RESUMO
In developing organisms, spatially prescribed cell identities are thought to be determined by the expression levels of multiple genes. Quantitative tests of this idea, however, require a theoretical framework capable of exposing the rules and precision of cell specification over developmental time. We use the gap gene network in the early fly embryo as an example to show how expression levels of the four gap genes can be jointly decoded into an optimal specification of position with 1% accuracy. The decoder correctly predicts, with no free parameters, the dynamics of pair-rule expression patterns at different developmental time points and in various mutant backgrounds. Precise cellular identities are thus available at the earliest stages of development, contrasting the prevailing view of positional information being slowly refined across successive layers of the patterning network. Our results suggest that developmental enhancers closely approximate a mathematically optimal decoding strategy.
Assuntos
Proteínas Ativadoras de GTPase/genética , Regulação da Expressão Gênica no Desenvolvimento/genética , Redes Reguladoras de Genes/genética , Animais , Padronização Corporal/genética , Diferenciação Celular/genética , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/genética , Drosophila melanogaster/metabolismo , Embrião não Mamífero/metabolismo , Desenvolvimento Embrionário/genética , Proteínas Ativadoras de GTPase/metabolismo , Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Modelos Genéticos , Fatores de Transcrição/metabolismoRESUMO
A conserved feature of the midblastula transition (MBT) is a requirement for a functional DNA replication checkpoint to coordinate cell-cycle remodeling and zygotic genome activation (ZGA). We have investigated what triggers this checkpoint during Drosophila embryogenesis. We find that the magnitude of the checkpoint scales with the quantity of transcriptionally engaged DNA. Measuring RNA polymerase II (Pol II) binding at 20 min intervals over the course of ZGA reveals that the checkpoint coincides with widespread de novo recruitment of Pol II that precedes and does not require a functional checkpoint. This recruitment drives slowing or stalling of DNA replication at transcriptionally engaged loci. Reducing Pol II recruitment in zelda mutants both reduces replication stalling and bypasses the requirement for a functional checkpoint. This suggests a model where the checkpoint functions as a feedback mechanism to remodel the cell cycle in response to nascent ZGA.
Assuntos
Replicação do DNA , Drosophila melanogaster/embriologia , Drosophila melanogaster/genética , Zigoto/metabolismo , Animais , Blástula/citologia , Blástula/metabolismo , Ciclo Celular , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/citologia , Drosophila melanogaster/metabolismo , Embrião não Mamífero/citologia , Embrião não Mamífero/metabolismo , Feminino , Masculino , Proteínas Nucleares , Regiões Promotoras Genéticas , RNA Polimerase II/metabolismo , Proteína de Replicação A/metabolismo , Fatores de Transcrição/metabolismoRESUMO
During early Drosophila embryogenesis, a network of gene regulatory interactions orchestrates terminal patterning, playing a critical role in the subsequent formation of the gut. We utilized CRISPR gene editing at endogenous loci to create live reporters of transcription and light-sheet microscopy to monitor the individual components of the posterior gut patterning network across 90 min prior to gastrulation. We developed a computational approach for fusing imaging datasets of the individual components into a common multivariable trajectory. Data fusion revealed low intrinsic dimensionality of posterior patterning and cell fate specification in wild-type embryos. The simple structure that we uncovered allowed us to construct a model of interactions within the posterior patterning regulatory network and make testable predictions about its dynamics at the protein level. The presented data fusion strategy is a step toward establishing a unified framework that would explore how stochastic spatiotemporal signals give rise to highly reproducible morphogenetic outcomes.
Assuntos
Padronização Corporal , Proteínas de Drosophila , Drosophila melanogaster , Endoderma , Redes Reguladoras de Genes , Animais , Padronização Corporal/genética , Proteínas de Drosophila/genética , Drosophila melanogaster/genética , Drosophila melanogaster/crescimento & desenvolvimento , Endoderma/crescimento & desenvolvimento , Regulação da Expressão Gênica no DesenvolvimentoRESUMO
In the regulation of gene expression, information of relevance to the organism is represented by the concentrations of transcription factor molecules. To extract this information the cell must effectively "measure" these concentrations, but there are physical limits to the precision of these measurements. We use the gap gene network in the early fly embryo as an example of the tradeoff between the precision of concentration measurements and the transmission of relevant information. For thresholded measurements we find that lower thresholds are more important, and fine tuning is not required for near-optimal information transmission. We then consider general sensors, constrained only by a limit on their information capacity, and find that thresholded sensors can approach true information theoretic optima. The information theoretic approach allows us to identify the optimal sensor for the entire gap gene network and to argue that the physical limitations of sensing necessitate the observed multiplicity of enhancer elements, with sensitivities to combinations rather than single transcription factors.
