RESUMEN
Dpp/BMP acts as a morphogen to provide positional information in the Drosophila wing disc. Key cell-surface molecules to control Dpp morphogen gradient formation and signaling are heparan sulfate proteoglycans (HSPGs). In the wing disc, two HSPGs, the glypicans Division abnormally delayed (Dally) and Dally-like (Dlp) have been suggested to act redundantly to control these processes through direct interaction of their heparan sulfate (HS) chains with Dpp. Based on this assumption, a number of models on how glypicans control Dpp gradient formation and signaling have been proposed, including facilitating or hindering Dpp spreading, stabilizing Dpp on the cell surface, or recycling Dpp. However, how distinct HSPGs act remains largely unknown. Here, we generate genome-engineering platforms for the two glypicans and find that only Dally is critical for Dpp gradient formation and signaling through interaction of its core protein with Dpp. We also find that this interaction is not sufficient and that the HS chains of Dally are essential for these functions largely without interacting with Dpp. We provide evidence that the HS chains of Dally are not essential for spreading or recycling of Dpp but for stabilizing Dpp on the cell surface by antagonizing receptor-mediated Dpp internalization. These results provide new insights into how distinct HSPGs control morphogen gradient formation and signaling during development.
Asunto(s)
Proteínas de Drosophila , Drosophila , Proteoglicanos de Heparán Sulfato , Glicoproteínas de Membrana , Proteoglicanos , Animales , Membrana Celular , Drosophila/crecimiento & desarrollo , Glipicanos , Heparitina SulfatoRESUMEN
Retrograde bone morphogenetic protein (BMP) signaling at the Drosophila neuromuscular junction (NMJ) has served as a paradigm to study TGF-ß-dependent synaptic function and maturation. Yet, how retrograde BMP signaling transcriptionally regulates these functions remains unresolved. Here, we uncover a gene network, enriched for neurotransmission-related genes, that is controlled by retrograde BMP signaling in motor neurons through two Smad-binding cis-regulatory motifs, the BMP-activating (BMP-AE) and silencer (BMP-SE) elements. Unpredictably, both motifs mediate direct gene activation, with no involvement of the BMP derepression pathway regulators Schnurri and Brinker. Genome editing of candidate BMP-SE and BMP-AE within the locus of the active zone gene bruchpilot, and a novel Ly6 gene witty, demonstrated the role of these motifs in upregulating genes required for the maturation of pre- and post-synaptic NMJ compartments. Our findings uncover how Smad-dependent transcriptional mechanisms specific to motor neurons directly orchestrate a gene network required for synaptic maturation by retrograde BMP signaling.
Asunto(s)
Proteínas Morfogenéticas Óseas/metabolismo , Proteínas de Drosophila , Drosophila/metabolismo , Redes Reguladoras de Genes , Unión Neuromuscular/metabolismo , Animales , Animales Modificados Genéticamente , Drosophila/genética , Proteínas de Drosophila/metabolismo , Factor de Crecimiento Transformador beta/genética , Factor de Crecimiento Transformador beta/metabolismoRESUMEN
Cellular development and function rely on highly dynamic molecular interactions among proteins distributed in all cell compartments. Analysis of these interactions has been one of the main topics in cellular and developmental research, and has been mostly achieved by the manipulation of proteins of interest (POIs) at the genetic level. Although genetic strategies have significantly contributed to our current understanding, targeting specific interactions of POIs in a time- and space-controlled manner or analysing the role of POIs in dynamic cellular processes, such as cell migration or cell division, would benefit from more-direct approaches. The recent development of specific protein binders, which can be expressed and function intracellularly, along with advancement in synthetic biology, have contributed to the creation of a new toolbox for direct protein manipulations. Here, we have selected a number of short-tag epitopes for which protein binders from different scaffolds have been generated and showed that single copies of these tags allowed efficient POI binding and manipulation in living cells. Using Drosophila, we also find that single short tags can be used for POI manipulation in vivo.
