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1.
Insect Sci ; 27(1): 2-13, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-29943911

RESUMO

The silkworm Bombyx mori is an important lepidopteran model insect in which many kinds of natural mutants have been identified. However, molecular mechanisms of most of these mutants remain to be explored. Here we report the identification of a gene Bm-app is responsible for the silkworm minute wing (mw) mutation which exhibits exceedingly small wings during pupal and adult stages. Compared with the wild type silkworm, relative messenger RNA expression of Bm-app is significantly decreased in the u11 mutant strain which shows mw phenotype. A 10 bp insertion in the putative promoter region of the Bm-app gene in mw mutant strain was identified and the dual luciferase assay revealed that this insertion decreased Bm-app promoter activity. Furthermore, clustered regularly interspaced short palindromic repeats/RNA-guided Cas9 nucleases-mediated depletion of the Bm-app induced similar wing defects which appeared in the mw mutant, demonstrating that Bm-app controls wing development in B. mori. Bm-app encodes a palmitoyltransferase and is responsible for the palmitoylation of selected cytoplasmic proteins, indicating that it is required for cell mitosis and growth during wing development. We also discuss the possibility that Bm-app regulates wing development through the Hippo signaling pathway in B. mori.


Assuntos
Aciltransferases/genética , Bombyx/genética , Proteínas de Insetos/genética , Asas de Animais/crescimento & desenvolvimento , Aciltransferases/metabolismo , Animais , Bombyx/crescimento & desenvolvimento , Proteínas de Insetos/metabolismo , Pupa/genética , Pupa/crescimento & desenvolvimento
2.
Insect Sci ; 27(1): 14-21, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31246335

RESUMO

Manipulating an exogenous or endogenous gene of interest at a defined level is critical for a wide variety of experiments. The Gal4/UAS system has been widely used to direct gene expression for studying complex genetic and biological problems in Drosophila melanogaster and other model organisms. Driven by a given tissue-specific Gal4, expressing UAS-transgene or UAS-RNAi (RNA interference) could be used to up- or down-regulate target gene expression, respectively. However, the efficiency of the Gal4/UAS system is roughly predefined by properties of transposon vector constructs and the insertion site in the transgenic stock. Here, we describe a simple way to modulate optomotor blind (omb) expression levels in its endogenous expression region of the wing disc. We co-expressed UAS-omb and UAS-omb-RNAi together under the control of dpp-Gal4 driver which is expressed in the omb expression region of the wing pouch. The repression effect is more sensitive to temperature than that of overexpression. At low temperature, overexpression plays a dominant role but the efficiency is attenuated by UAS-omb-RNAi. In contrast, at high temperature RNAi predominates in gene expression regulation. By this strategy, we could manipulate omb expression levels at a moderate level. It allows us to manipulate omb expression levels in the same tissue between overexpression and repression at different stages by temperature control.


Assuntos
Proteínas de Drosophila/genética , Drosophila melanogaster/crescimento & desenvolvimento , Drosophila melanogaster/genética , Regulação da Expressão Gênica no Desenvolvimento , Proteínas do Tecido Nervoso/genética , Proteínas com Domínio T/genética , Asas de Animais/crescimento & desenvolvimento , Animais , Animais Geneticamente Modificados/genética , Animais Geneticamente Modificados/crescimento & desenvolvimento , Animais Geneticamente Modificados/metabolismo , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Transdução de Sinais , Proteínas com Domínio T/metabolismo
3.
PLoS Genet ; 15(8): e1007877, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31425502

RESUMO

Patterned expression of many developmental genes is specified by transcription factor gene expression, but is thought to be refined by chromatin-mediated repression. Regulatory DNA sequences called Polycomb Response Elements (PREs) are required to repress some developmental target genes, and are widespread in genomes, suggesting that they broadly affect developmental programs. While PREs in transgenes can nucleate trimethylation on lysine 27 of the histone H3 tail (H3K27me3), none have been demonstrated to be necessary at endogenous chromatin domains. This failure is thought to be due to the fact that most endogenous H3K27me3 domains contain many PREs, and individual PREs may be redundant. In contrast to these ideas, we show here that PREs near the wing selector gene vestigial have distinctive roles at their endogenous locus, even though both PREs are repressors in transgenes. First, a PRE near the promoter is required for vestigial activation and not for repression. Second, only the distal PRE contributes to H3K27me3, but even removal of both PREs does not eliminate H3K27me3 across the vestigial domain. Thus, endogenous chromatin domains appear to be intrinsically marked by H3K27me3, and PREs appear required to enhance this chromatin modification to high levels at inactive genes.


