RESUMEN
Dental enamel formation depends upon the transcellular transport of Ca(2+) by ameloblasts, but little is known about the molecular mechanism, or even if the same process is operative during the secretory and maturation stages of amelogenesis. Identifying mutations in genes involved in Ca(2+) homeostasis that cause inherited enamel defects can provide insights into the molecular participants and potential mechanisms of Ca(2+) handling by ameloblasts. Stromal Interaction Molecule 1 (STIM1) is an ER transmembrane protein that activates membrane-specific Ca(2+) influx in response to the depletion of ER Ca(2+) stores. Solute carrier family 24, member 4 (SLC24A4), is a Na(+)/K(+)/Ca(2+) transporter that exchanges intracellular Ca(2+) and K(+) for extracellular Na(+). We identified a proband with syndromic hypomaturation enamel defects caused by a homozygous C to T transition (g.232598C>T c.1276C>T p.Arg426Cys) in STIM1, and a proband with isolated hypomaturation enamel defects caused by a homozygous C to T transition (g.124552C>T; c.437C>T; p.Ala146Val) in SLC24A4. Immunohistochemistry of developing mouse molars and incisors showed positive STIM1 and SLC24A4 signal specifically in maturation-stage ameloblasts. We conclude that enamel maturation is dependent upon STIM1 and SLC24A4 function, and that there are important differences in the Ca(2+) transcellular transport systems used by secretory- and maturation-stage ameloblasts.
Asunto(s)
Amelogénesis/fisiología , Antiportadores/fisiología , Proteínas de la Membrana/fisiología , Proteínas de Neoplasias/fisiología , Alanina/genética , Ameloblastos/fisiología , Amelogénesis/genética , Animales , Antiportadores/genética , Arginina/genética , Señalización del Calcio/fisiología , Niño , Preescolar , Consanguinidad , Cisteína/genética , Citosina , Hipoplasia del Esmalte Dental/genética , Femenino , Variación Genética/genética , Homocigoto , Humanos , Proteínas de la Membrana/genética , Ratones , Mutación Missense/genética , Proteínas de Neoplasias/genética , Linaje , Molécula de Interacción Estromal 1 , Timina , Valina/genéticaRESUMEN
We identified two families with an autosomal-recessive disorder manifested by severe enamel hypoplasia, delayed and failed tooth eruption, misshapen teeth, intrapulpal calcifications, and localized gingival hyperplasia. Genetic analyses identified novel FAM20A mutations associated with the disease phenotype in both families. The proband of Family 1 had an altered splice junction in Intron 1 (g.502011G>C; c.405-1G>C) and a missense mutation in Exon 8 (g.65094G>A; c.1207G>A; p.D403N). The missense mutation is notable because D(403) is strictly conserved among FAM20A homologues, and the corresponding defect in FAM20C caused osteosclerotic bone dysplasia and a loss of kinase activity. The proband at age 12 yrs tested negative for nephrocalcinosis. The proband and her affected father in Family 2 were homozygous for a single nucleotide deletion that altered a splice junction in Intron 10 (g.66622del; c.1361+4del). Minigene analyses demonstrated that this alteration precluded normal splicing. Immunohistochemistry (IHC) of mouse maxillary first molars localized FAM20A in secretory-stage ameloblasts, in odontoblasts, and in the eruption pathway. IHC of kidneys localized FAM20A in the renal tubules. We conclude that FAM20A is likely a secretory pathway kinase and that loss-of-function mutations cause pathology where its phosphorylations are necessary for normal development or homeostasis.
Asunto(s)
Amelogénesis Imperfecta/genética , Proteínas del Esmalte Dental/genética , Mutación/genética , Nefrocalcinosis/genética , Adenosina , Animales , Niño , Preescolar , Citosina , Hipoplasia del Esmalte Dental/genética , Calcificaciones de la Pulpa Dental/genética , Exones/genética , Femenino , Estudios de Seguimiento , Genes Recesivos/genética , Vectores Genéticos/genética , Hiperplasia Gingival/genética , Guanina , Células HEK293 , Homocigoto , Humanos , Intrones/genética , Masculino , Ratones , Mutación Missense/genética , Fenotipo , Polimorfismo de Nucleótido Simple/genética , Eliminación de Secuencia/genética , Anomalías Dentarias/genética , Erupción Dental/genéticaRESUMEN
Non-syndromic amelogenesis imperfecta (AI) is a collection of isolated inherited enamel malformations that follow X-linked, autosomal-dominant, or autosomal-recessive patterns of inheritance. The AI phenotype is also found in syndromes. We hypothesized that whole-exome sequencing of AI probands showing simplex or recessive patterns of inheritance would identify causative mutations among the known candidate genes for AI. DNA samples obtained from 12 unrelated probands with AI were analyzed. Disease-causing mutations were identified in three of the probands: a novel single-nucleotide deletion in both KLK4 alleles (g.6930delG; c.245delG; p.Gly82Alafs*87) that shifted the reading frame, a novel missense transition mutation in both MMP20 alleles (g.15390A>G; c.611A>G; p.His204Arg) that substituted arginine for an invariant histidine known to coordinate a structural zinc ion, and a previously described nonsense transition mutation in a single allele of FAM83H (c.1379G>A; g.5663G>A; p.W460*). Erupted molars and cross-sections from unerupted parts of the mandibular incisors of Mmp20 null mice were characterized by scanning electron microscopy. Their enamel malformations closely correlated with the enamel defects displayed by the proband with the MMP20 mutation. We conclude that whole-exome sequencing is an effective means of identifying disease-causing mutations in kindreds with AI, and this technique should prove clinically useful for this purpose.