Assuntos
Redes Reguladoras de Genes/genética , Animais , Dípteros/genética , Regulação da Expressão Gênica/genética , Modelos Biológicos , Fatores de Transcrição/genéticaRESUMO
Tissue morphogenesis relies on repeated use of dynamic behaviors at the levels of intracellular structures, individual cells, and cell groups. Rapidly accumulating live imaging datasets make it increasingly important to formalize and automate the task of mapping recurrent dynamic behaviors (motifs), as it is done in speech recognition and other data mining applications. Here, we present a "template-based search" approach for accurate mapping of sub- to multi-cellular morphogenetic motifs using a time series data mining framework. We formulated the task of motif mapping as a subsequence matching problem and solved it using dynamic time warping, while relying on high throughput graph-theoretic algorithms for efficient exploration of the search space. This formulation allows our algorithm to accurately identify the complete duration of each instance and automatically label different stages throughout its progress, such as cell cycle phases during cell division. To illustrate our approach, we mapped cell intercalations during germband extension in the early Drosophila embryo. Our framework enabled statistical analysis of intercalary cell behaviors in wild-type and mutant embryos, comparison of temporal dynamics in contracting and growing junctions in different genotypes, and the identification of a novel mode of iterative cell intercalation. Our formulation of tissue morphogenesis using time series opens new avenues for systematic decomposition of tissue morphogenesis.
Assuntos
Biologia Computacional/métodos , Processamento de Imagem Assistida por Computador/métodos , Morfogênese/fisiologia , Algoritmos , Animais , Divisão Celular/fisiologia , Mineração de Dados/métodos , Drosophila/citologia , Drosophila/embriologia , Embrião não Mamífero/citologia , Embrião não Mamífero/embriologia , Feminino , Masculino , Microscopia Confocal , Fatores de TempoRESUMO
Many models of morphogenesis are forced to assume specific mechanical properties of cells, because the actual mechanical properties of living tissues are largely unknown. Here, we measure the rheology of epithelial cells in the cellularizing Drosophila embryo by injecting magnetic particles and studying their response to external actuation. We establish that, on timescales relevant to epithelial morphogenesis, the cytoplasm is predominantly viscous, whereas the cellular cortex is elastic. The timescale of elastic stress relaxation has a lower bound of 4 min, which is comparable to the time required for internalization of the ventral furrow during gastrulation. The cytoplasm was measured to be â¼103-fold as viscous as water. We show that elasticity depends on the actin cytoskeleton and conclude by discussing how these results relate to existing mechanical models of morphogenesis.
Assuntos
Drosophila melanogaster/embriologia , Embrião não Mamífero/fisiologia , Células Epiteliais/fisiologia , Nanopartículas de Magnetita , Imãs , Animais , Citoplasma/efeitos dos fármacos , Citoplasma/fisiologia , Citoesqueleto/fisiologia , Elasticidade , Embrião não Mamífero/ultraestrutura , Gastrulação/fisiologia , Células Gigantes/fisiologia , Magnetismo , Microinjeções , Modelos Biológicos , Morfogênese , Reologia , Estresse Mecânico , ViscosidadeRESUMO
During tissue morphogenesis, simple epithelial sheets undergo folding to form complex structures. The prevailing model underlying epithelial folding involves cell shape changes driven by myosin-dependent apical constriction. Here we describe an alternative mechanism that requires differential positioning of adherens junctions controlled by modulation of epithelial apical-basal polarity. Using live embryo imaging, we show that before the initiation of dorsal transverse folds during Drosophila gastrulation, adherens junctions shift basally in the initiating cells, but maintain their original subapical positioning in the neighbouring cells. Junctional positioning in the dorsal epithelium depends on the polarity proteins Bazooka and Par-1. In particular, the basal shift that occurs in the initiating cells is associated with a progressive decrease in Par-1 levels. We show that uniform reduction of the activity of Bazooka or Par-1 results in uniform apical or lateral positioning of junctions and in each case dorsal fold initiation is abolished. In addition, an increase in the Bazooka/Par-1 ratio causes formation of ectopic dorsal folds. The basal shift of junctions not only alters the apical shape of the initiating cells, but also forces the lateral membrane of the adjacent cells to bend towards the initiating cells, thereby facilitating tissue deformation. Our data thus establish a direct link between modification of epithelial polarity and initiation of epithelial folding.
