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1.
JCI Insight ; 2024 Jul 11.
Artigo em Inglês | MEDLINE | ID: mdl-38990653

RESUMO

The Neurofibromatosis Type 1 (NF1) RASopathy is associated with persistent fibrotic nonunions (pseudarthrosis) in human and mouse skeletal tissue. Here, we first performed spatial transcriptomics to define the molecular signatures across normal endochondral healing following fracture in mice. Within the control fracture callus, we observed spatially restricted activation of morphogenetic pathways, such as TGF-ß, WNT, and BMP. To investigate the molecular mechanisms contributing to Nf1-deficient delayed fracture healing, we performed spatial transcriptomic analysis on a Postn-cre;Nf1flox/- (Nf1Postn) fracture callus. Transcriptional analyses, subsequently confirmed through p-SMAD1/5/8 immunohistochemistry, demonstrated a lack of BMP pathway induction in Nf1Postn mice. To further inform the human disease, we performed spatial transcriptomic analysis of fracture pseudarthrosis tissue from a NF1 patient. Analyses detected increased MAPK signaling at the fibrocartilaginous-osseus junction. Similar to the Nf1Postn fracture, BMP pathway activation was absent within the pseudarthrosis tissue. Our results demonstrate the feasibility to delineate the molecular and tissue-specific heterogeneity inherent in complex regenerative processes, such as fracture healing, and to reconstruct phase transitions representing endochondral bone formation in vivo. Furthermore, our results provide in situ molecular evidence of impaired BMP signaling underlying NF1 pseudarthrosis, potentially informing the clinical relevance of off-label BMP2 as a therapeutic intervention.

2.
Elife ; 122024 Jan 26.
Artigo em Inglês | MEDLINE | ID: mdl-38277211

RESUMO

Adolescent idiopathic scoliosis (AIS) is a common and progressive spinal deformity in children that exhibits striking sexual dimorphism, with girls at more than fivefold greater risk of severe disease compared to boys. Despite its medical impact, the molecular mechanisms that drive AIS are largely unknown. We previously defined a female-specific AIS genetic risk locus in an enhancer near the PAX1 gene. Here, we sought to define the roles of PAX1 and newly identified AIS-associated genes in the developmental mechanism of AIS. In a genetic study of 10,519 individuals with AIS and 93,238 unaffected controls, significant association was identified with a variant in COL11A1 encoding collagen (α1) XI (rs3753841; NM_080629.2_c.4004C>T; p.(Pro1335Leu); p=7.07E-11, OR = 1.118). Using CRISPR mutagenesis we generated Pax1 knockout mice (Pax1-/-). In postnatal spines we found that PAX1 and collagen (α1) XI protein both localize within the intervertebral disc-vertebral junction region encompassing the growth plate, with less collagen (α1) XI detected in Pax1-/- spines compared to wild-type. By genetic targeting we found that wild-type Col11a1 expression in costal chondrocytes suppresses expression of Pax1 and of Mmp3, encoding the matrix metalloproteinase 3 enzyme implicated in matrix remodeling. However, the latter suppression was abrogated in the presence of the AIS-associated COL11A1P1335L mutant. Further, we found that either knockdown of the estrogen receptor gene Esr2 or tamoxifen treatment significantly altered Col11a1 and Mmp3 expression in chondrocytes. We propose a new molecular model of AIS pathogenesis wherein genetic variation and estrogen signaling increase disease susceptibility by altering a PAX1-COL11a1-MMP3 signaling axis in spinal chondrocytes.


Adolescent idiopathic scoliosis (AIS) is a twisting deformity of the spine that occurs during periods of rapid growth in children worldwide. Children with severe cases of AIS require surgery to stop it from getting worse, presenting a significant financial burden to health systems and families. Although AIS is known to cluster in families, its genetic causes and its inheritance pattern have remained elusive. Additionally, AIS is known to be more prevalent in females, a bias that has not been explained. Advances in techniques to study the genetics underlying diseases have revealed that certain variations that increase the risk of AIS affect cartilage and connective tissue. In humans, one such variation is near a gene called Pax1, and it is female-specific. The extracellular matrix is a network of proteins and other molecules in the space between cells that help connect tissues together, and it is particularly important in cartilage and other connective tissues. One of the main components of the extracellular matrix is collagen. Yu, Kanshour, Ushiki et al. hypothesized that changes in the extracellular matrix could affect the cartilage and connective tissues of the spine, leading to AIS. To show this, the scientists screened over 100,000 individuals and found that AIS is associated with variants in two genes coding for extracellular matrix proteins. One of these variants was found in a gene called Col11a1, which codes for one of the proteins that makes up collagen. To understand the relationship between Pax1 and Col11a1, Yu, Kanshour, Ushiki et al. genetically modified mice so that they would lack the Pax1 gene. In these mice, the activation of Col11a1 was reduced in the mouse spine. They also found that the form of Col11a1 associated with AIS could not suppress the activation of a gene called Mmp3 in mouse cartilage cells as effectively as unmutated Col11a1. Going one step further, the researchers found that lowering the levels of an estrogen receptor altered the activation patterns of Pax1, Col11a1, and Mmp3 in mouse cartilage cells. These findings suggest a possible mechanism for AIS, particularly in females. The findings of Yu, Kanshour, Ushiki et al. highlight that cartilage cells in the spine are particularly relevant in AIS. The results also point to specific molecules within the extracellular matrix as important for maintaining proper alignment in the spine when children are growing rapidly. This information may guide future therapies aimed at maintaining healthy spinal cells in adolescent children, particularly girls.


