , Análise Mutacional de DNA , Evolução Fatal , Estudos de Associação Genética , Humanos , Lactente , Recém-Nascido , Masculino , Linhagem , Raízes Nervosas Espinhais/ultraestrutura , Sequenciamento do Exoma
19.
Am J Respir Cell Mol Biol ; 55(5): 716-721, 2016 11.
Artigo em Inglês | MEDLINE | ID: mdl-27374344

RESUMO

Mutations in the ATP-binding cassette transporter A3 gene (ABCA3) result in severe neonatal respiratory distress syndrome and childhood interstitial lung disease. As most ABCA3 mutations are rare or private, determination of mutation pathogenicity is often based on results from in silico prediction tools, identification in unrelated diseased individuals, statistical association studies, or expert opinion. Functional biologic studies of ABCA3 mutations are needed to confirm mutation pathogenicity and inform clinical decision making. Our objective was to functionally characterize two ABCA3 mutations (p.R288K and p.R1474W) identified among term and late-preterm infants with respiratory distress syndrome with unclear pathogenicity in a genetically versatile model system. We performed transient transfection of HEK293T cells with wild-type or mutant ABCA3 alleles to assess protein processing with immunoblotting. We used transduction of A549 cells with adenoviral vectors, which concurrently silenced endogenous ABCA3 and expressed either wild-type or mutant ABCA3 alleles (p.R288K and p.R1474W) to assess immunofluorescent localization, ATPase activity, and organelle ultrastructure. Both ABCA3 mutations (p.R288K and p.R1474W) encoded proteins with reduced ATPase activity but with normal intracellular localization and protein processing. Ultrastructural phenotypes of lamellar body-like vesicles in A549 cells transduced with mutant alleles were similar to wild type. Mutant proteins encoded by ABCA3 mutations p.R288K and p.R1474W had reduced ATPase activity, a biologically plausible explanation for disruption of surfactant metabolism by impaired phospholipid transport into the lamellar body. These results also demonstrate the usefulness of a genetically versatile, human model system for functional characterization of ABCA3 mutations with unclear pathogenicity.


Assuntos
Transportadores de Cassetes de Ligação de ATP/genética , Mutação/genética , Síndrome do Desconforto Respiratório do Recém-Nascido/genética , Células A549 , Adenosina Trifosfatases/metabolismo , Adenoviridae/metabolismo , Imunofluorescência , Células HEK293 , Humanos , Immunoblotting , Lactente , Proteínas Mutantes/metabolismo , Organelas/metabolismo , Organelas/ultraestrutura , Frações Subcelulares/metabolismo
20.
Am J Respir Crit Care Med ; 189(12): 1538-43, 2014 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-24871971

RESUMO

RATIONALE: Recessive mutations in the ATP-binding cassette transporter A3 (ABCA3) cause lethal neonatal respiratory failure and childhood interstitial lung disease. Most ABCA3 mutations are private. OBJECTIVES: To determine genotype-phenotype correlations for recessive ABCA3 mutations. METHODS: We reviewed all published and unpublished ABCA3 sequence and phenotype data from our prospective genetic studies of symptomatic infants and children at Washington and Johns Hopkins Universities. Mutations were classified based on their predicted disruption of protein function: frameshift and nonsense mutations were classified as "null," whereas missense, predicted splice site mutations, and insertion/deletions were classified as "other." We compared age of presentation and outcomes for the three genotypes: null/null, null/other, and other/other. MEASUREMENTS AND MAIN RESULTS: We identified 185 infants and children with homozygous or compound heterozygous ABCA3 mutations and lung disease. All of the null/null infants presented with respiratory failure at birth compared with 75% of infants with null/other or other/other genotypes (P = 0.00011). By 1 year of age, all of the null/null infants had died or undergone lung transplantation compared with 62% of the null/other and other/other children (P < 0.0001). CONCLUSIONS: Genotype-phenotype correlations exist for homozygous or compound heterozygous mutations in ABCA3. Frameshift or nonsense ABCA3 mutations are predictive of neonatal presentation and poor outcome, whereas missense, splice site, and insertion/deletions are less reliably associated with age of presentation and prognosis. Counseling and clinical decision making should acknowledge these correlations.


Assuntos
Transportadores de Cassetes de Ligação de ATP/deficiência , Estudos de Associação Genética , Doenças Pulmonares Intersticiais/genética , Mutação , Síndrome do Desconforto Respiratório do Recém-Nascido/genética , Transportadores de Cassetes de Ligação de ATP/genética , Criança , Pré-Escolar , Feminino , Marcadores Genéticos , Heterozigoto , Homozigoto , Humanos , Lactente , Recém-Nascido , Doenças Pulmonares Intersticiais/mortalidade , Doenças Pulmonares Intersticiais/cirurgia , Transplante de Pulmão , Masculino , Síndrome do Desconforto Respiratório do Recém-Nascido/mortalidade , Síndrome do Desconforto Respiratório do Recém-Nascido/cirurgia , Análise de Sequência de DNA
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