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1.
Cancers (Basel) ; 13(4)2021 Feb 20.
Artigo em Inglês | MEDLINE | ID: mdl-33672471

RESUMO

The Tribbles (Trib) family of pseudokinase proteins regulate cell growth, proliferation, and differentiation during normal development and in response to environmental stress. Mutations in human Trib isoforms (Trib1, 2, and 3) have been associated with metabolic disease and linked to leukemia and the formation of solid tumors, including melanomas, hepatomas, and lung cancers. Drosophila Tribbles (Trbl) was the first identified member of this sub-family of pseudokinases and shares a conserved structure and similar functions to bind and direct the degradation of key mediators of cell growth and proliferation. Common Trib targets include Akt kinase (also known as protein kinase B), C/EBP (CAAT/enhancer binding protein) transcription factors, and Cdc25 phosphatases, leading to the notion that Trib family members stand athwart multiple pathways modulating their growth-promoting activities. Recent work using the Drosophila model has provided important insights into novel facets of conserved Tribbles functions in stem cell quiescence, tissue regeneration, metabolism connected to insulin signaling, and tumor formation linked to the Hippo signaling pathway. Here we highlight some of these recent studies and discuss their implications for understanding the complex roles Tribs play in cancers and disease pathologies.

2.
Fly (Austin) ; 12(1): 23-33, 2018 01 02.
Artigo em Inglês | MEDLINE | ID: mdl-29189094

RESUMO

Epithelial cells are defined by apical-basal and planar cell polarity (PCP) signaling, the latter of which establishes an orthogonal plane of polarity in the epithelial sheet. PCP signaling is required for normal cell migration, differentiation, stem cell generation and tissue repair, and defects in PCP have been associated with developmental abnormalities, neuropathologies and cancers. While the molecular mechanism of PCP is incompletely understood, the deepest insights have come from Drosophila, where PCP is manifest in hairs and bristles across the adult cuticle and organization of the ommatidia in the eye. Fly wing cells are marked by actin-rich trichome structures produced at the distal edge of each cell in the developing wing epithelium and in a mature wing the trichomes orient collectively in the distal direction. Genetic screens have identified key PCP signaling pathway components that disrupt trichome orientation, which has been measured manually in a tedious and error prone process. Here we describe a set of image processing and pattern-recognition macros that can quantify trichome arrangements in micrographs and mark these directly by color, arrow or colored arrow to indicate trichome location, length and orientation. Nearest neighbor calculations are made to exploit local differences in orientation to better and more reliably detect and highlight local defects in trichome polarity. We demonstrate the use of these tools on trichomes in adult wing preps and on actin-rich developing trichomes in pupal wing epithelia stained with phalloidin. FijiWingsPolarity is freely available and will be of interest to a broad community of fly geneticists studying the effect of gene function on PCP.


Assuntos
Polaridade Celular , Drosophila melanogaster/citologia , Software , Asas de Animais/citologia , Animais , Drosophila melanogaster/crescimento & desenvolvimento , Drosophila melanogaster/metabolismo , Pupa/citologia , Tricomas/metabolismo
3.
Dis Model Mech ; 10(12): 1453-1464, 2017 12 19.
Artigo em Inglês | MEDLINE | ID: mdl-29025897

RESUMO

Members of the Tribbles family of proteins are conserved pseudokinases with diverse roles in cell growth and proliferation. Both Drosophila Tribbles (Trbl) and vertebrate Trib3 proteins bind to the kinase Akt (Akt1) to block its phosphorylation activation and reduce downstream insulin-stimulated anabolism. A single nucleotide polymorphism (SNP) variant in human TRIB3, which results in a glutamine (Q) to arginine (R) missense mutation in a conserved motif at position 84, confers stronger Akt binding, resulting in reduced Akt phosphorylation, and is associated with a predisposition to Type 2 diabetes, cardiovascular disease, diabetic nephropathy, chronic kidney disease and leukemogenesis. Here, we used a Drosophila model to understand the importance of the conserved R residue in several Trbl functions. In the fly fat body, misexpression of a site-directed Q mutation at position R141 resulted in weakened binding to Drosophila Akt (dAkt), leading to increased levels of phospho-dAkt, increased cell and tissue size, and increases in the levels of stored glycogen and triglycerides. Consistent with the functional conservation of this arginine in modulating Akt activity, mouse Trib3 R84 misexpressed in the fly fat body blocked dAkt phosphorylation with a strength similar to wild-type Trbl. Limited mutational analysis shows that the R141 site dictates the strength of Akt binding but does not affect other Trbl-dependent developmental processes, suggesting a specificity that could serve as a drug target for metabolic diseases.