Assuntos
Escoliose , Masculino , Animais , Criança , Camundongos , Humanos , Feminino , Adolescente , Escoliose/genética , Metaloproteinase 3 da Matriz/genética , Coluna Vertebral , Fatores de Transcrição/genética , Colágeno/genética , Variação Genética , Colágeno Tipo XI/genética
3.
J Clin Invest ; 134(2)2024 Jan 16.
Artigo em Inglês | MEDLINE | ID: mdl-37962965

RESUMO

Adolescent idiopathic scoliosis (AIS) is the most common form of spinal deformity, affecting millions of adolescents worldwide, but it lacks a defined theory of etiopathogenesis. Because of this, treatment of AIS is limited to bracing and/or invasive surgery after onset. Preonset diagnosis or preventive treatment remains unavailable. Here, we performed a genetic analysis of a large multicenter AIS cohort and identified disease-causing and predisposing variants of SLC6A9 in multigeneration families, trios, and sporadic patients. Variants of SLC6A9, which encodes glycine transporter 1 (GLYT1), reduced glycine-uptake activity in cells, leading to increased extracellular glycine levels and aberrant glycinergic neurotransmission. Slc6a9 mutant zebrafish exhibited discoordination of spinal neural activities and pronounced lateral spinal curvature, a phenotype resembling human patients. The penetrance and severity of curvature were sensitive to the dosage of functional glyt1. Administration of a glycine receptor antagonist or a clinically used glycine neutralizer (sodium benzoate) partially rescued the phenotype. Our results indicate a neuropathic origin for "idiopathic" scoliosis, involving the dysfunction of synaptic neurotransmission and central pattern generators (CPGs), potentially a common cause of AIS. Our work further suggests avenues for early diagnosis and intervention of AIS in preadolescents.


Assuntos
Escoliose , Animais , Humanos , Adolescente , Escoliose/genética , Escoliose/diagnóstico , Escoliose/cirurgia , Glicina/genética , Peixe-Zebra , Transmissão Sináptica
4.
Am J Hum Genet ; 110(12): 2103-2111, 2023 Dec 07.
Artigo em Inglês | MEDLINE | ID: mdl-37924809

RESUMO

Hereditary spastic parapareses (HSPs) are clinically heterogeneous motor neuron diseases with variable age of onset and severity. Although variants in dozens of genes are implicated in HSPs, much of the genetic basis for pediatric-onset HSP remains unexplained. Here, we re-analyzed clinical exome-sequencing data from siblings with HSP of unknown genetic etiology and identified an inherited nonsense mutation (c.523C>T [p.Arg175Ter]) in the highly conserved RAB1A. The mutation is predicted to produce a truncated protein with an intact RAB GTPase domain but without two C-terminal cysteine residues required for proper subcellular protein localization. Additional RAB1A mutations, including two frameshift mutations and a mosaic missense mutation (c.83T>C [p.Leu28Pro]), were identified in three individuals with similar neurodevelopmental presentations. In rescue experiments, production of the full-length, but not the truncated, RAB1a rescued Golgi structure and cell proliferation in Rab1-depleted cells. In contrast, the missense-variant RAB1a disrupted Golgi structure despite intact Rab1 expression, suggesting a dominant-negative function of the mosaic missense mutation. Knock-down of RAB1A in cultured human embryonic stem cell-derived neurons resulted in impaired neuronal arborization. Finally, RAB1A is located within the 2p14-p15 microdeletion syndrome locus. The similar clinical presentations of individuals with RAB1A loss-of-function mutations and the 2p14-p15 microdeletion syndrome implicate loss of RAB1A in the pathogenesis of neurodevelopmental manifestations of this microdeletion syndrome. Our study identifies a RAB1A-related neurocognitive disorder with speech and motor delay, demonstrates an essential role for RAB1a in neuronal differentiation, and implicates RAB1A in the etiology of the neurodevelopmental sequelae associated with the 2p14-p15 microdeletion syndrome.