Assuntos
Junções Aderentes/fisiologia , Polaridade Celular , Drosophila melanogaster/citologia , Drosophila melanogaster/embriologia , Células Epiteliais/citologia , Epitélio/embriologia , Gastrulação/fisiologia , Junções Aderentes/ultraestrutura , Animais , Forma Celular , Coristoma , Proteínas de Drosophila/deficiência , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/genética , Drosophila melanogaster/metabolismo , Células Epiteliais/metabolismo , Células Epiteliais/ultraestrutura , Epitélio/metabolismo , Epitélio/ultraestrutura , Gástrula/citologia , Gástrula/embriologia , Gástrula/metabolismo , Gástrula/ultraestrutura , Quinase 3 da Glicogênio Sintase , Peptídeos e Proteínas de Sinalização Intracelular/deficiência , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Proteínas Serina-Treonina Quinases/metabolismoRESUMO
Understanding the cellular and mechanical processes that underlie the shape changes of individual cells and their collective behaviors in a tissue during dynamic and complex morphogenetic events is currently one of the major frontiers in developmental biology. The advent of high-speed time-lapse microscopy and its use in monitoring the cellular events in fluorescently labeled developing organisms demonstrate tremendous promise in establishing detailed descriptions of these events and could potentially provide a foundation for subsequent hypothesis-driven research strategies. However, obtaining quantitative measurements of dynamic shapes and behaviors of cells and tissues in a rapidly developing metazoan embryo using time-lapse 3D microscopy remains technically challenging, with the main hurdle being the shortage of robust imaging processing and analysis tools. We have developed EDGE4D, a software tool for segmenting and tracking membrane-labeled cells using multi-photon microscopy data. Our results demonstrate that EDGE4D enables quantification of the dynamics of cell shape changes, cell interfaces and neighbor relations at single-cell resolution during a complex epithelial folding event in the early Drosophila embryo. We expect this tool to be broadly useful for the analysis of epithelial cell geometries and movements in a wide variety of developmental contexts.
Assuntos
Padronização Corporal , Drosophila melanogaster/embriologia , Epitélio/embriologia , Gastrulação , Animais , Forma Celular , Rastreamento de Células , Drosophila melanogaster/citologia , Células Epiteliais/citologia , Células Epiteliais/metabolismo , Processamento de Imagem Assistida por Computador , SoftwareRESUMO
Cells in a developing embryo have no direct way of "measuring" their physical position. Through a variety of processes, however, the expression levels of multiple genes come to be correlated with position, and these expression levels thus form a code for "positional information." We show how to measure this information, in bits, using the gap genes in the Drosophila embryo as an example. Individual genes carry nearly two bits of information, twice as much as would be expected if the expression patterns consisted only of on/off domains separated by sharp boundaries. Taken together, four gap genes carry enough information to define a cell's location with an error bar of ~1 along the anterior/posterior axis of the embryo. This precision is nearly enough for each cell to have a unique identity, which is the maximum information the system can use, and is nearly constant along the length of the embryo. We argue that this constancy is a signature of optimality in the transmission of information from primary morphogen inputs to the output of the gap gene network.
Assuntos
Movimento Celular/fisiologia , Drosophila/embriologia , Desenvolvimento Embrionário/fisiologia , Regulação da Expressão Gênica no Desenvolvimento/genética , Redes Reguladoras de Genes/genética , Modelos Biológicos , Proteínas/metabolismo , AnimaisRESUMO
Apical constriction facilitates epithelial sheet bending and invagination during morphogenesis. Apical constriction is conventionally thought to be driven by the continuous purse-string-like contraction of a circumferential actin and non-muscle myosin-II (myosin) belt underlying adherens junctions. However, it is unclear whether other force-generating mechanisms can drive this process. Here we show, with the use of real-time imaging and quantitative image analysis of Drosophila gastrulation, that the apical constriction of ventral furrow cells is pulsed. Repeated constrictions, which are asynchronous between neighbouring cells, are interrupted by pauses in which the constricted state of the cell apex is maintained. In contrast to the purse-string model, constriction pulses are powered by actin-myosin network contractions that occur at the medial apical cortex and pull discrete adherens junction sites inwards. The transcription factors Twist and Snail differentially regulate pulsed constriction. Expression of snail initiates actin-myosin network contractions, whereas expression of twist stabilizes the constricted state of the cell apex. Our results suggest a new model for apical constriction in which a cortical actin-myosin cytoskeleton functions as a developmentally controlled subcellular ratchet to reduce apical area incrementally.
