RESUMO
Tibial hemimelia is a rare, debilitating and often sporadic congenital deficiency. In syndromic cases, mutations of a Sonic hedgehog (SHH) enhancer have been identified. Here we describe an ~5 kb deletion within the SHH repressor GLI3 in two patients with bilateral tibial hemimelia. This deletion results in a truncated GLI3 protein that lacks a DNA-binding domain and cannot repress hedgehog signaling. These findings strengthen the concept that tibial hemimelia arises because of failure to restrict SHH activity to the posterior aspect of the limb bud.
Assuntos
Ectromelia/diagnóstico , Ectromelia/genética , Fatores de Transcrição Kruppel-Like , Mutação , Proteínas do Tecido Nervoso , Fenótipo , Tíbia/anormalidades , Animais , Linhagem Celular , Biologia Computacional/métodos , Variações do Número de Cópias de DNA , Éxons , Estudos de Associação Genética , Humanos , Mutação INDEL , Camundongos , Polimorfismo de Nucleotídeo Único , Esqueleto/diagnóstico por imagem , Esqueleto/patologia , Proteína Gli3 com Dedos de ZincoRESUMO
The tricellular junction (TCJ) forms at the convergence of bicellular junctions from three adjacent cells in polarized epithelia and is necessary for maintaining the transepithelial barrier. In the fruitfly Drosophila, the TCJ is generated at the meeting point of bicellular septate junctions. Gliotactin was the first identified component of the TCJ and is necessary for TCJ and septate junction development. Gliotactin is a member of the neuroligin family and associates with the PDZ protein discs large. Beyond this interaction, little is known about the mechanisms underlying Gliotactin localization and function at the TCJ. In this study, we show that Gliotactin is phosphorylated at conserved tyrosine residues, a process necessary for endocytosis and targeting to late endosomes and lysosomes for degradation. Regulation of Gliotactin levels through phosphorylation and endocytosis is necessary as overexpression results in displacement of Gliotactin away from the TCJ throughout the septate junction domain. Excessive Gliotactin in polarized epithelia leads to delamination, paired with subsequent migration, and apoptosis. The apoptosis and the resulting compensatory proliferation resulting from high levels of Gliotactin are mediated by the Drosophila JNK pathway. Therefore, Gliotactin levels within the cell membrane are regulated to ensure correct protein localization and cell survival.
Assuntos
Polaridade Celular , Drosophila/metabolismo , Endocitose , Células Epiteliais/metabolismo , Junções Intercelulares/metabolismo , Proteínas de Membrana/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Motivos de Aminoácidos , Animais , Proliferação de Células , Sobrevivência Celular , Drosophila/química , Drosophila/citologia , Drosophila/genética , Endossomos/genética , Endossomos/metabolismo , Células Epiteliais/química , Células Epiteliais/citologia , Junções Intercelulares/química , Proteínas de Membrana/química , Proteínas de Membrana/genética , Proteínas do Tecido Nervoso/química , Proteínas do Tecido Nervoso/genética , Fosforilação , Transporte ProteicoRESUMO
Chromatin remodeling by Polycomb group (PcG) and trithorax group (trxG) proteins regulates gene expression in all metazoans. Two major complexes, Polycomb repressive complexes 1 and 2 (PRC1 and PRC2), are thought to mediate PcG-dependent repression in flies and mammals. In Drosophila, PcG/trxG protein complexes are recruited by PcG/trxG response elements (PREs). However, it has been unclear how PcG/trxG are recruited in vertebrates. Here we have identified a vertebrate PRE, PRE-kr, that regulates expression of the mouse MafB/Kreisler gene. PRE-kr recruits PcG proteins in flies and mouse F9 cells and represses gene expression in a PcG/trxG-dependent manner. PRC1 and 2 bind to a minimal PRE-kr region, which can recruit stable PRC1 binding but only weak PRC2 binding when introduced ectopically, suggesting that PRC1 and 2 have different binding requirements. Thus, we provide evidence that similar to invertebrates, PREs act as entry sites for PcG/trxG chromatin remodeling in vertebrates.