Assuntos
Haploinsuficiência , Paraplegia Espástica Hereditária , Criança , Humanos , Haploinsuficiência/genética , Mutação , Mutação de Sentido Incorreto/genética , Proteínas rab de Ligação ao GTP/genética , Proteínas rab de Ligação ao GTP/metabolismo , Complexo de Golgi/metabolismo , Paraplegia Espástica Hereditária/genética
5.
J Pediatr Orthop ; 43(10): 615-619, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-37694695

RESUMO

BACKGROUND: Socioeconomic disparities in musculoskeletal care are increasingly recognized, however, no studies to date have investigated the role of the insurance carrier on outcomes after posterior spinal fusion (PSF) with segmental spinal instrumentation for adolescent idiopathic scoliosis (AIS). METHODS: A US insurance dataset was queried using the PearlDiver Mariner software for all patients aged 10 to 18 undergoing PSF for a primary diagnosis of AIS between 2010 and 2020. Age, sex, geographic region, number of levels fused, and baseline medical comorbidities were queried. Complications occurring within 90 days of the index surgery were queried using the International Classification of Diseases, Ninth Revision (ICD-9) and International Classification of Diseases, 10th Revision (ICD-10) codes. Revision surgery was also queried up to 5 years after the index PSF. Categorical variables were compared using the Fisher χ 2 tests and continuous variables were compared using independent t tests. All-cause revision within 5 years was compared using the Kaplan-Meier analysis and a log-rank test. Significance was set at P -value <0.05. RESULTS: A total of 10,794 patients were identified with 9006 (83.4%) patients with private insurance and 1788 (16.6%) patients insured by Medicaid. The mean follow-up in the database was 5.36±3 years for patients with private insurance and 4.78±2.9 years for patients with Medicaid insurance ( P <0.001). Children with AIS and Medicaid insurance had a significantly higher prevalence of asthma, hypertension, and obesity. A larger percentage of children with Medicaid insurance (41.3%) underwent a ≥13-level PSF compared with privately insured children (34.5%) ( P <0.001). Medicaid patients did not experience higher odds of postoperative complications; in addition, revision surgeries occurred in 1.1% and 1.8% of patients with private insurance and Medicaid insurance, respectively at 5 years postoperatively ( P =0.223). CONCLUSION: Despite worse baseline comorbidities and longer fusion constructs, AIS patients insured with Medicaid did not have higher rates of complications or revisions at 5-year follow-up versus privately insured patients. LEVEL OF EVIDENCE: Level III-retrospective cohort study.


Assuntos
Escoliose , Fusão Vertebral , Adolescente , Estados Unidos/epidemiologia , Humanos , Criança , Medicaid , Estudos Retrospectivos , Cobertura do Seguro , Comorbidade , Escoliose/cirurgia , Escoliose/epidemiologia
6.
bioRxiv ; 2023 Nov 22.
Artigo em Inglês | MEDLINE | ID: mdl-37292598

RESUMO

Adolescent idiopathic scoliosis (AIS) is a common and progressive spinal deformity in children that exhibits striking sexual dimorphism, with girls at more than five-fold greater risk of severe disease compared to boys. Despite its medical impact, the molecular mechanisms that drive AIS are largely unknown. We previously defined a female-specific AIS genetic risk locus in an enhancer near the PAX1 gene. Here we sought to define the roles of PAX1 and newly-identified AIS-associated genes in the developmental mechanism of AIS. In a genetic study of 10,519 individuals with AIS and 93,238 unaffected controls, significant association was identified with a variant in COL11A1 encoding collagen (α1) XI (rs3753841; NM_080629.2_c.4004C>T; p.(Pro1335Leu); P=7.07e-11, OR=1.118). Using CRISPR mutagenesis we generated Pax1 knockout mice (Pax1-/-). In postnatal spines we found that PAX1 and collagen (α1) XI protein both localize within the intervertebral disc (IVD)-vertebral junction region encompassing the growth plate, with less collagen (α1) XI detected in Pax1-/- spines compared to wildtype. By genetic targeting we found that wildtype Col11a1 expression in costal chondrocytes suppresses expression of Pax1 and of Mmp3, encoding the matrix metalloproteinase 3 enzyme implicated in matrix remodeling. However, this suppression was abrogated in the presence of the AIS-associated COL11A1P1335L mutant. Further, we found that either knockdown of the estrogen receptor gene Esr2, or tamoxifen treatment, significantly altered Col11a1 and Mmp3 expression in chondrocytes. We propose a new molecular model of AIS pathogenesis wherein genetic variation and estrogen signaling increase disease susceptibility by altering a Pax1-Col11a1-Mmp3 signaling axis in spinal chondrocytes.

