RESUMO
Epithelial organs, including the lung, are known to possess regenerative abilities through activation of endogenous stem cell populations, but the molecular pathways regulating stem cell expansion and regeneration are not well understood. Here we show that Gata6 regulates the temporal appearance and number of bronchioalveolar stem cells (BASCs) in the lung, its absence in Gata6-null lung epithelium leading to the precocious appearance of BASCs and concurrent loss in epithelial differentiation. This expansion of BASCs was the result of a pronounced increase in canonical Wnt signaling in lung epithelium upon loss of Gata6. Expression of the noncanonical Wnt receptor Fzd2 was downregulated in Gata6 mutants and increased Fzd2 or decreased beta-catenin expression rescued, in part, the lung epithelial defects in Gata6 mutants. During lung epithelial regeneration, canonical Wnt signaling was activated in the niche containing BASCs and forced activation of Wnt signaling led to a large increase in BASC numbers. Moreover, Gata6 was required for proper lung epithelial regeneration, and postnatal loss of Gata6 led to increased BASC expansion and decreased differentiation. Together, these data demonstrate that Gata6-regulated Wnt signaling controls the balance between progenitor expansion and epithelial differentiation required for both lung development and regeneration.
Assuntos
Fator de Transcrição GATA6/fisiologia , Regeneração/genética , Mucosa Respiratória/fisiologia , Células-Tronco/fisiologia , Proteínas Wnt/fisiologia , Animais , Proliferação de Células , Embrião de Mamíferos , Células Epiteliais/fisiologia , Receptores Frizzled/fisiologia , Fator de Transcrição GATA6/genética , Perfilação da Expressão Gênica , Pulmão/embriologia , Pulmão/metabolismo , Pulmão/fisiologia , Camundongos , Camundongos Transgênicos , Modelos Biológicos , Dados de Sequência Molecular , Análise de Sequência com Séries de Oligonucleotídeos , Receptores Acoplados a Proteínas G/fisiologia , Regeneração/fisiologia , Mucosa Respiratória/metabolismo , Fenômenos Fisiológicos Respiratórios , Transdução de Sinais/fisiologia , Células-Tronco/metabolismo , beta Catenina/antagonistas & inibidoresRESUMO
We have previously demonstrated that PM-Scl-75, a component of the human exosome complex involved in RNA maturation and mRNA decay, can specifically interact with RNAs containing an AU-rich instability element. Through the analysis of a series of deletion mutants, we have now shown that a 266 amino acid fragment representing the RNase PH domain is responsible for the sequence-specific binding to AU-rich elements. Furthermore, we found that the RNase PH domains from two other exosomal components, OIP2 and RRP41, as well as from Escherichia coli polynucleotide phosphorylase, are all capable of specifically interacting with RNAs containing an AU-rich element with similar affinities. Finally, we demonstrate that the interaction of the RNase PH domain of PM-Scl-75 is readily competed by poly(U), but only inefficiently using other homopolymeric RNAs. These data demonstrate that RNase PH domains in general have an affinity for U- and AU-rich sequences, and broaden the potential role in RNA biology of proteins containing these domains.
Assuntos
Exorribonucleases/química , Exorribonucleases/metabolismo , RNA/genética , RNA/metabolismo , Composição de Bases , Sequência de Bases , Sítios de Ligação/genética , Exorribonucleases/genética , Humanos , Técnicas In Vitro , Plasmídeos/genética , Poli U/metabolismo , Estrutura Terciária de Proteína , RNA/química , Estabilidade de RNA , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Deleção de SequênciaRESUMO
Guanosine triphosphate (GTP)-binding proteins are known to function as molecular switches that cycle between GTP-bound and guanosine diphosphate (GDP)-bound states. Switching is achieved by the fact that G-proteins in the GTP-bound conformation can interact with a certain set of effector molecules while they interact with a different set of partners in their GDP-bound conformation. The antiviral properties of the interferon-induced MxA protein are critically dependent on the ability of MxA to bind GTP. Using limited proteolysis we analyzed the conformations of the MxA protein under nucleotide-free, GDP-bound, and GTP-bound conditions. We find that whereas the conformations of nucleotide-free MxA and GDP-bound MxA are essentially similar, GTP-binding causes a dramatic change in the conformation of MxA.