Assuntos
Proteínas de Ciclo Celular/genética , Drosophila melanogaster/metabolismo , Resistência à Insulina , Polimorfismo de Nucleotídeo Único/genética , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Proteínas Repressoras/genética , Sequência de Aminoácidos , Animais , Proteínas de Ciclo Celular/química , Proliferação de Células , Análise Mutacional de DNA , Modelos Animais de Doenças , Proteínas de Drosophila/química , Proteínas de Drosophila/genética , Drosophila melanogaster/citologia , Ativação Enzimática , Corpo Adiposo/metabolismo , Humanos , Insulina/metabolismo , Larva/crescimento & desenvolvimento , Camundongos , Mutação/genética , Ligação Proteica , Proteínas Serina-Treonina Quinases/química , Proteínas Serina-Treonina Quinases/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteínas Repressoras/química , Transdução de Sinais
4.
Biochem Soc Trans ; 43(5): 1057-62, 2015 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-26517923

RESUMO

Fruit flies have emerged as a powerful tool to investigate metabolism. Not only are gene structures and gene networks that control metabolism conserved through evolution, but the interactions among organs to store and process metabolites have strong similarities between flies and humans. Accordingly, the Drosophila system has the potential to address human disorders associated with metabolic dysfunction including obesity, type 2 diabetes and lipotoxicity.


Assuntos
Drosophila melanogaster/metabolismo , Resistência à Insulina/fisiologia , Transdução de Sinais/fisiologia , Vertebrados/metabolismo , Animais , Evolução Biológica , Diabetes Mellitus Tipo 2/genética , Diabetes Mellitus Tipo 2/metabolismo , Diabetes Mellitus Tipo 2/fisiopatologia , Drosophila melanogaster/genética , Humanos , Resistência à Insulina/genética , Modelos Biológicos , Obesidade/genética , Obesidade/metabolismo , Obesidade/fisiopatologia , Transdução de Sinais/genética , Vertebrados/genética
5.
PLoS One ; 9(10): e109530, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25329475

RESUMO

Drosophila Tribbles (Trbl) is the founding member of the Trib family of kinase-like docking proteins that modulate cell signaling during proliferation, migration and growth. In a wing misexpression screen for Trbl interacting proteins, we identified the Ser/Thr protein kinase Akt1. Given the central role of Akt1 in insulin signaling, we tested the function of Trbl in larval fat body, a tissue where rapid increases in size are exquisitely sensitive to insulin/insulin-like growth factor levels. Consistent with a role in antagonizing insulin-mediated growth, trbl RNAi knockdown in the fat body increased cell size, advanced the timing of pupation and increased levels of circulating triglyceride. Complementarily, overexpression of Trbl reduced fat body cell size, decreased overall larval size, delayed maturation and lowered levels of triglycerides, while circulating glucose levels increased. The conserved Trbl kinase domain is required for function in vivo and for interaction with Akt in a yeast two-hybrid assay. Consistent with direct regulation of Akt, overexpression of Trbl in the fat body decreased levels of activated Akt (pSer505-Akt) while misexpression of trbl RNAi increased phospho-Akt levels, and neither treatment affected total Akt levels. Trbl misexpression effectively suppressed Akt-mediated wing and muscle cell size increases and reduced phosphorylation of the Akt target FoxO (pSer256-FoxO). Taken together, these data show that Drosophila Trbl has a conserved role to bind Akt and block Akt-mediated insulin signaling, and implicate Trib proteins as novel sites of signaling pathway integration that link nutrient availability with cell growth and proliferation.