7.
J Bone Miner Res ; 38(2): 288-299, 2023 02.
Artigo em Inglês | MEDLINE | ID: mdl-36459048

RESUMO

Neurofibromatosis type 1 (NF1) is a tumor predisposition syndrome caused by heterozygous NF1 gene mutations. Patients with NF1 present with pleiotropic somatic secondary manifestations, including development of bone pseudarthrosis after fracture. Somatic NF1 gene mutations were reproducibly identified in patient-derived pseudarthrosis specimens, suggesting a local mosaic cell population including somatic pathologic cells. The somatic cellular pathogenesis of NF1 pseudarthroses remains unclear, though defects in osteogenesis have been posited. Here, we applied time-series single-cell RNA-sequencing (scRNA-seq) to patient-matched control and pseudarthrosis-derived primary bone stromal cells (BSCs). We show that osteogenic specification to an osteoblast progenitor cell population was evident for control bone-derived cells and haploinsufficient pseudarthrosis-derived cells. Similar results were observed for somatic patient fracture-derived NF1-/- cells; however, expression of genetic pathways associated with skeletal mineralization were significantly reduced in NF1-/- cells compared with fracture-derived NF1+/- cells. In mice, we show that Nf1 expressed in bone marrow osteoprogenitors is required for the maintenance of the adult skeleton. Results from our study implicate impaired Clec11a-Itga11-Wnt signaling in the pathogenesis of NF1-associated skeletal disease. © 2022 The Authors. Journal of Bone and Mineral Research published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research (ASBMR).


Assuntos
Fraturas Ósseas , Neurofibromatose 1 , Pseudoartrose , Camundongos , Animais , Neurofibromatose 1/complicações , Neurofibromatose 1/genética , Neurofibromatose 1/patologia , Pseudoartrose/genética , Pseudoartrose/metabolismo , Pseudoartrose/patologia , Fraturas Ósseas/patologia , Osteoblastos/metabolismo , Osteogênese/genética
8.
Clin Case Rep ; 8(8): 1452-1457, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32884773

RESUMO

We present the use of whole-genome sequencing to correctly diagnose progressive pseudorheumatoid dysplasia in patients with atypical clinical and radiologic findings and prior diagnosis of juvenile idiopathic arthritis.

9.
Genes Dev ; 30(19): 2158-2172, 2016 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-27737959

RESUMO

Compaction of chromosomes is essential for accurate segregation of the genome during mitosis. In vertebrates, two condensin complexes ensure timely chromosome condensation, sister chromatid disentanglement, and maintenance of mitotic chromosome structure. Here, we report that biallelic mutations in NCAPD2, NCAPH, or NCAPD3, encoding subunits of these complexes, cause microcephaly. In addition, hypomorphic Ncaph2 mice have significantly reduced brain size, with frequent anaphase chromatin bridge formation observed in apical neural progenitors during neurogenesis. Such DNA bridges also arise in condensin-deficient patient cells, where they are the consequence of failed sister chromatid disentanglement during chromosome compaction. This results in chromosome segregation errors, leading to micronucleus formation and increased aneuploidy in daughter cells. These findings establish "condensinopathies" as microcephalic disorders, with decatenation failure as an additional disease mechanism for microcephaly, implicating mitotic chromosome condensation as a key process ensuring mammalian cerebral cortex size.


Assuntos
Adenosina Trifosfatases/genética , Proteínas de Ligação a DNA/genética , Microcefalia/genética , Mitose/genética , Complexos Multiproteicos/genética , Mutação/genética , Aneuploidia , Animais , Catenanos/metabolismo , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Células Cultivadas , Instabilidade Cromossômica/genética , Segregação de Cromossomos/genética , Feminino , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Micronúcleos com Defeito Cromossômico , Neurônios/patologia , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Células-Tronco
10.
Sci Rep ; 6: 27684, 2016 06 13.
Artigo em Inglês | MEDLINE | ID: mdl-27291587

RESUMO

We report identification and genetic characterization of a rare skeletal disorder that remained unidentified for decades in a village of Jammu and Kashmir, India. The population residing in this region is highly consanguineous and a lack of understanding of the disorder has hindered clinical management and genetic counseling for the many affected individuals in the region. We collected familial information and identified two large extended multiplex pedigrees displaying apparent autosomal recessive inheritance of an uncharacterized skeletal dysplasia. Whole exome sequencing (WES) in members of one pedigree revealed a rare mutation in WISP3:c.156C > A (NP_003871.1:p.Cys52Ter), that perfectly segregated with the disease in the family. To our surprise, Sanger sequencing the WISP3 gene in the second family identified a distinct, novel splice site mutation c.643 + 1G > A, that perfectly segregated with the disease. Combining our next generation sequencing data with careful clinical documentation (familial histories, genetic data, clinical and radiological findings), we have diagnosed the families with Progressive Pseudorheumatoid Dysplasia (PPD). Our results underscore the utility of WES in arriving at definitive diagnoses for rare skeletal dysplasias. This genetic characterization will aid in genetic counseling and management, critically required to curb this rare disorder in the families.