Assuntos
Cromatina/genética , Proteínas de Drosophila/genética , Regulação da Expressão Gênica no Desenvolvimento , Proteínas Nucleares/genética , Proteínas do Grupo Polycomb/metabolismo , Elementos de Resposta/genética , Animais , Animais Geneticamente Modificados , Metilação de DNA , Drosophila melanogaster/fisiologia , Feminino , Histonas/genética , Masculino , Mutagênese Sítio-Dirigida , Ativação Transcricional/genética , Asas de Animais/crescimento & desenvolvimento
4.
PLoS Genet ; 15(8): e1008133, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31425511

RESUMO

Coordinated intra- and inter-organ growth during animal development is essential to ensure a correctly proportioned individual. The Drosophila wing has been a valuable model system to reveal the existence of a stress response mechanism involved in the coordination of growth between adjacent cell populations and to identify a role of the fly orthologue of p53 (Dmp53) in this process. Here we identify the molecular mechanisms used by Dmp53 to regulate growth and proliferation in a non-autonomous manner. First, Dmp53-mediated transcriptional induction of Eiger, the fly orthologue of TNFα ligand, leads to the cell-autonomous activation of JNK. Second, two distinct signaling events downstream of the Eiger/JNK axis are induced in order to independently regulate tissue size and cell number in adjacent cell populations. Whereas expression of the hormone dILP8 acts systemically to reduce growth rates and tissue size of adjacent cell populations, the production of Reactive Oxygen Species-downstream of Eiger/JNK and as a consequence of apoptosis induction-acts in a non-cell-autonomous manner to reduce proliferation rates. Our results unravel how local and systemic signals act concertedly within a tissue to coordinate growth and proliferation, thereby generating well-proportioned organs and functionally integrated adults.


Assuntos
Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Peptídeos e Proteínas de Sinalização Intercelular/genética , Proteínas de Membrana/genética , Espécies Reativas de Oxigênio/metabolismo , Proteína Supressora de Tumor p53/metabolismo , Animais , Animais Geneticamente Modificados , Apoptose/genética , Proliferação de Células/genética , Drosophila melanogaster/fisiologia , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Larva/crescimento & desenvolvimento , Sistema de Sinalização das MAP Quinases/genética , Proteínas de Membrana/metabolismo , Modelos Animais , Tamanho do Órgão/genética , Asas de Animais/crescimento & desenvolvimento
5.
Insect Biochem Mol Biol ; 112: 103206, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31425850

RESUMO

Wings are an indispensable structure in many insects for their foraging, courtship, escape from predators, and migration. Cuticular proteins are major components of the insect cuticle and wings, but there is limited information on how cuticular proteins may play an essential role in wing morphogenesis. We identified a wing-specific cuticular protein, LmACP7, which belongs to the RR-2 subfamily of CPR chitin-binding proteins in the migratory locust. LmACP7 was initially produced in epidermal cells and subsequently migrated to the exocuticle at the pre-ecdysial stage in adult wings. Depletion of LmACP7 transcripts by RNA interference markedly reduced its protein amounts, which consequently led to abnormal wing morphogenesis. The deformed wings were curved, wrinkled, and failed to fully expand. We further demonstrated that the deformation was caused by both severe damage of the endocuticle and death of the epidermal cells in the wings. Based on these data, we propose that LmACP7 not only serves as an essential structural protein in the wing but is also required for the integrity of wing epithelial cells. LmACP7 contributes to production of the wing endocuticle and to the morphogenesis of functional wings in the migratory locust.


Assuntos
Proteínas de Insetos/genética , Locusta migratoria/genética , Asas de Animais/crescimento & desenvolvimento , Animais , Quitina/metabolismo , Células Epidérmicas/metabolismo , Proteínas de Insetos/metabolismo , Locusta migratoria/crescimento & desenvolvimento , Metamorfose Biológica/genética , Morfogênese/genética , Ninfa/genética , Ninfa/crescimento & desenvolvimento , Interferência de RNA , Asas de Animais/anormalidades
6.
Biochim Biophys Acta Mol Cell Res ; 1866(10): 1520-1532, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31326540

