RESUMEN
Osteogenesis imperfecta (OI), or brittle bone disease, is a disorder characterized by bone fragility and increased fracture incidence. All forms of OI also feature short stature, implying an effect on endochondral ossification. Using the Aga2+/- mouse, which has a mutation in type I collagen, we show an affected growth plate primarily due to a shortened proliferative zone. We used single-cell RNA-Seq analysis of tibial and femoral growth plate tissues to understand transcriptional consequences on growth plate cell types. We show that perichondrial cells, which express abundant type I procollagen, and growth plate chondrocytes, which were found to express low amounts of type I procollagen, had ER stress and dysregulation of the same unfolded protein response pathway as previously demonstrated in osteoblasts. Aga2+/- proliferating chondrocytes showed increased FGF and MAPK signaling, findings consistent with accelerated differentiation. There was also increased Sox9 expression throughout the growth plate, which is expected to accelerate early chondrocyte differentiation but reduce late hypertrophic differentiation. These data reveal that mutant type I collagen expression in OI has an impact on the cartilage growth plate. These effects on endochondral ossification indicate that OI is a biologically complex phenotype going beyond its known impacts on bone to negatively affect linear growth.
Asunto(s)
Osteogénesis Imperfecta , Animales , Ratones , Cartílago/metabolismo , Colágeno Tipo I/genética , Colágeno Tipo I/metabolismo , Expresión Génica , Osteogénesis Imperfecta/metabolismoRESUMEN
Defects in the biosynthesis and/or function of primary cilia cause a spectrum of disorders collectively referred to as ciliopathies. A subset of these disorders is distinguished by profound abnormalities of the skeleton that include a long narrow chest with markedly short ribs, extremely short limbs, and polydactyly. These include the perinatal lethal short-rib polydactyly syndromes (SRPS) and the less severe asphyxiating thoracic dystrophy (ATD), Ellis-van Creveld (EVC) syndrome, and cranioectodermal dysplasia (CED) phenotypes. To identify new genes and define the spectrum of mutations in the skeletal ciliopathies, we analyzed 152 unrelated families with SRPS, ATD, and EVC. Causal variants were discovered in 14 genes in 120 families, including one newly associated gene and two genes previously associated with other ciliopathies. These three genes encode components of three different ciliary complexes; FUZ, which encodes a planar cell polarity complex molecule; TRAF3IP1, which encodes an anterograde ciliary transport protein; and LBR, which encodes a nuclear membrane protein with sterol reductase activity. The results established the molecular basis of SRPS type IV, in which mutations were identified in four different ciliary genes. The data provide systematic insight regarding the genotypes associated with a large cohort of these genetically heterogeneous phenotypes and identified new ciliary components required for normal skeletal development.
Asunto(s)
Ciliopatías/diagnóstico , Ciliopatías/genética , Estudios de Asociación Genética , Variación Genética , Fenotipo , Esqueleto/anomalías , Dineínas Citoplasmáticas/genética , Marcadores Genéticos , Genotipo , Humanos , Péptidos y Proteínas de Señalización Intercelular , Mutación , Proteínas/genética , Radiografía , Secuenciación del ExomaRESUMEN
Multiple Epiphyseal Dysplasia (MED) is a relatively mild skeletal dysplasia characterized by mild short stature, joint pain, and early-onset osteoarthropathy. Dominantly inherited mutations in COMP, MATN3, COL9A1, COL9A2, and COL9A3, and recessively inherited mutations in SLC26A2, account for the molecular basis of disease in about 80-85% of the cases. In two families with recurrent MED of an unknown molecular basis, we used exome sequencing and candidate gene analysis to identify homozygosity for recessively inherited missense mutations in CANT1, which encodes calcium-activated nucleotidase 1. The MED phenotype is thus allelic to the more severe Desbuquois dysplasia phenotype and the results identify CANT1 as a second locus for recessively inherited MED.