Assuntos
Actinas/metabolismo , Drosophila melanogaster/citologia , Drosophila melanogaster/embriologia , Gastrulação , Miosina Tipo II/metabolismo , Actinas/química , Junções Aderentes/química , Junções Aderentes/metabolismo , Animais , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/genética , Regulação da Expressão Gênica no Desenvolvimento , Miosina Tipo II/química , Periodicidade , Fatores de Transcrição da Família Snail , Fatores de Transcrição/metabolismo , Proteína 1 Relacionada a Twist/metabolismoRESUMO
Tissue morphogenesis is the process in which coordinated movements and shape changes of large numbers of cells form tissues, organs, and the internal body structure. Understanding morphogenetic movements requires precise measurements of whole-cell shape changes over time. Tissue folding and invagination are thought to be facilitated by apical constriction, but the mechanism by which changes near the apical cell surface affect changes along the entire apical-basal axis of the cell remains elusive. Here, we developed Embryo Development Geometry Explorer, an approach for quantifying rapid whole-cell shape changes over time, and we combined it with deep-tissue time-lapse imaging based on fast two-photon microscopy to study Drosophila ventral furrow formation. We found that both the cell lengthening along the apical-basal axis and the movement of the nucleus to the basal side proceeded stepwise and were correlated with apical constriction. Moreover, cell volume lost apically due to constriction largely balanced the volume gained basally by cell lengthening. The volume above the nucleus was conserved during its basal movement. Both apical volume loss and cell lengthening were absent in mutants showing deficits in the contractile cytoskeleton underlying apical constriction. We conclude that a single mechanical mechanism involving volume conservation and apical constriction-induced basal movement of cytoplasm accounts quantitatively for the cell shape changes and the nucleus movement in Drosophila ventral furrow formation. Our study provides a comprehensive quantitative analysis of the fast dynamics of whole-cell shape changes during tissue folding and points to a simplified model for Drosophila gastrulation.
Assuntos
Núcleo Celular/metabolismo , Forma Celular , Tamanho Celular , Drosophila melanogaster/citologia , Drosophila melanogaster/crescimento & desenvolvimento , Organogênese , Animais , Polaridade Celular , Citoplasma/metabolismo , Drosophila melanogaster/embriologia , Embrião não Mamífero/anatomia & histologia , Embrião não Mamífero/citologia , MovimentoRESUMO
Covalent modification cycles (systems in which the activity of a substrate is regulated by the action of two opposing enzymes) and ligand/receptor interactions are ubiquitous in signaling systems and their steady-state properties are well understood. However, the behavior of such systems far from steady state remains unclear. Here, we analyze the properties of covalent modification cycles and ligand/receptor interactions driven by the accumulation of the activating enzyme and the ligand, respectively. We show that for a large range of parameters both systems produce sharp switchlike response and yet allow for temporal integration of the signal, two desirable signaling properties. Ultrasensitivity is observed also in a region of parameters where the steady-state response is hyperbolic. The temporal integration properties are tunable by regulating the levels of the deactivating enzyme and receptor, as well as by adjusting the rate of accumulation of the activating enzyme and ligand. We propose that this tunability is used to generate precise responses in signaling systems.
Assuntos
Modelos Biológicos , Processamento de Proteína Pós-Traducional , Receptores de Superfície Celular/metabolismo , Transdução de Sinais , Enzimas/metabolismo , CinéticaRESUMO
The Bicoid morphogen gradient directs the patterning of cell fates along the anterior-posterior axis of the syncytial Drosophila embryo and serves as a paradigm of morphogen-mediated patterning. The simplest models of gradient formation rely on constant protein synthesis and diffusion from anteriorly localized source mRNA, coupled with uniform protein degradation. However, currently such models cannot account for all known gradient characteristics. Recent work has proposed that bicoid mRNA spatial distribution is sufficient to produce the observed protein gradient, minimizing the role of protein transport. Here, we adapt a novel method of fluorescent in situ hybridization to quantify the global spatio-temporal dynamics of bicoid mRNA particles. We determine that >90% of all bicoid mRNA is continuously present within the anterior 20% of the embryo. bicoid mRNA distribution along the body axis remains nearly unchanged despite dynamic mRNA translocation from the embryo core to the cortex. To evaluate the impact of mRNA distribution on protein gradient dynamics, we provide detailed quantitative measurements of nuclear Bicoid levels during the formation of the protein gradient. We find that gradient establishment begins 45 minutes after fertilization and that the gradient requires about 50 minutes to reach peak levels. In numerical simulations of gradient formation, we find that incorporating the actual bicoid mRNA distribution yields a closer prediction of the observed protein dynamics compared to modeling protein production from a point source at the anterior pole. We conclude that the spatial distribution of bicoid mRNA contributes to, but cannot account for, protein gradient formation, and therefore that protein movement, either active or passive, is required for gradient formation.