RESUMO

Activation and repression of Notch target genes is mediated by transcription factor CSL, known as Suppressor of Hairless (Su(H)) in Drosophila and CBF1 or RBPJ in human. CSL associates either with co-activator Notch or with co-repressors such as Drosophila Hairless. The nuclear translocation of transcription factor CSL relies on co-factor association, both in mammals and in Drosophila. The Drosophila CSL orthologue Su(H) requires Hairless for repressor complex formation. Based on its role in transcriptional silencing, H protein would be expected to be strictly nuclear. However, H protein is also cytosolic, which may relate to its role in the stabilization and nuclear translocation of Su(H) protein. Here, we investigate the function of the predicted nuclear localization signals (NLS 1-3) and single nuclear export signal (NES) of co-repressor Hairless using GFP-fusion proteins, reporter assays and in vivo analyses using Hairless wild type and shuttling-defective Hairless mutants. We identify NLS3 and NES to be critical for Hairless function. In fact, H⁎NLS3 mutant flies match H null mutants, whereas H⁎NLS3⁎NES double mutants display weaker phenotypes in agreement with a crucial role for NES in H export. As expected for a transcriptional repressor, Notch target genes are deregulated in H⁎NLS3 mutant cells, demonstrating nuclear requirement for its activity. Importantly, we reveal that Su(H) protein strictly follows Hairless protein localization. Together, we propose that shuttling between the nucleo-cytoplasmic compartments provides the possibility to fine tune the regulation of Notch target gene expression by balancing of Su(H) protein availability for Notch activation.


Assuntos
Citoplasma/metabolismo , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/genética , Drosophila melanogaster/metabolismo , Receptores Notch/metabolismo , Fatores de Transcrição/metabolismo , Animais , Proteínas de Drosophila/genética , Feminino , Sinais de Exportação Nuclear/genética , Sinais de Localização Nuclear/genética , Fenótipo , Receptores Notch/genética , Fatores de Transcrição/genética , Asas de Animais/crescimento & desenvolvimento
7.
Life Sci Alliance ; 2(4)2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31331981

RESUMO

One of the fundamental issues in biology is understanding how organ size is controlled. Tissue growth has to be carefully regulated to generate well-functioning organs, and defects in growth control can result in tumor formation. The Hippo signaling pathway is a universal growth regulator and has been implicated in cancer. In Drosophila, the Hippo pathway acts through the miRNA bantam to regulate cell proliferation and apoptosis. Even though the bantam targets regulating apoptosis have been determined, the target genes controlling proliferation have not been identified thus far. In this study, we identify the gene tribbles as a direct bantam target gene. Tribbles limits cell proliferation by suppressing G2/M transition. We show that tribbles regulation by bantam is central in controlling tissue growth and tumorigenesis. We expand our study to other cell cycle regulators and show that deregulated G2/M transition can collaborate with oncogene activation driving tumor formation.


Assuntos
Carcinogênese/genética , Proteínas de Ciclo Celular/genética , Proteínas de Drosophila/genética , Drosophila melanogaster/crescimento & desenvolvimento , MicroRNAs/genética , Proteínas Serina-Treonina Quinases/genética , Animais , Apoptose , Proliferação de Células , Regulação para Baixo , Drosophila melanogaster/genética , Regulação da Expressão Gênica no Desenvolvimento , Discos Imaginais/crescimento & desenvolvimento , Tamanho do Órgão , Transdução de Sinais , Asas de Animais/crescimento & desenvolvimento
8.
Int J Mol Sci ; 20(13)2019 Jul 04.
Artigo em Inglês | MEDLINE | ID: mdl-31277421

RESUMO

Lipin is evolutionarily conserved from yeast to mammals. Although its roles in lipid metabolism in adipocyte tissue, skeletal muscle, and the liver, and as a transcriptional co-activator are known, its functions during development are still under investigation. In this study, we analyzed the role of Drosophila lipin (dLipin) in development. Specifically, we showed that the tissue-selective knockdown of dLipin in the wing pouch led to an atrophied wing. Elevated DNA damage was observed in the wing imaginal disc of dLipin-knockdown flies. dLipin dysfunction induced accumulation of cells in S phase and significantly reduced the number of mitotic cells, indicating DNA damage-induced activation of the G2/M checkpoint. Reduced expression of cyclin B, which is critical for the G2 to M transition, was observed in the margin of the wing imaginal disc of dLipin-knockdown flies. The knockdown of dLipin led to increased apoptotic cell death in the wing imaginal disc. Thus, our results suggest that dLipin is involved in DNA replication during normal cell cycle progression in wing development of Drosophila melanogaster.