Assuntos
Proteínas de Sinalização Intercelular CCN/genética , Exoma , Artropatias/congênito , Adulto , Criança , Consanguinidade , Feminino , Genes Recessivos , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Índia , Artropatias/etnologia , Artropatias/genética , Masculino , Pessoa de Meia-Idade , Mutação , Linhagem , Fenótipo , Adulto Jovem
11.
Am J Hum Genet ; 97(6): 837-47, 2015 Dec 03.
Artigo em Inglês | MEDLINE | ID: mdl-26637977

RESUMO

The periosteum contributes to bone repair and maintenance of cortical bone mass. In contrast to the understanding of bone development within the epiphyseal growth plate, factors that regulate periosteal osteogenesis have not been studied as intensively. Osteofibrous dysplasia (OFD) is a congenital disorder of osteogenesis and is typically sporadic and characterized by radiolucent lesions affecting the cortical bone immediately under the periosteum of the tibia and fibula. We identified germline mutations in MET, encoding a receptor tyrosine kinase, that segregate with an autosomal-dominant form of OFD in three families and a mutation in a fourth affected subject from a simplex family and with bilateral disease. Mutations identified in all families with dominant inheritance and in the one simplex subject with bilateral disease abolished the splice inclusion of exon 14 in MET transcripts, which resulted in a MET receptor (MET(Δ14)) lacking a cytoplasmic juxtamembrane domain. Splice exclusion of this domain occurs during normal embryonic development, and forced induction of this exon-exclusion event retarded osteoblastic differentiation in vitro and inhibited bone-matrix mineralization. In an additional subject with unilateral OFD, we identified a somatic MET mutation, also affecting exon 14, that substituted a tyrosine residue critical for MET receptor turnover and, as in the case of the MET(Δ14) mutations, had a stabilizing effect on the mature protein. Taken together, these data show that aberrant MET regulation via the juxtamembrane domain subverts core MET receptor functions that regulate osteogenesis within cortical diaphyseal bone.


Assuntos
Doenças do Desenvolvimento Ósseo/genética , Éxons , Mutação em Linhagem Germinativa , Osteogênese/genética , Periósteo/metabolismo , Proteínas Proto-Oncogênicas c-met/genética , Adulto , Sequência de Bases , Doenças do Desenvolvimento Ósseo/metabolismo , Doenças do Desenvolvimento Ósseo/patologia , Diferenciação Celular , Criança , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Genes Dominantes , Humanos , Masculino , Pessoa de Meia-Idade , Dados de Sequência Molecular , Osteoblastos/metabolismo , Osteoblastos/patologia , Linhagem , Periósteo/crescimento & desenvolvimento , Periósteo/patologia , Cultura Primária de Células , Estrutura Terciária de Proteína , Proteínas Proto-Oncogênicas c-met/metabolismo , Splicing de RNA
12.
J Bone Miner Res ; 29(12): 2636-42, 2014 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-24932921

RESUMO

Neurofibromatosis type 1 (NF1) is an autosomal dominant disease caused by mutations in NF1. Among the earliest manifestations is tibial pseudoarthrosis and persistent nonunion after fracture. To further understand the pathogenesis of pseudoarthrosis and the underlying bone remodeling defect, pseudoarthrosis tissue and cells cultured from surgically resected pseudoarthrosis tissue from NF1 individuals were analyzed using whole-exome and whole-transcriptome sequencing as well as genomewide microarray analysis. Genomewide analysis identified multiple genetic mechanisms resulting in somatic biallelic NF1 inactivation; no other genes with recurring somatic mutations were identified. Gene expression profiling identified dysregulated pathways associated with neurofibromin deficiency, including phosphoinositide 3-kinase (PI3K) and mitogen-activated protein kinase (MAPK) signaling pathways. Unlike aggressive NF1-associated malignancies, tibial pseudoarthrosis tissue does not harbor a high frequency of somatic mutations in oncogenes or other tumor-suppressor genes, such as p53. However, gene expression profiling indicates that pseudoarthrosis tissue has a tumor-promoting transcriptional pattern, despite lacking tumorigenic somatic mutations. Significant overexpression of specific cancer-associated genes in pseudoarthrosis highlights a potential for receptor tyrosine kinase inhibitors to target neurofibromin-deficient pseudoarthrosis and promote proper bone remodeling and fracture healing.


