RESUMO
Development of an organism requires accurate coordination between the growth of a tissue and orientation of cells within the tissue. The large cadherin Fat has been shown to play a role in both of these processes. Fat is involved in the establishment of planar cell polarity and regulates growth through the Hippo pathway, a developmental cascade that controls proliferation and apoptosis. Both Fat and the Hippo pathway are known to regulate transcription of four-jointed, although the nature of this regulation is unknown. In this study, we test whether Fat affects four-jointed transcription via or independently of Hippo pathway. Our analysis of the four-jointed regulatory region reveals a 1.2â¯kb element that functions as an enhancer for graded expression of Four-jointed in the eye imaginal disc. Within this enhancer element, we identify a 20 bp fragment that is critical for regulation by Fat but not by Hippo. Our findings suggest that Fat and the Hippo pathway control four-jointed expression independently of each other and none of the transcription factors known to function downstream of the Hippo pathway are required to regulate four-jointed expression through the 1.2â¯kb element.
Assuntos
Moléculas de Adesão Celular , Proteínas de Drosophila , Elementos Facilitadores Genéticos , Regulação da Expressão Gênica no Desenvolvimento , Peptídeos e Proteínas de Sinalização Intracelular , Glicoproteínas de Membrana , Proteínas Serina-Treonina Quinases , Transdução de Sinais , Transcrição Gênica , Animais , Moléculas de Adesão Celular/genética , Moléculas de Adesão Celular/metabolismo , Proteínas de Drosophila/biossíntese , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Drosophila melanogaster , Genes Reporter , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Glicoproteínas de Membrana/biossíntese , Glicoproteínas de Membrana/genética , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismoRESUMO
The Drosophila tumor suppressor gene fat encodes a large cadherin that regulates growth and a form of tissue organization known as planar cell polarity (PCP). Fat regulates growth via the Hippo kinase pathway, which controls expression of genes promoting cell proliferation and inhibiting apoptosis (reviewed in). The Hippo pathway is highly conserved and is implicated in the regulation of mammalian growth and cancer development. Genetic studies suggest that Fat activity is regulated by binding to another large cadherin, Dachsous (Ds). The tumor suppressor discs overgrown (dco)/Casein Kinase I delta/epsilon also regulates Hippo activity and PCP. The biochemical nature of how Fat, Ds, and Dco interact to regulate these pathways is poorly understood. Here we demonstrate that Fat is cleaved to generate 450 kDa and 110 kDa fragments (Fat(450) and Fat(110)). Fat(110) contains the cytoplasmic and transmembrane domain. The cytoplasmic domain of Fat binds Dco and is phosphorylated by Dco at multiple sites. Importantly, we show Fat forms cis-dimers and that Fat phosphorylation is regulated by Dachsous and Dco in vivo. We propose that Ds regulates Dco-dependent phosphorylation of Fat and Fat-associated proteins to control Fat signaling in growth and PCP.
Assuntos
Caderinas/metabolismo , Caseína Quinase 1 épsilon/metabolismo , Moléculas de Adesão Celular/metabolismo , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/fisiologia , Sequência de Aminoácidos , Animais , Caderinas/genética , Caseína Quinase 1 épsilon/genética , Caseína Quinase Idelta/genética , Caseína Quinase Idelta/metabolismo , Moléculas de Adesão Celular/genética , Linhagem Celular , Polaridade Celular , Proteínas de Drosophila/genética , Drosophila melanogaster/genética , Células-Tronco Embrionárias/citologia , Células-Tronco Embrionárias/fisiologia , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Ligantes , Camundongos , Dados de Sequência Molecular , Fosforilação , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Processamento de Proteína Pós-Traducional , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , TransgenesRESUMO
High levels of interspecies conservation characterise all signal transduction cascades and demonstrate the significance of these pathways over evolutionary time. Here, we review advances in the field of JAK/STAT signalling, focusing on recent developments in Drosophila. In particular, recent results from genetic and genome-wide RNAi screens, as well as studies into the developmental roles played by this pathway, highlight striking levels of physical and functional conservation in processes such as cellular proliferation, immune responses and stem cell maintenance. These insights underscore the value of model organisms for improving our understanding of this human disease-relevant pathway.
Assuntos
Proteínas de Drosophila/metabolismo , Drosophila melanogaster/fisiologia , Proteínas Tirosina Quinases/metabolismo , Fatores de Transcrição STAT/metabolismo , Transdução de Sinais/fisiologia , Fatores de Transcrição/metabolismo , Animais , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Proteínas de Drosophila/genética , Drosophila melanogaster/anatomia & histologia , Drosophila melanogaster/embriologia , Drosophila melanogaster/crescimento & desenvolvimento , Humanos , Janus Quinases , Proteínas Inibidoras de STAT Ativados/genética , Proteínas Inibidoras de STAT Ativados/metabolismo , Proteínas Tirosina Quinases/genética , Fatores de Transcrição STAT/genética , Células-Tronco/citologia , Células-Tronco/fisiologia , Proteínas Supressoras da Sinalização de Citocina/genética , Proteínas Supressoras da Sinalização de Citocina/metabolismo , Fatores de Transcrição/genéticaRESUMO
A limited number of evolutionarily conserved signal transduction pathways are repeatedly reused during development to regulate a wide range of processes. Here we describe a new negative regulator of JAK/STAT signaling and identify a potential mechanism by which the pleiotropy of responses resulting from pathway activation is generated in vivo. As part of a genetic interaction screen, we have identified Ken & Barbie (Ken) , which is an ortholog of the mammalian proto-oncogene BCL6 , as a negative regulator of the JAK/STAT pathway. Ken genetically interacts with the pathway in vivo and recognizes a DNA consensus sequence overlapping that of STAT92E in vitro. Tissue culture-based assays demonstrate the existence of Ken-sensitive and Ken-insensitive STAT92E binding sites, while ectopically expressed Ken is sufficient to downregulate a subset of JAK/STAT pathway target genes in vivo. Finally, we show that endogenous Ken specifically represses JAK/STAT-dependent expression of ventral veins lacking (vvl) in the posterior spiracles. Ken therefore represents a novel regulator of JAK/STAT signaling whose dynamic spatial and temporal expression is capable of selectively modulating the transcriptional repertoire elicited by activated STAT92E in vivo.