Assuntos
Proteínas de Drosophila/metabolismo , Drosophila melanogaster/crescimento & desenvolvimento , Drosophila melanogaster/metabolismo , Asas de Animais/crescimento & desenvolvimento , Asas de Animais/metabolismo , Animais , Apoptose/genética , Divisão Celular , Ciclina B/genética , Ciclina B/metabolismo , Dano ao DNA , Regulação para Baixo/genética , Drosophila melanogaster/genética , Feminino , Discos Imaginais/crescimento & desenvolvimento , Discos Imaginais/metabolismo , Masculino , Fase S
9.
PLoS Genet ; 15(6): e1008235, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-31242182

RESUMO

Polyphenism is a successful strategy adopted by organisms to adapt to environmental changes. Brown planthoppers (BPH, Nilaparvata lugens) develop two wing phenotypes, including long-winged (LW) and short-winged (SW) morphs. Though insulin receptor (InR) and juvenile hormone (JH) have been known to regulate wing polyphenism in BPH, the interaction between these regulators remains largely elusive. Here, we discovered that a conserved microRNA, miR-34, modulates a positive autoregulatory feedback loop of JH and insulin/IGF signaling (IIS) pathway to control wing polyphenism in BPH. Nlu-miR-34 is abundant in SW BPHs and suppresses NlInR1 by targeting at two binding sites in the 3'UTR of NlInR1. Overexpressing miR-34 in LW BPHs by injecting agomir-34 induces the development towards SW BPHs, whereas knocking down miR-34 in SW BPHs by injecting antagomir-34 induces more LW BPHs when another NlInR1 suppressor, NlInR2, is also suppressed simultaneously. A cis-response element of Broad Complex (Br-C) is found in the promoter region of Nlu-miR-34, suggesting that 20-hydroxyecdysone (20E) might be involved in wing polyphenism regulation. Topic application of 20E downregulates miR-34 expression but does not change wing morphs. On the other hand, JH application upregulates miR-34 expression and induces more SW BPHs. Moreover, knocking down genes in IIS pathway changes JH titers and miR-34 abundance. In all, we showed that miRNA mediates the cross talk between JH, 20E and IIS pathway by forming a positive feedback loop, uncovering a comprehensive regulation mechanism which integrates almost all known regulators controlling wing polyphenism in insects.


Assuntos
Hemípteros/genética , MicroRNAs/genética , Receptor de Insulina/genética , Asas de Animais/crescimento & desenvolvimento , Animais , Antagomirs/genética , Ecdisterona/genética , Regulação da Expressão Gênica/genética , Hemípteros/crescimento & desenvolvimento , Hormônios Juvenis/genética , Fenótipo , Regiões Promotoras Genéticas/genética , Asas de Animais/metabolismo
10.
Insect Biochem Mol Biol ; 110: 128-135, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31108167

RESUMO

The chitin modifying deacetylases (CDA) CDA1 and CDA2 have been reported to play partially redundant roles during insect cuticle formation and molting and tracheal morphogenesis in various insect species. In order to distinguish possible functional differences between these two enzymes, we analyzed their function during wing development in the fruit fly Drosophila melanogaster. In tissue-specific RNA interference experiments, we demonstrate that DmCDA1 (Serpentine, Serp) and DmCDA2 (Vermiform, Verm) have distinct functions during Drosophila adult wing cuticle differentiation. Chitosan staining revealed that Serp is the major enzyme responsible for chitin deacetylation during wing cuticle formation, while Verm does not seem to be needed for this process. Indeed, it is questionable whether Verm is a chitin deacetylase at all. Atomic force microscopy suggested that Serp and Verm have distinct roles in establishing the shape of nanoscale bumps at the wing surface. Moreover, our data indicate that Verm but not Serp is required for the laminar arrangement of chitin. Both enzymes participate in the establishment of the cuticular inward barrier against penetration of xenobiotics. Taken together, correct differentiation of the wing cuticle involves both Serp and Verm in parallel in largely non-overlapping functions.


Assuntos
Amidoidrolases/genética , Proteínas de Drosophila/genética , Drosophila melanogaster/crescimento & desenvolvimento , Drosophila melanogaster/genética , Asas de Animais/crescimento & desenvolvimento , Amidoidrolases/metabolismo , Animais , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/enzimologia , Larva/enzimologia , Larva/genética , Larva/crescimento & desenvolvimento , Pupa/enzimologia , Pupa/genética , Pupa/crescimento & desenvolvimento , Interferência de RNA , Asas de Animais/enzimologia
11.
Med Vet Entomol ; 33(4): 476-484, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31125148

RESUMO

Wing geometry helps to identify mosquito species, even cryptic ones. On the other hand, temperature has a well-known effect on insect metric properties. Can such effects blur the taxonomic signal embedded in the wing? Two strains of Aedes albopictus (laboratory and field strain) were examined under three different rearing temperatures (26, 30 and 33 °C) using landmark- and outline-based morphometric approaches. The wings of each experimental line were compared with Aedes aegypti. Both approaches indicated similar associations between wing size and temperature. For the laboratory strain, the wing size significantly decreased as the temperature increased. For the field strain, the largest wings were observed at the intermediate temperature. The two morphometric approaches describing shape showed different sensibilities to temperature. For both strains and sexes, the landmark-based approach disclosed significant wing shape changes with temperature changes. The outline-based approach showed lesser effects, detecting significant changes only in laboratory females and in field males. Despite the size and shape changes induced by temperature, the two strains of Ae. albopictus were always distinguished from Ae. aegypti. The present study confirms the lability of size. However, it also suggests that, despite environmentally-induced variation, the architecture of the wing still provides a strong taxonomic signal.