Assuntos
Regulação da Expressão Gênica , Neurofibromatose 1 , Neurofibromina 1/deficiência , Pseudoartrose , Fraturas da Tíbia , Transcrição Gênica , Adolescente , Remodelação Óssea/genética , Pré-Escolar , Feminino , Consolidação da Fratura/genética , Perfilação da Expressão Gênica , Humanos , Lactente , Sistema de Sinalização das MAP Quinases/genética , Masculino , Neurofibromatose 1/genética , Neurofibromatose 1/metabolismo , Neurofibromatose 1/patologia , Neurofibromatose 1/terapia , Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/metabolismo , Pseudoartrose/genética , Pseudoartrose/metabolismo , Pseudoartrose/patologia , Pseudoartrose/terapia , Fraturas da Tíbia/genética , Fraturas da Tíbia/metabolismo , Fraturas da Tíbia/patologia , Fraturas da Tíbia/terapia
14.
Hum Mutat ; 35(1): 76-85, 2014 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-24123394

RESUMO

Ligase IV syndrome is a rare differential diagnosis for Nijmegen breakage syndrome owing to a shared predisposition to lympho-reticular malignancies, significant microcephaly, and radiation hypersensitivity. Only 16 cases with mutations in LIG4 have been described to date with phenotypes varying from malignancy in developmentally normal individuals, to severe combined immunodeficiency and early mortality. Here, we report the identification of biallelic truncating LIG4 mutations in 11 patients with microcephalic primordial dwarfism presenting with restricted prenatal growth and extreme postnatal global growth failure (average OFC -10.1 s.d., height -5.1 s.d.). Subsequently, most patients developed thrombocytopenia and leucopenia later in childhood and many were found to have previously unrecognized immunodeficiency following molecular diagnosis. None have yet developed malignancy, though all patients tested had cellular radiosensitivity. A genotype-phenotype correlation was also noted with position of truncating mutations corresponding to disease severity. This work extends the phenotypic spectrum associated with LIG4 mutations, establishing that extreme growth retardation with microcephaly is a common presentation of bilallelic truncating mutations. Such growth failure is therefore sufficient to consider a diagnosis of LIG4 deficiency and early recognition of such cases is important as bone marrow failure, immunodeficiency, and sometimes malignancy are long term sequelae of this disorder.


Assuntos
DNA Ligases/deficiência , DNA Ligases/genética , Nanismo/genética , Retardo do Crescimento Fetal/genética , Leucopenia/genética , Trombocitopenia/genética , Anormalidades Múltiplas/genética , Imunidade Adaptativa , Adolescente , Linhagem Celular , Criança , Pré-Escolar , DNA Ligase Dependente de ATP , Exoma , Feminino , Retardo do Crescimento Fetal/etiologia , Variação Genética , Genótipo , Heterozigoto , Humanos , Lactente , Masculino , Microcefalia/genética , Neoplasias/genética , Síndrome de Quebra de Nijmegen/genética , Linhagem , Fenótipo , Polimorfismo de Nucleotídeo Único , Síndrome
16.
Hum Mol Genet ; 22(3): 444-51, 2013 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-23100325

RESUMO

Macrodactyly is a discrete congenital anomaly consisting of enlargement of all tissues localized to the terminal portions of a limb, typically within a 'nerve territory'. The classic terminology for this condition is 'lipofibromatous hamartoma of nerve' or Type I macrodactyly. The peripheral nerve, itself, is enlarged both in circumference and in length. It is not related to neurofibromatosis (NF1), nor is it associated with vascular malformations, such as in the recently reported CLOVES syndrome. The specific nerve pathophysiology in this form of macrodactyly has not been well described and a genetic etiology for this specific form of enlargement is unknown. To identify the genetic cause of macrodactyly, we used whole-exome sequencing to identify somatic mutations present in the affected nerve of a single patient. We confirmed a novel mutation in PIK3CA (R115P) present in the patient's affected nerve tissue but not in blood DNA. Sequencing PIK3CA exons identified gain-of-function mutations (E542K, H1047L or H1047R) in the affected tissue of five additional unrelated patients; mutations were absent in blood DNA available from three patients. Immunocytochemistry confirmed AKT activation in cultured cells from the nerve of a macrodactyly patient. Additionally, we found that the most abnormal structure within the involved nerve in a macrodactylous digit is the perineurium, with additional secondary effects on the axon number and size. Thus, isolated congenital macrodactyly is caused by somatic activation of the PI3K/AKT cell-signaling pathway and is genetically and biochemically related to other overgrowth syndromes.