Assuntos
Proteínas de Ciclo Celular/genética , Proliferação de Células/genética , Proteínas de Drosophila/genética , Insulina/metabolismo , Proteínas Serina-Treonina Quinases/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Animais , Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/genética , Drosophila melanogaster/crescimento & desenvolvimento , Fosforilação , Ligação Proteica , Proteínas Serina-Treonina Quinases/metabolismo , Interferência de RNA , Transdução de Sinais/genética
6.
G3 (Bethesda) ; 3(8): 1443-9, 2013 Aug 07.
Artigo em Inglês | MEDLINE | ID: mdl-23797110

RESUMO

Development requires coordination between cell proliferation and cell growth to pattern the proper size of tissues, organs, and whole organisms. The Drosophila wing has landmark features, such as the location of veins patterned by cell groups and trichome structures produced by individual cells, that are useful to examine the genetic contributions to both tissue and cell size. Wing size and trichome density have been measured manually, which is tedious and error prone, and although image processing and pattern-recognition software can quantify features in micrographs, this approach has not been applied to insect wings. Here we present FijiWings, a set of macros designed to perform semiautomated morphophometric analysis of a wing photomicrograph. FijiWings uses plug-ins installed in the Fiji version of ImageJ to detect and count trichomes and measure wing area either to calculate trichome density of a defined region selected by the user or generate a heat map of overall trichome densities. For high-throughput screens we have developed a macro that directs a trainable segmentation plug-in to detect wing vein locations either to measure trichome density in specific intervein regions or produce a heat map of relative intervein areas. We use wing GAL4 drivers and UAS-regulated transgenes to confirm the ability of these tools to detect changes in overall tissue growth and individual cell size. FijiWings is freely available and will be of interest to a broad community of fly geneticists studying both the effect of gene function on wing patterning and the evolution of wing morphology.


Assuntos
Drosophila/metabolismo , Software , Asas de Animais/crescimento & desenvolvimento , Animais , Automação , Internet , Morfogênese , Interface Usuário-Computador
7.
Dev Biol ; 375(1): 33-44, 2013 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-23305818

RESUMO

Drosophila Tribbles (Trbl) encodes the founding member of the Trib family of kinase-like proteins that regulate cell migration, proliferation, growth and homeostasis. Trbl was identified in a misexpression screen in the ovary as an antagonist of border cell migration and acts in part by directing turnover of the C/EBP protein encoded by the gene slow border cells (slbo). The ability of mammalian Trib isoforms to promote C/EBP turnover during tissue differentiation indicates that this function is highly conserved. To better understand the role of Trbl in cell migration, we tested specific Trbl antisera, a trbl null allele and Trbl transgenes bearing site-directed mutations. Trbl is expressed at high levels in the nuclei of follicle cell epithelia and is downregulated in delaminating epithelia as expression of Slbo (C/EBP) is upregulated. This complementary pattern of expression during subsequent cell migration is achieved by negative feedback whereby slbo represses Trbl expression and trbl is necessary and sufficient to promote Slbo protein turnover. A series of point mutations that scan the conserved kinase domain of Trbl reveal that the conserved DLK catalytic loop is required for Trbl-Slbo binding and turnover, as well as for interactions between Trbl subunits, suggesting a mechanism of Trbl function.


Assuntos
Proteínas Estimuladoras de Ligação a CCAAT/metabolismo , Proteínas de Ciclo Celular/metabolismo , Movimento Celular , Proteínas de Drosophila/metabolismo , Drosophila/citologia , Proteínas Serina-Treonina Quinases/metabolismo , Animais , Proteínas Estimuladoras de Ligação a CCAAT/genética , Domínio Catalítico , Proteínas de Ciclo Celular/química , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/imunologia , Diferenciação Celular , Movimento Celular/genética , Proliferação de Células , Drosophila/metabolismo , Proteínas de Drosophila/química , Proteínas de Drosophila/genética , Proteínas de Drosophila/imunologia , Retroalimentação Fisiológica , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Masculino , Mutação , Oogênese/genética , Ovário/citologia , Ovário/metabolismo , Proteínas Serina-Treonina Quinases/química , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/imunologia , Interferência de RNA , RNA Interferente Pequeno , Transgenes
8.
Dev Dyn ; 241(8): 1239-48, 2012 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-22711497