Assuntos
Aedes/anatomia & histologia , Temperatura Alta , Mosquitos Vetores/anatomia & histologia , Asas de Animais/anatomia & histologia , Aedes/crescimento & desenvolvimento , Animais , Feminino , Masculino , Mosquitos Vetores/crescimento & desenvolvimento , Tailândia , Asas de Animais/crescimento & desenvolvimento
12.
Molecules ; 24(8)2019 Apr 23.
Artigo em Inglês | MEDLINE | ID: mdl-31018591

RESUMO

Sonic hedgehog (Shh) signaling plays a tumor-promoting role in many epithelial cancers. Cancer cells produce soluble a Shh that signals to distant stromal cells that express the receptor Patched (Ptc). These receiving cells respond by producing other soluble factors that promote cancer cell growth, generating a positive feedback loop. To interfere with reinforced Shh signaling, we examined the potential of defined heparin and heparan sulfate (HS) polysaccharides to block Shh solubilization and Ptc receptor binding. We confirm in vitro and in vivo that proteolytic cleavage of the N-terminal Cardin-Weintraub (CW) amino acid motif is a prerequisite for Shh solubilization and function. Consistent with the established binding of soluble heparin or HS to the Shh CW target motif, both polysaccharides impaired proteolytic Shh processing and release from source cells. We also show that HS and heparin bind to, and block, another set of basic amino acids required for unimpaired Shh binding to Ptc receptors on receiving cells. Both modes of Shh activity downregulation depend more on HS size and overall charge than on specific HS sulfation modifications. We conclude that heparin oligosaccharide interference in the physiological roles of HS in Shh release and reception may be used to expand the field of investigation to pharmaceutical intervention of tumor-promoting Shh functions.


Assuntos
Proteínas de Drosophila/química , Proteínas de Drosophila/genética , Proteínas Hedgehog/química , Heparina/farmacologia , Heparitina Sulfato/farmacologia , Receptor Patched-1/genética , Receptores de Superfície Celular/genética , Sequência de Aminoácidos , Animais , Sítios de Ligação , Ligação Competitiva , Linhagem Celular Tumoral , Proteínas de Drosophila/antagonistas & inibidores , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/genética , Drosophila melanogaster/crescimento & desenvolvimento , Drosophila melanogaster/metabolismo , Embrião não Mamífero , Retroalimentação Fisiológica , Regulação da Expressão Gênica no Desenvolvimento , Células HeLa , Proteínas Hedgehog/antagonistas & inibidores , Proteínas Hedgehog/genética , Proteínas Hedgehog/metabolismo , Heparina/química , Heparitina Sulfato/química , Humanos , Modelos Moleculares , Receptor Patched-1/metabolismo , Ligação Proteica , Domínios e Motivos de Interação entre Proteínas , Estrutura Secundária de Proteína , Receptores de Superfície Celular/metabolismo , Transdução de Sinais , Solubilidade , Asas de Animais/crescimento & desenvolvimento , Asas de Animais/metabolismo
13.
J Therm Biol ; 81: 25-32, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30975420

RESUMO

In seasonal environments, natural selection should favor genotypes that acclimate to slow and predictable changes in temperature. Selective pressure for acclimation should be especially strong for animals that complete many generations per year, because seasonal warming or cooling causes offspring to experience different temperatures than their parents did. Here, we studied variation in acclimation capacity among three populations of Drosophila melanogaster. We used a reverse acclimation design to see whether developmental acclimation persisted throughout adulthood. Flies developed from fertilization to adulthood at either 16° or 26 °C. Then, flies either remained at the same temperature or moved to the other temperature for 7 days. We measured fecundity at seven temperatures ranging from 14° to 36°C. Genotypes from North Carolina and Vermont laid more eggs at 16 °C after spending the larval and adult stages at 16 °C, instead of 26 °C. In both populations, the benefit of acclimation to 16 °C during development was erased by acclimation to 26 °C during adulthood. In contrast to our prediction, genotypes from Indiana laid fewer eggs at 16 °C or 26 °C after developing at this temperature. Overall, these data provide only weak support for the models of optimal acclimation in seasonal environments.