Asunto(s)
Genes Recesivos , Nucleotidasas/genética , Osteocondrodisplasias/genética , Adulto , Secuencia de Bases , Niño , Preescolar , Exoma/genética , Femenino , Humanos , Masculino , Mutación Missense/genética , Osteocondrodisplasias/diagnóstico por imagen , Osteocondrodisplasias/fisiopatología , Linaje , RadiografíaRESUMEN
Shohat-type spondyloepimetaphyseal dysplasia (SEMD) is a skeletal dysplasia that affects cartilage development. Similar skeletal disorders, such as spondyloepiphyseal dysplasias, are linked to mutations in type II collagen (COL2A1), but the causative gene in SEMD is not known. Here, we have performed whole-exome sequencing to identify a recurrent homozygous c.408+1G>A donor splice site loss-of-function mutation in DDRGK domain containing 1 (DDRGK1) in 4 families affected by SEMD. In zebrafish, ddrgk1 deficiency disrupted craniofacial cartilage development and led to decreased levels of the chondrogenic master transcription factor sox9 and its downstream target, col2a1. Overexpression of sox9 rescued the zebrafish chondrogenic and craniofacial phenotype generated by ddrgk1 knockdown, thus identifying DDRGK1 as a regulator of SOX9. Consistent with these results, Ddrgk1-/- mice displayed delayed limb bud chondrogenic condensation, decreased SOX9 protein expression and Col2a1 transcript levels, and increased apoptosis. Furthermore, we determined that DDRGK1 can directly bind to SOX9 to inhibit its ubiquitination and proteasomal degradation. Taken together, these data indicate that loss of DDRGK1 decreases SOX9 expression and causes a human skeletal dysplasia, identifying a mechanism that regulates chondrogenesis via modulation of SOX9 ubiquitination.
Asunto(s)
Proteínas Portadoras/fisiología , Osteocondrodisplasias/genética , Factor de Transcripción SOX9/metabolismo , Ubiquitinación , Proteínas Adaptadoras Transductoras de Señales , Animales , Cartílago/crecimiento & desarrollo , Células Cultivadas , Condrogénesis , Colágeno Tipo II/genética , Colágeno Tipo II/metabolismo , Expresión Génica , Estudios de Asociación Genética , Predisposición Genética a la Enfermedad , Homocigoto , Humanos , Ratones Noqueados , Mutación , Osteocondrodisplasias/metabolismo , Linaje , Isoformas de Proteínas/fisiología , Estabilidad Proteica , Sitios de Empalme de ARN , Pez CebraRESUMEN
[This corrects the article DOI: 10.1371/journal.pgen.1006307.].
RESUMEN
The acrofacial dysostoses (AFD) are a genetically heterogeneous group of inherited disorders with craniofacial and limb abnormalities. Rodriguez syndrome is a severe, usually perinatal lethal AFD, characterized by severe retrognathia, oligodactyly and lower limb abnormalities. Rodriguez syndrome has been proposed to be a severe form of Nager syndrome, a non-lethal AFD that results from mutations in SF3B4, a component of the U2 small nuclear ribonucleoprotein particle (U2 snRNP). Furthermore, a case with a phenotype intermediate between Rodriguez and Nager syndromes has been shown to have an SF3B4 mutation. We identified heterozygosity for SF3B4 mutations in Rodriguez syndrome, confirming that the phenotype is a dominant disorder that is allelic with Nager syndrome. The mutations led to reduced SF3B4 synthesis and defects in mRNA splicing, primarily exon skipping. The mutations also led to reduced expression in growth plate chondrocytes of target genes, including the DLX5, DLX6, SOX9, and SOX6 transcription factor genes, which are known to be important for skeletal development. These data provide mechanistic insight toward understanding how SF3B4 mutations lead to the skeletal abnormalities observed in the acrofacial dysostoses.