Asunto(s)
Drosophila melanogaster/genética , Epítopos/genética , Péptidos/genética , Proteínas/genética , Animales , Línea Celular , Células Cultivadas , Péptidos/química , Unión Proteica/genética , Proteínas/química , Biología SintéticaRESUMEN
Precise control of stem cell (SC) proliferation ensures tissue homeostasis. In the Drosophila intestine, injury-induced regeneration involves initial activation of intestinal SC (ISC) proliferation and subsequent return to quiescence. These two phases of the regenerative response are controlled by differential availability of the BMP type I receptor Thickveins (Tkv), yet how its expression is dynamically regulated remains unclear. Here we show that during homeostasis, the E3 ubiquitin ligase Highwire and the ubiquitin-proteasome system maintain low Tkv protein expression. After ISC activation, Tkv is stabilized by proteasome inhibition and undergoes endocytosis due to the induction of the nucleoside diphosphate kinase Abnormal Wing Disc (AWD). Tkv internalization is required for the activation of the Smad protein Mad, and for the return to quiescence after a regenerative episode. Our data provide insight into the mechanisms ensuring tissue homeostasis by dynamic control of somatic stem cell activity.
Asunto(s)
Proteínas Morfogenéticas Óseas/metabolismo , Proteínas de Drosophila/metabolismo , Nucleósido-Difosfato Quinasa/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Receptores de Superficie Celular/metabolismo , Transducción de Señal/fisiología , Células Madre/metabolismo , Animales , Proteínas de Unión al ADN/metabolismo , Drosophila melanogaster , Femenino , Homeostasis/fisiología , Intestinos/citología , Modelos Animales , Proteínas del Tejido Nervioso/metabolismo , Regeneración , Factores de Transcripción/metabolismoRESUMEN
Intercellular signaling pathways activate transcription factors, which, along with tissue-specific co-factors, regulate expression of target genes. Responses to TGFß/BMP signals are mediated by Smad proteins, which form complexes and accumulate in the nucleus to directly bind and regulate enhancers of BMP targets upon signaling. In Drosophila, gene activation by BMP signaling often requires, in addition to direct input by Smads, the signal-dependent removal of the transcriptional repressor Brk. Previous studies on enhancers of BMP-activated genes have defined a BMP-responsive motif, the AE, which integrates activatory and repressive input by the Smad complex and Brk, respectively. Here, we address whether sequence variations within the core AE sequences might endow the motif with additional properties accounting for qualitative and quantitative differences in BMP responses, including tissue specificity of transcriptional activation and differential sensitivity to Smad and Brk inputs. By analyzing and cross-comparing three distinct BMP-responsive enhancers from the genes wit and Dad in two different epithelia, the wing imaginal disc and the follicular epithelium, we demonstrate that differences in the AEs contribute neither to the observed tissue-restriction of BMP responses nor to differences in the utilization of the Smad and Brk branches for transcriptional activation. Rather, our results suggest that the cis-environment of the BMP-response elements not only dictates tissue specificity but also differential sensitivity to the two BMP mediators.