RESUMO

The gene tribbles (trbl), identified 12 years ago in genetic screens for mutations that control both cell division and cell migration during embryonic Drosophila development, is the founding member of the Tribbles (Trib) family of kinase-like proteins that have diverse roles in cell signaling, tissue homeostasis, and cancer. Trib proteins share three motifs: (1) a divergent kinase region (Trib domain) with undetermined catalytic activity, (2) a COP1 site used to direct key target proteins to the proteosome for degradation, and (3) a MEK1 site that binds and modulates MAPKK kinase activity. The notion that Tribs act as scaffolding proteins to balance signaling levels in multiple pathways retains an attractive simplicity, but given recent data showing that divergent kinases act by means of novel catalytic mechanisms, the enzymatic activity of Tribs remains untested. Here, we focus on the role of Tribs during development. Developmental analysis of Drosophila trbl phenotypes reveals tissue-specific, sometimes contradictory roles. In mammals, multiple Trib isoforms exhibit overlapping and tissue-specific functions. Recent data indicate the mechanism of Trib activity is conserved and requires the Trib domain. Finally, we discuss the connections between Tribs in disease and cancer that have implications for their normal roles during organogenesis.


Assuntos
Proteínas de Ciclo Celular/metabolismo , Proteínas de Drosophila/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Animais , Proteínas de Ciclo Celular/química , Proteínas de Ciclo Celular/genética , Proteínas de Drosophila/química , Proteínas de Drosophila/genética , Humanos , Proteínas Serina-Treonina Quinases/química , Proteínas Serina-Treonina Quinases/genética , Transdução de Sinais
9.
Mech Dev ; 124(7-8): 559-69, 2007 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-17600691

RESUMO

The bunched (bun) gene encodes the Drosophila member of the TSC-22/GILZ family of leucine zipper transcriptional regulators. The bun locus encodes multiple BUN protein isoforms and has diverse roles during patterning of the eye, wing margin, dorsal notum and eggshell. Here we report the construction and activity of a dominant negative allele (BunDN) of the BUN-B isoform. In the ovary, BunDN expression in the follicle cells (FC) resulted in epithelial defects including aberrant accumulation of DE-cadherin and failure to rearrange into columnar FC cell shapes. BunDN expression in the posterior FC led to loss of epithelial integrity associated with extensive apoptosis. BunDN FC phenotypes collectively resemble loss-of-function bun mutant phenotypes. BunDN expression using tissue-specific imaginal disk drivers resulted in characteristic cuticular patterning defects that were enhanced by bun mutations and suppressed by co-expression of the BUN-B protein isoform. These data indicate that BunDN has dominant negative activity useful to identify bun functions and genetic interactions that occur during tissue patterning.


Assuntos
Proteínas de Drosophila/metabolismo , Drosophila/fisiologia , Animais , Padronização Corporal , Caderinas/metabolismo , Forma Celular , Drosophila/embriologia , Drosophila/metabolismo , Proteínas de Drosophila/genética , Epitélio/anormalidades , Epitélio/embriologia , Epitélio/fisiologia , Feminino , Mutação , Folículo Ovariano/citologia , Folículo Ovariano/metabolismo
10.
Dev Biol ; 243(1): 166-75, 2002 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-11846485

RESUMO

Signal transducer and activator of transcription (STAT) proteins are transcription factors that play a critical role in the response of a variety of eukaryotic cells to cytokine and growth factor signaling. In Drosophila, the STAT homolog encoded by the stat92E gene is required for the normal development of multiple tissues, including embryonic segmentation, imaginal discs, blood cells, male germ cells, and sex determination. We used multiple approaches to study the role of stat92E in oogenesis. Stat92E RNA expression is strongest in the differentiating follicle cells in the germarium, as determined by in situ hybridization. We generated an ethylmethane sulfonate-induced, temperature-sensitive allele, stat92E(F), in which the mutant protein contains a P506S substitution, located in the DNA binding domain. At the restrictive temperature, mutant females are sterile. Mutant ovaries have multiple defects, including fused egg chambers and an absence of interfollicular stalks cells and functional polar follicle cells. An analysis of mosaic clones, using an apparent null stat92E allele, indicates that Stat92E is required in the polar/stalk follicle cell lineage. We conclude that stat92E is necessary for the early differentiation of follicle cells and for proper germ line cell encapsulation during Drosophila oogenesis.


Assuntos
Proteínas de Ligação a DNA/fisiologia , Proteínas de Drosophila , Drosophila/fisiologia , Oogênese/fisiologia , Transativadores/fisiologia , Animais , Diferenciação Celular/fisiologia , Proteínas de Ligação a DNA/genética , Feminino , Mutação , Folículo Ovariano/citologia , Folículo Ovariano/fisiologia , Fatores de Transcrição STAT , Transativadores/genética
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