Asunto(s)
Condrocitos/metabolismo , Deformidades Congénitas de la Mano/genética , Disostosis Mandibulofacial/genética , Mutación , Factores de Empalme de ARN/genética , Empalme del ARN , Adulto , Células Cultivadas , Femenino , Deformidades Congénitas de la Mano/diagnóstico por imagen , Deformidades Congénitas de la Mano/patología , Proteínas de Homeodominio/genética , Proteínas de Homeodominio/metabolismo , Humanos , Recién Nacido , Masculino , Disostosis Mandibulofacial/diagnóstico por imagen , Disostosis Mandibulofacial/patología , Linaje , Fenotipo , Factores de Empalme de ARN/metabolismo , Factores de Transcripción SOXD/genética , Factores de Transcripción SOXD/metabolismo , Factores de Transcripción/genética , Factores de Transcripción/metabolismoRESUMEN
The short-rib polydactyly syndromes (SRPS) encompass a radiographically and genetically heterogeneous group of skeletal ciliopathies that are characterized by a long narrow chest, short extremities, and variable occurrence of polydactyly. Radiographic abnormalities include undermineralization of the calvarium, shortened and bowed appendicular bones, trident shaped acetabula and polydactyly. In a case of SRPS we identified compound heterozygosity for mutations in IFT52, which encodes a component of the anterograde intraflagellar transport complex. The IFT52 mutant cells synthesized a significantly reduced amount of IFT52 protein, leading to reduced synthesis of IFT74, IFT81, IFT88 and ARL13B, other key anterograde complex members. Ciliogenesis was also disrupted in the mutant cells, with a 60% reduction in the presence of cilia on mutant cells and loss of cilia length regulation for the cells with cilia. These data demonstrate that IFT52 is essential for anterograde complex integrity and for the biosynthesis and maintenance of cilia. The data identify a new locus for SRPS and show that IFT52 mutations result in a ciliopathy with primary effects on the skeleton.
Asunto(s)
Proteínas Portadoras/genética , Cilios/genética , Ciliopatías/genética , Síndrome de Costilla Pequeña y Polidactilia/genética , Cilios/metabolismo , Ciliopatías/fisiopatología , Proteínas del Citoesqueleto/genética , Flagelos/genética , Flagelos/patología , Humanos , Péptidos y Proteínas de Señalización Intracelular , Complejos Multiproteicos/genética , Proteínas Musculares/genética , Mutación/genética , Síndrome de Costilla Pequeña y Polidactilia/fisiopatología , Esqueleto/crecimiento & desarrollo , Esqueleto/metabolismo , Esqueleto/patología , Proteínas Supresoras de Tumor/genéticaAsunto(s)
Sitios Genéticos , Hidropesía Fetal/genética , Mutación , Osteocondrodisplasias/genética , Monoéster Fosfórico Hidrolasas/genética , Exoma , Expresión Génica , Edad Gestacional , Homocigoto , Humanos , Hidropesía Fetal/patología , Recién Nacido , Masculino , Osteocondrodisplasias/patología , Fosfatidilinositol-3,4,5-Trifosfato 5-Fosfatasas , MortinatoRESUMEN
Osteogenesis imperfecta (OI) is a genetic disorder that results in low bone mineral density and brittle bones. Most cases result from dominant mutations in the type I procollagen genes, but mutations in a growing number of genes have been identified that produce autosomal recessive forms of the disease. Among these include mutations in the genes SERPINH1 and FKBP10, which encode the type I procollagen chaperones HSP47 and FKBP65, respectively, and predominantly produce a moderately severe form of OI. Little is known about the biochemical consequences of the mutations and how they produce OI. We have identified a new OI mutation in SERPINH1 that results in destabilization and mislocalization of HSP47 and secondarily has similar effects on FKBP65. We found evidence that HSP47 and FKBP65 act cooperatively during posttranslational maturation of type I procollagen and that FKBP65 and HSP47 but fail to properly interact in mutant HSP47 cells. These results thus reveal a common cellular pathway in cases of OI caused by HSP47 and FKBP65 deficiency.
Asunto(s)
Colágeno Tipo I/biosíntesis , Proteínas del Choque Térmico HSP47/metabolismo , Osteogénesis Imperfecta/metabolismo , Procolágeno/biosíntesis , Proteínas de Unión a Tacrolimus/metabolismo , Adulto , Secuencia de Aminoácidos , Secuencia de Bases , Preescolar , Femenino , Proteínas del Choque Térmico HSP47/química , Proteínas del Choque Térmico HSP47/genética , Humanos , Masculino , Datos de Secuencia Molecular , Osteogénesis Imperfecta/genética , Linaje , Transporte de Proteínas , Alineación de Secuencia , Proteínas de Unión a Tacrolimus/química , Proteínas de Unión a Tacrolimus/genética , Adulto JovenRESUMEN
This report identifies human skeletal diseases associated with mutations in WNT1. In 10 family members with dominantly inherited, early-onset osteoporosis, we identified a heterozygous missense mutation in WNT1, c.652TâG (p.Cys218Gly). In a separate family with 2 siblings affected by recessive osteogenesis imperfecta, we identified a homozygous nonsense mutation, c.884CâA, p.Ser295*. In vitro, aberrant forms of the WNT1 protein showed impaired capacity to induce canonical WNT signaling, their target genes, and mineralization. In mice, Wnt1 was clearly expressed in bone marrow, especially in B-cell lineage and hematopoietic progenitors; lineage tracing identified the expression of the gene in a subset of osteocytes, suggesting the presence of altered cross-talk in WNT signaling between the hematopoietic and osteoblastic lineage cells in these diseases.