Asunto(s)
Proteínas Morfogenéticas Óseas/metabolismo , Proteínas de Drosophila/fisiología , Drosophila melanogaster , Elementos de Respuesta/fisiología , Animales , Animales Modificados Genéticamente , Secuencia de Bases/fisiología , Sitios de Unión/genética , Proteínas Morfogenéticas Óseas/fisiología , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/embriología , Drosophila melanogaster/genética , Drosophila melanogaster/crecimiento & desarrollo , Embrión no Mamífero , Femenino , Regulación del Desarrollo de la Expresión Génica , Larva , Receptores de Superficie Celular/genética , Receptores de Superficie Celular/metabolismo , Transducción de Señal/genética , Activación Transcripcional/genéticaRESUMEN
Retrograde Bone Morphogenetic Protein (BMP) signaling in neurons is essential for the differentiation and synaptic function of many neuronal subtypes. BMP signaling regulates these processes via Smad transcription factor activity, yet the scope and nature of Smad-dependent gene regulation in neurons are mostly unknown. Here, we applied a computational approach to predict Smad-binding cis-regulatory BMP-Activating Elements (BMP-AEs) in Drosophila, followed by transgenic in vivo reporter analysis to test their neuronal subtype enhancer activity in the larval central nervous system (CNS). We identified 34 BMP-AE-containing genomic fragments that are responsive to BMP signaling in neurons, and showed that the embedded BMP-AEs are required for this activity. RNA-seq analysis identified BMP-responsive genes in the CNS and revealed that BMP-AEs selectively enrich near BMP-activated genes. These data suggest that functional BMP-AEs control nearby BMP-activated genes, which we validated experimentally. Finally, we demonstrated that the BMP-AE motif mediates a conserved Smad-responsive function in the Drosophila and vertebrate CNS. Our results provide evidence that BMP signaling controls neuronal function by directly coordinating the expression of a battery of genes through widespread deployment of a conserved Smad-responsive cis-regulatory motif.
Asunto(s)
Proteínas Morfogenéticas Óseas/fisiología , Proteínas de Drosophila/fisiología , Neuronas/metabolismo , Elementos de Respuesta , Transducción de Señal , Activación Transcripcional , Animales , Antígenos Ly/genética , Antígenos Ly/metabolismo , Embrión de Pollo , Proteínas de Unión al ADN/metabolismo , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/genética , Drosophila melanogaster/metabolismo , Evolución Molecular , Proteínas Ligadas a GPI/genética , Proteínas Ligadas a GPI/metabolismo , Proteínas Smad/metabolismo , Proteína Smad4/metabolismo , Factores de Transcripción/metabolismoRESUMEN
A common path to the formation of complex 3D structures starts with an epithelial sheet that is patterned by inductive cues that control the spatiotemporal activities of transcription factors. These activities are then interpreted by the cis-regulatory regions of the genes involved in cell differentiation and tissue morphogenesis. Although this general strategy has been documented in multiple developmental contexts, the range of experimental models in which each of the steps can be examined in detail and evaluated in its effect on the final structure remains very limited. Studies of the Drosophila eggshell patterning provide unique insights into the multiscale mechanisms that connect gene regulation and 3D epithelial morphogenesis. Here we review the current understanding of this system, emphasizing how the recent identification of cis-regulatory regions of genes within the eggshell patterning network enables mechanistic analysis of its spatiotemporal dynamics and evolutionary diversification. It appears that cis-regulatory changes can account for only some aspects of the morphological diversity of Drosophila eggshells, such as the prominent differences in the number of the respiratory dorsal appendages. Other changes, such as the appearance of the respiratory eggshell ridges, are caused by changes in the spatial distribution of inductive signals. Both types of mechanisms are at play in this rapidly evolving system, which provides an excellent model of developmental patterning and morphogenesis.
Asunto(s)
Drosophila/genética , Redes Reguladoras de Genes , Animales , Diferenciación Celular , Drosophila/crecimiento & desarrollo , Óvulo/citología , Óvulo/crecimiento & desarrollo , Óvulo/ultraestructuraRESUMEN
Alternative splicing (AS), the process which generates multiple RNA and protein isoforms from a single pre-mRNA, greatly contributes to transcript diversity and compensates for the fact that the gene number does not scale with organismal complexity. A number of genomic approaches have established that the extent of AS is much higher than previously expected, raising questions on its spatio-temporal regulation and function. In the present study, we address AS in the context of sex-specific neuronal development in the model Drosophila melanogaster. We report that at least 47 genes display sex-specific AS in the adult fly head. Unlike targets of the classical Sex lethal-dependent sex determination cascade, sex-specific isoforms of the vast majority of these genes are not present during larval development but start accumulating during metamorphosis or later, indicating the existence of novel mechanisms in the induction of sex-specific AS. We also established that sex-specific AS in the adult fly head is largely independent of the germline or the mating process. Finally, we investigated the role of sex-specific AS of the sulfotransferase Tango13 pre-mRNA and provide first evidence that differential expression of certain isoforms of this protein significantly affects courtship and mating behavior in male flies.