Assuntos
Deformidades Congênitas dos Membros/genética , Mutação , Fosfatidilinositol 3-Quinases/genética , Células Cultivadas , Pré-Escolar , Classe I de Fosfatidilinositol 3-Quinases , Feminino , Dedos/anormalidades , Estudo de Associação Genômica Ampla/métodos , Humanos , Imuno-Histoquímica , Lactente , Microscopia Eletrônica , Tecido Nervoso/patologia , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Análise de Sequência de DNA , Transdução de Sinais , Síndrome
17.
Am J Hum Genet ; 90(5): 796-808, 2012 May 04.
Artigo em Inglês | MEDLINE | ID: mdl-22521419

RESUMO

Psoriasis is a common inflammatory disorder of the skin and other organs. We have determined that mutations in CARD14, encoding a nuclear factor of kappa light chain enhancer in B cells (NF-kB) activator within skin epidermis, account for PSORS2. Here, we describe fifteen additional rare missense variants in CARD14, their distribution in seven psoriasis cohorts (>6,000 cases and >4,000 controls), and their effects on NF-kB activation and the transcriptome of keratinocytes. There were more CARD14 rare variants in cases than in controls (burden test p value = 0.0015). Some variants were only seen in a single case, and these included putative pathogenic mutations (c.424G>A [p.Glu142Lys] and c.425A>G [p.Glu142Gly]) and the generalized-pustular-psoriasis mutation, c.413A>C (p.Glu138Ala); these three mutations lie within the coiled-coil domain of CARD14. The c.349G>A (p.Gly117Ser) familial-psoriasis mutation was present at a frequency of 0.0005 in cases of European ancestry. CARD14 variants led to a range of NF-kB activities; in particular, putative pathogenic variants led to levels >2.5× higher than did wild-type CARD14. Two variants (c.511C>A [p.His171Asn] and c.536G>A [p.Arg179His]) required stimulation with tumor necrosis factor alpha (TNF-α) to achieve significant increases in NF-kB levels. Transcriptome profiling of wild-type and variant CARD14 transfectants in keratinocytes differentiated probably pathogenic mutations from neutral variants such as polymorphisms. Over 20 CARD14 polymorphisms were also genotyped, and meta-analysis revealed an association between psoriasis and rs11652075 (c.2458C>T [p.Arg820Trp]; p value = 2.1 × 10(-6)). In the two largest psoriasis cohorts, evidence for association increased when rs11652075 was conditioned on HLA-Cw*0602 (PSORS1). These studies contribute to our understanding of the genetic basis of psoriasis and illustrate the challenges faced in identifying pathogenic variants in common disease.


Assuntos
Proteínas Adaptadoras de Sinalização CARD/genética , Guanilato Ciclase/genética , Proteínas de Membrana/genética , NF-kappa B/genética , NF-kappa B/metabolismo , Psoríase/genética , Proteínas Adaptadoras de Sinalização CARD/metabolismo , Estudos de Casos e Controles , Epiderme/metabolismo , Regulação da Expressão Gênica , Predisposição Genética para Doença , Guanilato Ciclase/metabolismo , Antígenos HLA-C/genética , Antígenos HLA-C/metabolismo , Humanos , Queratinócitos , Proteínas de Membrana/metabolismo , Mutação de Sentido Incorreto , Polimorfismo Genético , Pele/patologia , Transcriptoma , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/metabolismo , População Branca/genética
18.
Science ; 332(6026): 240-3, 2011 Apr 08.
Artigo em Inglês | MEDLINE | ID: mdl-21474761

RESUMO

The spliceosome, a ribonucleoprotein complex that includes proteins and small nuclear RNAs (snRNAs), catalyzes RNA splicing through intron excision and exon ligation to produce mature messenger RNAs, which, in turn serve as templates for protein translation. We identified four point mutations in the U4atac snRNA component of the minor spliceosome in patients with brain and bone malformations and unexplained postnatal death [microcephalic osteodysplastic primordial dwarfism type 1 (MOPD 1) or Taybi-Linder syndrome (TALS); Mendelian Inheritance in Man ID no. 210710]. Expression of a subgroup of genes, possibly linked to the disease phenotype, and minor intron splicing were affected in cell lines derived from TALS patients. Our findings demonstrate a crucial role of the minor spliceosome component U4atac snRNA in early human development and postnatal survival.