Asunto(s)
Mutación , Osteogénesis Imperfecta/genética , Osteoporosis/genética , Proteína Wnt1/genética , Adolescente , Adulto , Edad de Inicio , Anciano , Animales , Niño , Femenino , Genes Dominantes , Genes Recesivos , Humanos , Masculino , Ratones , Ratones Transgénicos , Persona de Mediana Edad , Linaje , Proteína Wnt1/metabolismo , Adulto JovenRESUMEN
Familial idiopathic basal ganglia calcification (IBGC) or Fahr's disease is a rare neurodegenerative disorder characterized by calcium deposits in the basal ganglia and other brain regions, which is associated with neuropsychiatric and motor symptoms. Familial IBGC is genetically heterogeneous and typically transmitted in an autosomal dominant fashion. We performed a mutational analysis of SLC20A2, the first gene found to cause IBGC, to assess its genetic contribution to familial IBGC. We recruited 218 subjects from 29 IBGC-affected families of varied ancestry and collected medical history, neurological exam, and head CT scans to characterize each patient's disease status. We screened our patient cohort for mutations in SLC20A2. Twelve novel (nonsense, deletions, missense, and splice site) potentially pathogenic variants, one synonymous variant, and one previously reported mutation were identified in 13 families. Variants predicted to be deleterious cosegregated with disease in five families. Three families showed nonsegregation with clinical disease of such variants, but retrospective review of clinical and neuroimaging data strongly suggested previous misclassification. Overall, mutations in SLC20A2 account for as many as 41% of our familial IBGC cases. Our screen in a large series expands the catalog of SLC20A2 mutations identified to date and demonstrates that mutations in SLC20A2 are a major cause of familial IBGC. Non-perfect segregation patterns of predicted deleterious variants highlight the challenges of phenotypic assessment in this condition with highly variable clinical presentation.
Asunto(s)
Enfermedades de los Ganglios Basales/genética , Calcinosis/genética , Mutación , Enfermedades Neurodegenerativas/genética , Proteínas Cotransportadoras de Sodio-Fosfato de Tipo III/genética , Adulto , Anciano , Secuencia de Aminoácidos , Estudios de Cohortes , Análisis Mutacional de ADN , Familia , Femenino , Humanos , Desequilibrio de Ligamiento , Masculino , Persona de Mediana Edad , Modelos Biológicos , Datos de Secuencia Molecular , Mutación/fisiología , Estudios RetrospectivosRESUMEN
Fibrochondrogenesis is a severe, recessively inherited skeletal dysplasia shown to result from mutations in the gene encoding the proα1(XI) chain of type XI collagen, COL11A1. The first of two cases reported here was the affected offspring of first cousins and sequence analysis excluded mutations in COL11A1. Consequently, whole-genome SNP genotyping was performed to identify blocks of homozygosity, identical-by-descent, wherein the disease locus would reside. COL11A1 was not within a region of homozygosity, further excluding it as the disease locus, but the gene encoding the proα2(XI) chain of type XI collagen, COL11A2, was located within a large region of homozygosity. Sequence analysis identified homozygosity for a splice donor mutation in intron 18. Exon trapping demonstrated that the mutation resulted in skipping of exon 18 and predicted deletion of 18 amino acids from the triple helical domain of the protein. In the second case, heterozygosity for a de novo 9 bp deletion in exon 40 of COL11A2 was identified, indicating that there are autosomal dominant forms of fibrochondrogenesis. These findings thus demonstrate that fibrochondrogenesis can result from either recessively or dominantly inherited mutations in COL11A2.