Asunto(s)
Envejecimiento/genética , Drosophila melanogaster/genética , Células Germinativas/metabolismo , Caracteres Sexuales , Conducta Sexual Animal , Empalme Alternativo , Animales , Proteínas de Drosophila/metabolismo , Femenino , Regulación del Desarrollo de la Expresión Génica , Cabeza , Masculino , Neuronas/metabolismo , Pupa/genética , ARN Mensajero/genética , ARN Mensajero/metabolismoRESUMEN
Tight regulation of signalling activity is crucial for proper tissue patterning and growth. Here we investigate the function of Pentagone (Pent), a secreted protein that acts in a regulatory feedback during establishment and maintenance of BMP/Dpp morphogen signalling during Drosophila wing development. We show that Pent internalises the Dpp co-receptors, the glypicans Dally and Dally-like protein (Dlp), and propose that this internalisation is important in the establishment of a long range Dpp gradient. Pent-induced endocytosis and degradation of glypicans requires dynamin- and Rab5, but not clathrin or active BMP signalling. Thus, Pent modifies the ability of cells to trap and transduce BMP by fine-tuning the levels of the BMP reception system at the plasma membrane. In addition, and in accordance with the role of glypicans in multiple signalling pathways, we establish a requirement of Pent for Wg signalling. Our data propose a novel mechanism by which morphogen signalling is regulated.
Asunto(s)
Proteínas de Drosophila/metabolismo , Drosophila/embriología , Proteínas de la Matriz Extracelular/metabolismo , Proteoglicanos/metabolismo , Transducción de Señal , Alas de Animales/embriología , Animales , Dinaminas/metabolismo , Proteínas de Unión al GTP rab5/metabolismoRESUMEN
The protein kinase LKB1 regulates cell metabolism and growth and is implicated in intestinal and lung cancer. Bone morphogenetic protein (BMP) signaling regulates cell differentiation during development and tissue homeostasis. We demonstrate that LKB1 physically interacts with BMP type I receptors and requires Smad7 to promote downregulation of the receptor. Accordingly, LKB1 suppresses BMP-induced osteoblast differentiation and affects BMP signaling in Drosophila wing longitudinal vein morphogenesis. LKB1 protein expression and Smad1 phosphorylation analysis in a cohort of non-small cell lung cancer patients demonstrated a negative correlation predominantly in a subset enriched in adenocarcinomas. Lung cancer patient data analysis indicated strong correlation between LKB1 loss-of-function mutations and high BMP2 expression, and these two events further correlated with expression of a gene subset functionally linked to apoptosis and migration. This new mechanism of BMP receptor regulation by LKB1 has ramifications in physiological organogenesis and disease.