Assuntos
Mutação Puntual , Splicing de RNA , RNA Nuclear Pequeno/genética , Spliceossomos/genética , Pareamento de Bases , Linhagem Celular , Pré-Escolar , Cromossomos Humanos Par 2/genética , Nanismo/genética , Nanismo/metabolismo , Feminino , Retardo do Crescimento Fetal/genética , Retardo do Crescimento Fetal/metabolismo , Humanos , Lactente , Íntrons , Sequências Repetidas Invertidas , Masculino , Microcefalia/genética , Microcefalia/metabolismo , Proteínas Associadas aos Microtúbulos/genética , Conformação de Ácido Nucleico , Osteocondrodisplasias/genética , Osteocondrodisplasias/metabolismo , Linhagem , Sítios de Splice de RNA , RNA Nuclear Pequeno/química , RNA Nuclear Pequeno/metabolismo , Spliceossomos/metabolismo
19.
Nat Genet ; 41(2): 199-204, 2009 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-19169254

RESUMO

Psoriasis is a common immune-mediated disorder that affects the skin, nails and joints. To identify psoriasis susceptibility loci, we genotyped 438,670 SNPs in 1,409 psoriasis cases and 1,436 controls of European ancestry. We followed up 21 promising SNPs in 5,048 psoriasis cases and 5,041 controls. Our results provide strong support for the association of at least seven genetic loci and psoriasis (each with combined P < 5 x 10(-8)). Loci with confirmed association include HLA-C, three genes involved in IL-23 signaling (IL23A, IL23R, IL12B), two genes that act downstream of TNF-alpha and regulate NF-kappaB signaling (TNIP1, TNFAIP3) and two genes involved in the modulation of Th2 immune responses (IL4, IL13). Although the proteins encoded in these loci are known to interact biologically, we found no evidence for epistasis between associated SNPs. Our results expand the catalog of genetic loci implicated in psoriasis susceptibility and suggest priority targets for study in other auto-immune disorders.


Assuntos
Predisposição Genética para Doença , Interleucina-23/genética , NF-kappa B/genética , Psoríase/genética , Transdução de Sinais/genética , Adulto , Estudos de Casos e Controles , Proteínas de Ligação a DNA/genética , Feminino , Frequência do Gene , Estudo de Associação Genômica Ampla , Antígenos HLA-C/genética , Humanos , Subunidade p40 da Interleucina-12/genética , Interleucina-13/genética , Interleucina-4/genética , Peptídeos e Proteínas de Sinalização Intracelular/genética , Masculino , Pessoa de Meia-Idade , Proteínas Nucleares/genética , Polimorfismo de Nucleotídeo Único , Proteína 3 Induzida por Fator de Necrose Tumoral alfa , Adulto Jovem
20.
Am J Hum Genet ; 83(5): 616-22, 2008 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-18950742

RESUMO

Clubfoot is one of the most common severe musculoskeletal birth defects, with a worldwide incidence of 1 in 1000 live births. In the present study, we describe a five-generation family with asymmetric right-sided predominant idiopathic clubfoot segregating as an autosomal-dominant condition with incomplete penetrance. Other lower-limb malformations, including patellar hypoplasia, oblique talus, tibial hemimelia, developmental hip dysplasia, and preaxial polydactyly, were also present in some family members. Genome-wide linkage analysis with Affymetrix GeneChip Mapping 10K mapping data from 13 members of this family revealed a multipoint LOD(max) of 3.31 on chromosome 5q31. A single missense mutation (c.388G-->A) was identified in PITX1, a bicoid-related homeodomain transcription factor critical for hindlimb development, and segregated with disease in this family. The PITX1 E130K mutation is located in the highly conserved homeodomain and reduces the ability of PITX1 to transactivate a luciferase reporter. The PITX1 E130K mutation also suppresses wild-type PITX1 activity in a dose-dependent manner, suggesting dominant-negative effects on transcription. The propensity for right-sided involvement in tibial hemimelia and clubfoot suggests that PITX1, or pathways involving PITX1, may be involved in their etiology. Implication of a gene involved in early limb development in clubfoot pathogenesis also suggests additional pathways for future investigations of idiopathic clubfoot etiology in humans.


Assuntos
Anormalidades Congênitas/genética , Deformidades Congênitas das Extremidades Inferiores/genética , Mutação , Fatores de Transcrição Box Pareados/genética , Alelos , Sequência de Aminoácidos , Substituição de Aminoácidos , Estudos de Casos e Controles , Mapeamento Cromossômico , Cromossomos Humanos Par 5 , Sequência Conservada , Feminino , Frequência do Gene , Genes Dominantes , Ligação Genética , Marcadores Genéticos , Haplótipos , Heterozigoto , Humanos , Escore Lod , Deformidades Congênitas das Extremidades Inferiores/diagnóstico por imagem , Lisina/metabolismo , Masculino , Modelos Moleculares , Dados de Sequência Molecular , Mutação de Sentido Incorreto , Linhagem , Polimorfismo de Nucleotídeo Único , Radiografia , Fatores de Transcrição/genética
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