Assuntos
Drosophila melanogaster/fisiologia , Estações do Ano , Termotolerância , Animais , Temperatura Corporal , Drosophila melanogaster/genética , Drosophila melanogaster/crescimento & desenvolvimento , Feminino , Fertilidade , Genótipo , Masculino , Modelos Biológicos , Temperatura Ambiente , Asas de Animais/crescimento & desenvolvimento
14.
Cell Prolif ; 52(3): e12593, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30847993

RESUMO

OBJECTIVES: The evolutionary conserved JNK pathway plays crucial role in cell death, yet factors that modulate this signalling have not been fully disclosed. In this study, we aim to identify additional factors that regulate JNK signalling in cell death, and characterize the underlying mechanisms. MATERIALS AND METHODS: Drosophila were raised on standard media, and cross was carried out at 25°C. The Gal4/UAS system was used to express proteins or RNAi in a specific temporal and spatial pattern. Gene expression was revealed by GFP fluorescence, X-gal staining or immunostaining of 3rd instar larval eye and wing discs. Cell death was visualized by acridine orange (AO) staining. Images of fly eyes and wings were taken by OLYMPUS microscopes. RESULTS: We found that licorne (lic) encoding the Drosophila MKK3 is an essential regulator of JNK-mediated cell death. Firstly, loss of lic suppressed ectopic Egr-triggered JNK activation and cell death in eye and wing development. Secondary, lic is necessary for loss-of-cell polarity-induced, physiological JNK-dependent cell death in wing development. Thirdly, Lic overexpression is sufficient to initiate JNK-mediated cell death in developing eyes and wings. Furthermore, ectopic Lic activates JNK signalling by promoting JNK phosphorylation. Finally, genetic epistatic analysis confirmed that Lic acts in parallel with Hep in the Egr-JNK pathway. CONCLUSIONS: This study not only identified Lic as a novel component of the JNK signalling, but also disclosed the crucial roles and mechanism of Lic in cell death.


Assuntos
Morte Celular , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/citologia , Drosophila melanogaster/metabolismo , Sistema de Sinalização das MAP Quinases , Proteínas Quinases/metabolismo , Animais , Animais Geneticamente Modificados , Morte Celular/genética , Proteínas de Drosophila/antagonistas & inibidores , Proteínas de Drosophila/genética , Drosophila melanogaster/genética , Epistasia Genética , Olho/crescimento & desenvolvimento , Olho/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Genes de Insetos , Sistema de Sinalização das MAP Quinases/genética , Proteínas de Membrana/genética , Quinases de Proteína Quinase Ativadas por Mitógeno/genética , Proteínas Quinases/genética , Interferência de RNA , Asas de Animais/crescimento & desenvolvimento , Asas de Animais/metabolismo
15.
Dev Biol ; 449(2): 143-150, 2019 05 15.
Artigo em Inglês | MEDLINE | ID: mdl-30844377

RESUMO

The decapentaplegic (dpp) gene plays a variety of roles in diverse cellular and molecular processes of the growth and development. In insects, dpp is mainly required for dorsal-ventral patterning and appendage formation. The brown planthopper (BPH) Nilaparvata lugens, a major pest of rice, possesses two distinct wing morphs described as long-winged (LW) and short-winged (SW) morphs. With our lab-maintained stable strains of LW and SW BPH, RNA interference (RNAi) was used to research the functions of N. lugens dpp (Nldpp) on wing development. Silencing of Nldpp in the SW strain led to the significant lengthening of the forewing, while Nldpp-knockdown in the LW strain resulted in distorted wings. Moreover, knockdown of Nldpp caused the complete absence of wing veins. During the development of wing-pads, the Nldpp abundance in the terga of the SW strain was significantly higher than that of the LW strain. Through controlling the direction of wing morph transformation, we found that the expression level of Nldpp increased in the NlInR1-knockdown BPH (tending to SW) and abundance of Nldpp declined after dsNlInR2 injection (tending to LW). Our results showed that Nldpp is mainly responsible for the formation and development of veins in BPH. Also, Nldpp can be regulated by NlInR1/2 and participate in the wing morph transformation.