Asunto(s)
Receptores de Proteínas Morfogenéticas Óseas de Tipo 1/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Transducción de Señal , Proteína smad7/metabolismo , Quinasas de la Proteína-Quinasa Activada por el AMP , Animales , Animales Modificados Genéticamente , Receptores de Proteínas Morfogenéticas Óseas de Tipo 1/genética , Línea Celular , Línea Celular Tumoral , Células Cultivadas , Drosophila/genética , Drosophila/crecimiento & desarrollo , Drosophila/metabolismo , Expresión Génica , Células HEK293 , Humanos , Immunoblotting , Inmunohistoquímica , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patología , Ratones Noqueados , Unión Proteica , Proteínas Serina-Treonina Quinasas/genética , Pupa/genética , Pupa/crecimiento & desarrollo , Pupa/metabolismo , Interferencia de ARN , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Proteína smad7/genética , Alas de Animales/crecimiento & desarrollo , Alas de Animales/metabolismoRESUMEN
Bone Morphogenetic Proteins (BMPs) signal by activating Smad transcription factors to control a number of decisions during animal development. In Drosophila, signaling by the BMP ligand Decapentaplegic (Dpp) involves the activity of brinker (brk) which, in most contexts, is repressed by Dpp. Brk encodes a transcription factor which represses BMP signaling output by antagonizing Smad-dependent target gene activation. Here, we study BMP-dependent gene regulation during Drosophila oogenesis by following the signal transmission from Dpp to its target broad (br), a gene with a crucial function in eggshell patterning. We identify regulatory sequences that account for expression of both brk and br, and connect these to the transcription factors of the pathway. We show that Dpp directly regulates brk transcription through Smad- and Schnurri (Shn)-dependent repression. Brk is epistatic to Dpp in br expression and activates br indirectly, through removal of a repressor, which is yet to be identified. Our work provides first cis-regulatory insights into transcriptional interpretation of BMP signaling in eggshell morphogenesis and defines a transcriptional cascade that connects Dpp to target gene regulation.
Asunto(s)
Proteínas Morfogenéticas Óseas/metabolismo , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/embriología , Drosophila melanogaster/metabolismo , Animales , Tipificación del Cuerpo , Femenino , Regulación del Desarrollo de la Expresión Génica , Oogénesis , Folículo Ovárico/metabolismo , Proteínas Represoras/metabolismoRESUMEN
In a broad variety of bilaterian species the trunk central nervous system (CNS) derives from three primary rows of neuroblasts. The fates of these neural progenitor cells are determined in part by three conserved transcription factors: vnd/nkx2.2, ind/gsh and msh/msx in Drosophila melanogaster/vertebrates, which are expressed in corresponding non-overlapping patterns along the dorsal-ventral axis. While this conserved suite of "neural identity" gene expression strongly suggests a common ancestral origin for the patterning systems, it is unclear whether the original regulatory mechanisms establishing these patterns have been similarly conserved during evolution. In Drosophila, genetic evidence suggests that Bone Morphogenetic Proteins (BMPs) act in a dosage-dependent fashion to repress expression of neural identity genes. BMPs also play a dose-dependent role in patterning the dorsal and lateral regions of the vertebrate CNS, however, the mechanism by which they achieve such patterning has not yet been clearly established. In this report, we examine the mechanisms by which BMPs act on cis-regulatory modules (CRMs) that control localized expression of the Drosophila msh and zebrafish (Danio rerio) msxB in the dorsal central nervous system (CNS). Our analysis suggests that BMPs act differently in these organisms to regulate similar patterns of gene expression in the neuroectoderm: repressing msh expression in Drosophila, while activating msxB expression in the zebrafish. These findings suggest that the mechanisms by which the BMP gradient patterns the dorsal neuroectoderm have reversed since the divergence of these two ancient lineages.
Asunto(s)
Proteínas Morfogenéticas Óseas/metabolismo , Proteínas de Drosophila/genética , Drosophila/genética , Drosophila/metabolismo , Regulación del Desarrollo de la Expresión Génica , Placa Neural/metabolismo , Vertebrados/genética , Vertebrados/metabolismo , Animales , Sitios de Unión , Secuencia Conservada , Genómica , Proteína Homeobox Nkx-2.2 , Placa Neural/embriología , Unión Proteica , Transducción de Señal , Elementos Silenciadores Transcripcionales , Proteínas de Pez CebraRESUMEN
Decapentaplegic (Dpp), the fly homolog of the secreted mammalian BMP2/4 signaling molecules, is involved in almost all aspects of fly development. Dpp has critical functions at all developmental stages, from patterning of the eggshell to the determination of adult intestinal stem cell identity. Here, we focus on recent findings regarding the transcriptional regulatory logic of the pathway, on a new feedback regulator, Pentagone, and on Dpp's roles in scaling and growth of the Drosophila wing.
