RESUMO
EP300 is a member of the EP300/CBP family of lysine acetyltransferases (KATs) with multiple roles in development and physiology. Loss of EP300/CBP activity in humans causes a very rare congenital disorder called Rubinstein Taybi Syndrome (RSTS). The zebrafish genome has two co-orthologs of lysine acetyltransferase EP300 (KAT3B) in zebrafish viz. ep300a and ep300b. Chemical inhibition of Ep300 with C646, a competitive inhibitor and morpholino-based genetic knockdown of ep300a and ep300b cause defects in embryonic development reminiscent of the human RSTS syndrome. Remarkably, overexpression of Ep300a KAT domain results in near complete rescue of the jaw development defects, a characteristic feature of RSTS in human suggesting the dispensability of the protein-interaction and DNA-binding domains for at least some developmental roles of Ep300. We also perform a chemical screen and identify two inhibitors of deacetylases, CHIC35 and HDACi III, that can partially rescue the RSTS-like phenotypes. Thus, modeling rare human genetic disorders in zebrafish allows for functional understanding of the genes involved and can also yield small molecule candidates towards therapeutic goals.
Assuntos
Modelos Animais de Doenças , Proteína p300 Associada a E1A , Embrião não Mamífero/embriologia , Desenvolvimento Embrionário/genética , Técnicas de Silenciamento de Genes , Síndrome de Rubinstein-Taybi , Proteínas de Peixe-Zebra , Peixe-Zebra , Animais , Proteína p300 Associada a E1A/genética , Proteína p300 Associada a E1A/metabolismo , Humanos , Síndrome de Rubinstein-Taybi/embriologia , Síndrome de Rubinstein-Taybi/genética , Síndrome de Rubinstein-Taybi/patologia , Peixe-Zebra/embriologia , Peixe-Zebra/genética , Proteínas de Peixe-Zebra/genética , Proteínas de Peixe-Zebra/metabolismoRESUMO
Mutations in the coactivator CREB-binding protein (CBP) are a major cause of the human skeletal dysplasia Rubinstein-Taybi syndrome (RTS); however, the mechanism by which these mutations affect skeletal mineralization and patterning is unknown. Here, we report the identification of 3-phosphoinositide-dependent kinase 1 (PDK1) as a key regulator of CBP activity and demonstrate that its functions map to both osteoprogenitor cells and mature osteoblasts. In osteoblasts, PDK1 activated the CREB/CBP complex, which in turn controlled runt-related transcription factor 2 (RUNX2) activation and expression of bone morphogenetic protein 2 (BMP2). These pathways also operated in vivo, as evidenced by recapitulation of RTS spectrum phenotypes with osteoblast-specific Pdk1 deletion in mice (Pdk1osx mice) and by the genetic interactions observed in mice heterozygous for both osteoblast-specific Pdk1 deletion and either Runx2 or Creb deletion. Finally, treatment of Pdk1osx and Cbp+/- embryos with BMPs in utero partially reversed their skeletal anomalies at birth. These findings illustrate the in vivo function of the PDK1-AKT-CREB/CBP pathway in bone formation and provide proof of principle for in utero growth factor supplementation as a potential therapy for skeletal dysplasias.
Assuntos
Proteína Morfogenética Óssea 2/administração & dosagem , Proteína Morfogenética Óssea 7/administração & dosagem , Proteína de Ligação a CREB/genética , Terapias Fetais/métodos , Proteínas Serina-Treonina Quinases/genética , Síndrome de Rubinstein-Taybi/genética , Síndrome de Rubinstein-Taybi/terapia , Animais , Desenvolvimento Ósseo/genética , Desenvolvimento Ósseo/fisiologia , Proteína Morfogenética Óssea 2/genética , Proteína de Ligação a CREB/deficiência , Subunidade alfa 1 de Fator de Ligação ao Core/deficiência , Subunidade alfa 1 de Fator de Ligação ao Core/genética , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/deficiência , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/genética , Modelos Animais de Doenças , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Humanos , Insulina/metabolismo , Fator de Crescimento Insulin-Like I/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Mutantes , Mutação , Osteoblastos/efeitos dos fármacos , Osteoblastos/metabolismo , Osteoblastos/patologia , Gravidez , Proteínas Serina-Treonina Quinases/deficiência , Proteínas Serina-Treonina Quinases/metabolismo , Piruvato Desidrogenase Quinase de Transferência de Acetil , Proteínas Recombinantes/administração & dosagem , Síndrome de Rubinstein-Taybi/embriologia , Síndrome de Rubinstein-Taybi/metabolismo , Transdução de Sinais , ÚteroRESUMO
Nail changes may be marker lesions for complex systemic disorders and herald associated syndromes. Knowledge of the anatomy, embryology and biochemical properties of the nail apparatus is essential for understanding the pathogenesis of hereditary nail disorders. In the last few years significant progress has been made in the field of clinical and molecular pathology of human diseases. A considerable number of the genes responsible for genodermatoses have been identified. The homeobox master control genes, genes encoding for transcription factors, genes encoding for the maintenance of telomeres, or for structural molecules, such as the similarly evolutionary highly conserved a-helical rod domains of keratins, are involved in the embryogenesis and normal functioning of nails. Using nail changes in selected genodermatoses with a known genetic background, we try to elucidate the genesis of inherited nail disorders and review the resultant clinical manifestations.