Asunto(s)
Colágeno Tipo XI/genética , Enanismo/genética , Enanismo/patología , Osteocondrodisplasias/genética , Osteocondrodisplasias/patología , Sitios de Empalme de ARN/genética , Enanismo/diagnóstico , Exones , Genes Dominantes , Genes Recesivos , Genotipo , Humanos , Recién Nacido , Intrones , Osteocondrodisplasias/diagnóstico , Polimorfismo de Nucleótido Simple , Eliminación de SecuenciaRESUMEN
Fibrochondrogenesis is a severe, autosomal-recessive, short-limbed skeletal dysplasia. In a single case of fibrochondrogenesis, whole-genome SNP genotyping identified unknown ancestral consanguinity by detecting three autozygous regions. Because of the predominantly skeletal nature of the phenotype, the 389 genes localized to the autozygous intervals were prioritized for mutation analysis by correlation of their expression with known cartilage-selective genes via the UCLA Gene Expression Tool, UGET. The gene encoding the α1 chain of type XI collagen (COL11A1) was the only cartilage-selective gene among the three candidate intervals. Sequence analysis of COL11A1 in two genetically independent fibrochondrogenesis cases demonstrated that each was a compound heterozygote for a loss-of-function mutation on one allele and a mutation predicting substitution for a conserved triple-helical glycine residue on the other. The parents who were carriers of missense mutations had myopia. Early-onset hearing loss was noted in both parents who carried a loss-of-function allele, suggesting COL11A1 as a locus for mild, dominantly inherited hearing loss. These findings identify COL11A1 as a locus for fibrochondrogenesis and indicate that there might be phenotypic manifestations among carriers.
Asunto(s)
Colágeno Tipo XI/genética , Mutación , Osteocondrodisplasias/genética , Cartílago/patología , Pérdida Auditiva/genética , Humanos , Osteocondrodisplasias/patologíaRESUMEN
Diaphanospondylodysostosis (DSD) is a rare, recessively inherited, perinatal lethal skeletal disorder. The low frequency and perinatal lethality of DSD makes assembling a large set of families for traditional linkage-based genetic approaches challenging. By searching for evidence of unknown ancestral consanguinity, we identified two autozygous intervals, comprising 34 Mbps, unique to a single case of DSD. Empirically testing for ancestral consanguinity was effective in localizing the causative variant, thereby reducing the genomic space within which the mutation resides. High-throughput sequence analysis of exons captured from these intervals demonstrated that the affected individual was homozygous for a null mutation in BMPER, which encodes the bone morphogenetic protein-binding endothelial cell precursor-derived regulator. Mutations in BMPER were subsequently found in three additional DSD cases, confirming that defects in BMPER produce DSD. Phenotypic similarities between DSD and Bmper null mice indicate that BMPER-mediated signaling plays an essential role in vertebral segmentation early in human development.
Asunto(s)
Proteína Morfogenética Ósea 2/genética , Consanguinidad , Disostosis/genética , Transducción de Señal/genética , Columna Vertebral/embriología , Espondilólisis/genética , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Genes Recesivos/genética , Homocigoto , Humanos , Ratones , Datos de Secuencia Molecular , Mutación/genética , Linaje , Polimorfismo de Nucleótido Simple/genética , Análisis de Secuencia de ADNRESUMEN
Phagocytosis, which is essential for the immune response to pathogens, is initiated by specific interactions between pathogens and cell surface receptors expressed by phagocytes. This study identifies triggering receptor expressed on myeloid cells 2 (TREM-2) and its signaling counterpart DAP12 as a molecular complex that promotes phagocytosis of bacteria. Expression of TREM-2-DAP12 enables nonphagocytic Chinese hamster ovary cells to internalize bacteria. This function depends on actin cytoskeleton dynamics and the activity of the small guanosine triphosphatases Rac and Cdc42. Internalization also requires src kinase activity and tyrosine phosphorylation. In bone marrow-derived macrophages, phagocytosis is decreased in the absence of DAP12 and can be restored by expression of TREM-2-DAP12. Depletion of TREM-2 inhibits both binding and uptake of bacteria. Finally, TREM-2-dependent phagocytosis is impaired in Syk-deficient macrophages. This study highlights a novel role for TREM-2-DAP12 in the immune response to bacterial pathogens.