Asunto(s)
Proteínas de Drosophila/genética , Drosophila melanogaster/genética , Regulación del Desarrollo de la Expresión Génica , Transducción de Señal/genética , Animales , Proteínas Morfogenéticas Óseas/genética , Proteínas Morfogenéticas Óseas/metabolismo , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/embriología , Drosophila melanogaster/crecimiento & desarrollo , Proteínas de la Matriz Extracelular/genética , Proteínas de la Matriz Extracelular/metabolismo , Modelos Genéticos , Alas de Animales/embriología , Alas de Animales/crecimiento & desarrollo , Alas de Animales/metabolismoRESUMEN
During Drosophila oogenesis, activation of Notch signaling in the follicular epithelium (FE) around stage 6 of oogenesis is essential for entry into the endocycle and a series of other changes such as cell differentiation and migration of subsets of the follicle cells. Notch induces the expression of zinc finger protein Hindsight and suppresses homeodomain protein Cut to regulate the mitotic/endocycle (ME) switch. Here we report that broad (br), encoding a small group of zinc-finger transcription factors resulting from alternative splicing, is a transcriptional target of Notch nuclear effector Suppressor of Hairless (Su(H)). The early pattern of Br in the FE, uniformly expressed except in the polar cells, is established by Notch signaling around stage 6, through the binding of Su(H) to the br early enhancer (brE) region. Mutation of the Su(H) binding site leads to a significant reduction of brE reporter expression in follicle cells undergoing the endocycle. Chromatin immunoprecipitation results further confirm Su(H) binding to the br early enhancer. Consistent with its expression in follicle cells during midoogenesis, loss of br function results in a delayed entry into the endocycle. Our findings suggest an important role of br in the timing of follicle cell development, and its transcriptional regulation by the Notch pathway.
Asunto(s)
Proteínas de Drosophila/genética , Drosophila melanogaster/embriología , Folículo Ovárico/embriología , Receptores Notch/genética , Proteínas Represoras/genética , Factores de Transcripción/genética , Animales , Sitios de Unión/genética , Proteínas de Drosophila/biosíntesis , Drosophila melanogaster/genética , Epitelio/embriología , Epitelio/metabolismo , Femenino , Proteínas de Homeodominio/biosíntesis , Proteínas Nucleares/biosíntesis , Oogénesis/genética , Oogénesis/fisiología , Folículo Ovárico/citología , Folículo Ovárico/fisiología , Interferencia de ARN , ARN Interferente Pequeño , Transducción de Señal/genética , Factores de Transcripción/biosíntesis , Transcripción GenéticaRESUMEN
Although it is widely appreciated that a typical developmental control gene is regulated by multiple enhancers, coordination of enhancer activities remains poorly understood. We propose a mechanism for such coordination in Drosophila oogenesis, when the expression of the transcription factor Broad (BR) evolves from a uniform to a two-domain pattern that prefigures the formation of two respiratory eggshell appendages. This change reflects sequential activities of two enhancers of the br gene, early and late, both of which are controlled by the epidermal growth factor receptor (EGFR) pathway. The late enhancer controls br in the appendage-producing cells, but the function of the early enhancer remained unclear. We found that the early enhancer is essential for the activity of the late enhancer and induction of eggshell appendages. This requirement can be explained by a mechanism whereby the BR protein produced by the early enhancer protects the late enhancer from EGFR-dependent repression. We illustrate this complex mechanism using a computational model that correctly predicts the wild-type dynamics of BR expression and its response to genetic perturbations.