Assuntos
Regulação da Expressão Gênica no Desenvolvimento , Hemípteros/genética , Proteínas de Insetos/genética , Asas de Animais/metabolismo , Sequência de Aminoácidos , Animais , Sequência de Bases , Padronização Corporal/genética , Hemípteros/anatomia & histologia , Hemípteros/crescimento & desenvolvimento , Proteínas de Insetos/metabolismo , Fenótipo , Interferência de RNA , Análise de Sequência de DNA/métodos , Asas de Animais/anatomia & histologia , Asas de Animais/crescimento & desenvolvimento
16.
G3 (Bethesda) ; 9(4): 999-1008, 2019 04 09.
Artigo em Inglês | MEDLINE | ID: mdl-30733380

RESUMO

During morphogenesis, cells communicate with each other to shape tissues and organs. Several lines of recent evidence indicate that ion channels play a key role in cellular signaling and tissue morphogenesis. However, little is known about the scope of specific ion-channel types that impinge upon developmental pathways. The Drosophila melanogaster wing is an excellent model in which to address this problem as wing vein patterning is acutely sensitive to changes in developmental pathways. We conducted a screen of 180 ion channels expressed in the wing using loss-of-function mutant and RNAi lines. Here we identify 44 candidates that significantly impacted development of the Drosophila melanogaster wing. Calcium, sodium, potassium, chloride, and ligand-gated cation channels were all identified in our screen, suggesting that a wide variety of ion channel types are important for development. Ion channels belonging to the pickpocket family, the ionotropic receptor family, and the bestrophin family were highly represented among the candidates of our screen. Seven new ion channels with human orthologs that have been implicated in human channelopathies were also identified. Many of the human orthologs of the channels identified in our screen are targets of common general anesthetics, anti-seizure and anti-hypertension drugs, as well as alcohol and nicotine. Our results confirm the importance of ion channels in morphogenesis and identify a number of ion channels that will provide the basis for future studies to understand the role of ion channels in development.


Assuntos
Drosophila melanogaster/crescimento & desenvolvimento , Canais Iônicos/fisiologia , Asas de Animais/crescimento & desenvolvimento , Animais , Drosophila melanogaster/genética , Drosophila melanogaster/metabolismo , Canais Iônicos/genética , Canais Iônicos/metabolismo , Morfogênese/genética , Interferência de RNA , Asas de Animais/metabolismo
17.
Biophys J ; 116(4): 725-740, 2019 02 19.
Artigo em Inglês | MEDLINE | ID: mdl-30704858

RESUMO

The robust specification of organ development depends on coordinated cell-cell communication. This process requires signal integration among multiple pathways, relying on second messengers such as calcium ions. Calcium signaling encodes a significant portion of the cellular state by regulating transcription factors, enzymes, and cytoskeletal proteins. However, the relationships between the inputs specifying cell and organ development, calcium signaling dynamics, and final organ morphology are poorly understood. Here, we have designed a quantitative image-analysis pipeline for decoding organ-level calcium signaling. With this pipeline, we extracted spatiotemporal features of calcium signaling dynamics during the development of the Drosophila larval wing disc, a genetic model for organogenesis. We identified specific classes of wing phenotypes that resulted from calcium signaling pathway perturbations, including defects in gross morphology, vein differentiation, and overall size. We found four qualitative classes of calcium signaling activity. These classes can be ordered based on agonist stimulation strength Gαq-mediated signaling. In vivo calcium signaling dynamics depend on both receptor tyrosine kinase/phospholipase C γ and G protein-coupled receptor/phospholipase C ß activities. We found that spatially patterned calcium dynamics correlate with known differential growth rates between anterior and posterior compartments. Integrated calcium signaling activity decreases with increasing tissue size, and it responds to morphogenetic perturbations that impact organ growth. Together, these findings define how calcium signaling dynamics integrate upstream inputs to mediate multiple response outputs in developing epithelial organs.


Assuntos
Sinalização do Cálcio , Drosophila melanogaster/anatomia & histologia , Asas de Animais/citologia , Asas de Animais/crescimento & desenvolvimento , Animais , Drosophila melanogaster/crescimento & desenvolvimento , Tamanho do Órgão , Organogênese , Fenótipo
18.
J Insect Physiol ; 114: 35-44, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30776423