Asunto(s)
Proteínas de Drosophila/metabolismo , Drosophila/fisiología , Elementos de Facilitación Genéticos/genética , Receptores ErbB/metabolismo , Modelos Biológicos , Oogénesis/fisiología , Transducción de Señal/fisiología , Factores de Transcripción/metabolismo , Animales , Biología Computacional , Receptores ErbB/genética , Retroalimentación Fisiológica , Transducción de Señal/genéticaRESUMEN
Traditionally, the analysis of gene regulatory regions suffered from the caveat that it was restricted to artificial contexts (e.g. reporter constructs of limited size). With the advent of the BAC recombineering technique, genomic constructs can now be generated to test regulatory elements in their endogenous environment. The expression of the transcriptional repressor brinker (brk) is negatively regulated by Dpp signaling. Repression is mediated by small sequence motifs, the silencer elements (SEs), that are present in multiple copies in the regulatory region of brk. In this work, we manipulated the SEs in the brk locus. We precisely quantified the effects of the individual SEs on the Brk gradient in the wing disc by employing a 1D data extraction method, followed by the quantification of the data with reference to an internal control. We found that mutating the SEs results in an expansion of the brk expression domain. However, even after mutating all predicted SEs, repression could still be observed in regions of maximal Dpp levels. Thus, our data point to the presence of additional, low affinity binding sites in the brk locus.
Asunto(s)
Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Drosophila/crecimiento & desarrollo , Drosophila/genética , Regulación del Desarrollo de la Expresión Génica , Proteínas Represoras/genética , Elementos Silenciadores Transcripcionales , Animales , Secuencia de Bases , Regulación hacia Abajo , Drosophila/metabolismo , Proteínas de Drosophila/análisis , Genes de Insecto , Genes Reporteros , Sitios Genéticos , Proteínas Fluorescentes Verdes/análisis , Proteínas Fluorescentes Verdes/genética , Mutación , Proteínas Represoras/análisis , Transducción de Señal , Alas de Animales/crecimiento & desarrollo , Alas de Animales/metabolismoRESUMEN
The Drosophila eggshell is an elaborate structure that is derived from a monolayer of follicular epithelium surrounding the developing oocyte within the female ovary. The bone morphogenetic protein (BMP) signaling pathway is essential for controlling the patterning and morphogenesis of the eggshell. During oogenesis, the roles of patterning and morphogenesis by the BMP type I receptor thickveins (tkv) have been studied extensively. However, signaling through this pathway requires both type I and II receptors, and the latter has yet to be established in oogenesis. We focus on wishful thinking (wit), the Drosophila homolog to the mammalian BMP type II receptor, BMPRII. We found that wit is expressed dynamically in the FCs of D. melanogaster in an evolutionary conserved pattern. The expression patterns are highly correlated with the dynamics of the BMP signaling, which is consistent with our finding that wit is a target of BMP signaling. Furthermore, we established that WIT is necessary for BMP signaling, and loss of WIT is associated with cell autonomous loss of BMP responses. Of importance, we demonstrated that perturbations in WIT led to changes in eggshell morphologies in domains that are patterned by BMP signaling. Previous studies have shown a role for WIT in BMP signaling during neurogenesis; however, our results reveal a role for WIT in epithelial cells' development.
Asunto(s)
Receptores de Proteínas Morfogenéticas Óseas de Tipo II/metabolismo , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/metabolismo , Cáscara de Huevo/fisiología , Oocitos/citología , Oocitos/metabolismo , Oogénesis , Receptores de Superficie Celular/metabolismo , Animales , Tipificación del Cuerpo/genética , Receptores de Proteínas Morfogenéticas Óseas de Tipo II/genética , Proteínas Morfogenéticas Óseas/genética , Proteínas Morfogenéticas Óseas/metabolismo , Diferenciación Celular , Proteínas de Drosophila/genética , Drosophila melanogaster/genética , Femenino , Regulación del Desarrollo de la Expresión Génica , Morfogénesis , Ovario , Proteínas Serina-Treonina Quinasas/metabolismo , Receptores de Superficie Celular/genética , Transducción de SeñalRESUMEN
Epidermal growth factor receptor (EGFR) controls a wide range of developmental events, from body axes specification in insects to cardiac development in humans. During Drosophila oogenesis, a gradient of EGFR activation patterns the follicular epithelium. Multiple transcriptional targets of EGFR in this tissue have been identified, but their regulatory elements are essentially unknown. We report the regulatory elements of broad (br) and pipe (pip), two important targets of EGFR signaling in Drosophila oogenesis. br is expressed in a complex pattern that prefigures the formation of respiratory eggshell appendages. We found that this pattern is generated by dynamic activities of two regulatory elements, which display different responses to Pointed, Capicua, and Mirror, transcription factors involved in the EGFR-mediated gene expression. One of these elements is active in a pattern similar to pip, a gene repressed by EGFR and essential for establishing the dorsoventral polarity of the embryo. We demonstrate that this similarity of expression depends on a common sequence motif that binds Mirror in vitro and is essential for transcriptional repression in vivo.
Asunto(s)
Drosophila/fisiología , Receptores ErbB/metabolismo , Transducción de Señal , Transcripción Genética , Animales , Secuencia de Bases , ADN , Regulación de la Expresión Génica , OogénesisRESUMEN
The wing of the fruit fly, Drosophila melanogaster, with its simple, two-dimensional structure, is a model organ well suited for a systems biology approach. The wing arises from an epithelial sac referred to as the wing imaginal disc, which undergoes a phase of massive growth and concomitant patterning during larval stages. The Decapentaplegic (Dpp) morphogen plays a central role in wing formation with its ability to co-coordinately regulate patterning and growth. Here, we asked whether the Dpp signaling activity scales, i.e. expands proportionally, with the growing wing imaginal disc. Using new methods for spatial and temporal quantification of Dpp activity and its scaling properties, we found that the Dpp response scales with the size of the growing tissue. Notably, scaling is not perfect at all positions in the field and the scaling of target gene domains is ensured specifically where they define vein positions. We also found that the target gene domains are not defined at constant concentration thresholds of the downstream Dpp activity gradients P-Mad and Brinker. Most interestingly, Pentagone, an important secreted feedback regulator of the pathway, plays a central role in scaling and acts as an expander of the Dpp gradient during disc growth.
Asunto(s)
Proteínas de Drosophila/metabolismo , Drosophila melanogaster/crecimiento & desarrollo , Proteínas de la Matriz Extracelular/metabolismo , Alas de Animales/crecimiento & desarrollo , Animales , Proteínas de Unión al ADN/metabolismo , Drosophila melanogaster/metabolismo , Femenino , Proteínas Fluorescentes Verdes/metabolismo , Proteínas de Homeodominio/metabolismo , Larva/crecimiento & desarrollo , Larva/metabolismo , Masculino , Microscopía Fluorescente , Morfogénesis , Proteínas del Tejido Nervioso/metabolismo , Tamaño de los Órganos , Fosforilación , Proteínas Recombinantes de Fusión/metabolismo , Proteínas Represoras/metabolismo , Proteínas de Dominio T Box/metabolismo , Factores de Transcripción/metabolismo , Alas de Animales/irrigación sanguíneaRESUMEN
Maintaining a proportionate body plan requires the adjustment or scaling of organ pattern with organ size. Scaling is a general property of developmental systems, yet little is known about its underlying molecular mechanisms. Using theoretical modeling, we examine how the Dpp activation gradient in the Drosophila wing imaginal disc scales with disc size. We predict that scaling is achieved through an expansion-repression mechanism [1] whose mediator is the widely diffusible protein Pentagone (Pent). Central to this mechanism is the repression of pent expression by Dpp signaling, which provides an effective size measurement, and the Pent-dependent expansion of the Dpp gradient, which adjusts the gradient with tissue size. We validate this mechanism experimentally by demonstrating that scaling requires Pent and further, that scaling is abolished when pent is ubiquitously expressed. The expansion-repression circuit can be readily implemented by a variety of molecular interactions, suggesting its general utilization for scaling morphogen gradients during development.