RESUMO

Wing dimorphism is a common phenomenon in a wide range of insect taxa. In most insects, the two morphs are macropterous and micropterous, in extreme cases of the latter, wing shedding can occur. Wing dimorphism contributes significantly to the ecological success of many insect species. However, the molecular basis of wing dimorphism is not fully understood, especially for wing-shed. Here, differentially expressed genes over eight developmental stages of the house cricket Acheta domesticus, which undergoes wing-shed dimorphism, were studied. The results revealed a wing-shed peak during adult development in which many DEGs were highly upregulated and it's influenced by cricket population density. A weighted correlation network analysis (WGCNA) grouped 21,922 DEGs among 141,456 unigenes into 18 modules of different expression patterns. The module in which the gene expression pattern correlated with the wing-shed phenotypic data was selected for further analyses with STEM and Cytoscape, and three candidate genes (AdomHSP40: Heat shock protein 40, AdomCFDP: Craniofacial development protein, AdomDIS3L: DIS3 Like 3'-5' Exoribonuclease) were identified by gene network analysis as the DEGs most relevant to wing-shed occurrence. The RNA interference of these genes together with an insulin receptor and Nylanderia fulva virus showed that the silencing of AdomHSP40 significantly decreased wing-shed occurrence, whereas the silencing of other candidate genes did not, suggesting that AdomHSP40 plays a crucial role in the wing-shed of Acheta domesticus. These findings provide insights into the molecular mechanisms underlying wing dimorphism in the house crickets, which differ from those found in other insects such as the planthopper.


Assuntos
Gryllidae/crescimento & desenvolvimento , Proteínas de Choque Térmico HSP40/metabolismo , Asas de Animais/crescimento & desenvolvimento , Animais , Feminino , Perfilação da Expressão Gênica , Gryllidae/metabolismo , Masculino , Densidade Demográfica
19.
Genetics ; 211(4): 1429-1447, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30792267

RESUMO

Due to the complexity of genotype-phenotype relationships, simultaneous analyses of genomic associations with multiple traits will be more powerful and informative than a series of univariate analyses. However, in most cases, studies of genotype-phenotype relationships have been analyzed only one trait at a time. Here, we report the results of a fully integrated multivariate genome-wide association analysis of the shape of the Drosophila melanogaster wing in the Drosophila Genetic Reference Panel. Genotypic effects on wing shape were highly correlated between two different laboratories. We found 2396 significant SNPs using a 5% false discovery rate cutoff in the multivariate analyses, but just four significant SNPs in univariate analyses of scores on the first 20 principal component axes. One quarter of these initially significant SNPs retain their effects in regularized models that take into account population structure and linkage disequilibrium. A key advantage of multivariate analysis is that the direction of the estimated phenotypic effect is much more informative than a univariate one. We exploit this fact to show that the effects of knockdowns of genes implicated in the initial screen were on average more similar than expected under a null model. A subset of SNP effects were replicable in an unrelated panel of inbred lines. Association studies that take a phenomic approach, considering many traits simultaneously, are an important complement to the power of genomics.


Assuntos
Proteínas de Drosophila/genética , Estudo de Associação Genômica Ampla/métodos , Polimorfismo de Nucleotídeo Único , Asas de Animais/crescimento & desenvolvimento , Animais , Drosophila melanogaster , Estudo de Associação Genômica Ampla/normas , Padrões de Referência , Asas de Animais/metabolismo
20.
Oncogene ; 38(24): 4739-4754, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-30792539

RESUMO

Organ size is regulated by intercellular signaling for cell growth and proliferation. The TOR pathway mediates a key signaling mechanism for controlling cell size and number in organ growth. Chaperonin containing TCP-1 (CCT) is a complex that assists protein folding and function, but its role in animal development is largely unknown. Here we show that the CCT complex is required for organ growth by interacting with the TOR pathway in Drosophila. Reduction of CCT4 results in growth defects by affecting both cell size and proliferation. Loss of CCT4 causes preferential cell death anterior to the morphogenetic furrow in the eye disc and within wing pouch in the wing disc. Depletion of any CCT subunit in the eye disc results in headless phenotype. Overgrowth by active TOR signaling is suppressed by CCT RNAi. The CCT complex physically interacts with TOR signaling components including TOR, Rheb, and S6K. Loss of CCT leads to decreased phosphorylation of S6K and S6 while increasing phosphorylation of Akt. Insulin/TOR signaling is also necessary and sufficient for promoting CCT complex transcription. Our data provide evidence that the CCT complex regulates organ growth by directly interacting with the TOR signaling pathway.


Assuntos
Estruturas Animais/crescimento & desenvolvimento , Chaperonina com TCP-1/fisiologia , Chaperoninas/fisiologia , Insulina/metabolismo , Serina-Treonina Quinases TOR/metabolismo , Estruturas Animais/metabolismo , Animais , Animais Geneticamente Modificados , Chaperoninas/genética , Drosophila/embriologia , Drosophila/genética , Drosophila/crescimento & desenvolvimento , Drosophila/metabolismo , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Complexos Multiproteicos/fisiologia , Organogênese/genética , Transdução de Sinais/genética , Asas de Animais/crescimento & desenvolvimento , Asas de Animais/metabolismo
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