Assuntos
Padronização Corporal/fisiologia , Drosophila/embriologia , Proteínas de Homeodomínio/metabolismo , Hibridização in Situ Fluorescente/métodos , Modelos Biológicos , RNA Mensageiro/metabolismo , Transativadores/metabolismo , Animais , Simulação por Computador , Proteínas de Drosophila , Proteínas de Fluorescência Verde/metabolismo , Processamento de Imagem Assistida por Computador , Transporte Proteico/fisiologiaRESUMO
Transcription factor combinations play a key role in shaping cellular identity. However, the precise relationship between specific combinations and downstream effects remains elusive. Here, we investigate this relationship within the context of the Drosophila eve locus, which is controlled by gap genes. We measure spatiotemporal levels of four gap genes in heterozygous and homozygous gap mutant embryos and correlate them with the striped eve activity pattern. Although changes in gap gene expression extend beyond the manipulated gene, the spatial patterns of Eve expression closely mirror canonical activation levels in wild type. Interestingly, some combinations deviate from the wild-type repertoire but still drive eve activation. Although in homozygous mutants some Eve stripes exhibit partial penetrance, stripes consistently emerge at reproducible positions, even with varying gap gene levels. Our findings suggest a robust molecular canalization of cell fates in gap mutants and provide insights into the regulatory constraints governing multi-enhancer gene loci.
Assuntos
Proteínas de Drosophila , Drosophila , Animais , Drosophila/metabolismo , Regulação da Expressão Gênica no Desenvolvimento/genética , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Expressão Gênica , Proteínas de Homeodomínio/metabolismoRESUMO
Shape changes of epithelia during animal development, such as convergent extension, are achieved through concerted mechanical activity of individual cells. While much is known about the corresponding large scale tissue flow and its genetic drivers, key open questions regard the cell-scale mechanics, e.g. internal vs external driving forces, and coordination, e.g. bottom-up self-organization vs top-down genetic instruction. To address these questions, we develop a quantitative, model-based analysis framework to relate cell geometry to local tension in recently obtained timelapse imaging data of gastrulating Drosophila embryos. This analysis provides a systematic decomposition of cell shape changes and T1-rearrangements into internally driven, active, and externally driven, passive, contributions. Specifically, we find evidence that germ band extension is driven by active T1 processes that self-organize through positive feedback acting on tensions. More generally, our findings suggest that epithelial convergent extension results from controlled transformation of internal force balance geometry which we quantify with a novel quantification tool for local tension configurations.
RESUMO
Gastrulation movements in all animal embryos start with regulated deformations of patterned epithelial sheets, which are driven by cell divisions, cell shape changes, and cell intercalations. Each of these behaviors has been associated with distinct aspects of gastrulation1-4 and has been a subject of intense research using genetic, cell biological, and more recently, biophysical approaches.5-14 Most of these studies, however, focus either on cellular processes driving gastrulation or on large-scale tissue deformations.15-23 Recent advances in microscopy and image processing create a unique opportunity for integrating these complementary viewpoints.24-28 Here, we take a step toward bridging these complementary strategies and deconstruct the early stages of gastrulation in the entire Drosophila embryo. Our approach relies on an integrated computational framework for cell segmentation and tracking and on efficient algorithms for event detection. The detected events are then mapped back onto the blastoderm shell, providing an intuitive visual means to examine complex cellular activity patterns within the context of their initial anatomic domains. By analyzing these maps, we identified that the loss of nearly half of surface cells to invaginations is compensated primarily by transient mitotic rounding. In addition, by analyzing mapped cell intercalation events, we derived direct quantitative relations between intercalation frequency and the rate of axis elongation. This work is setting the stage for systems-level dissection of a pivotal step in animal development.
Assuntos
Embrião de Mamíferos , Gastrulação , Animais , Forma Celular , Drosophila , MorfogêneseRESUMO
During the maternal-to-zygotic transition, a developing embryo integrates post-transcriptional regulation of maternal mRNAs with transcriptional activation of its own genome. By combining chromosomal ablation in Drosophila with microarray analysis, we characterized the basis of this integration. We show that the expression profile for at least one third of zygotically active genes is coupled to the concomitant degradation of the corresponding maternal mRNAs. The embryo uses transcription and degradation to generate localized patterns of expression, and zygotic transcription to degrade distinct classes of maternal transcripts. Although degradation does not appear to involve a simple regulatory code, the activation of the zygotic genome starts from intronless genes sharing a common cis-element. This cis-element interacts with a single protein, the Bicoid stability factor, and acts as a potent enhancer capable of timing the activity of an exogenous transactivator. We propose that this regulatory mode links morphogen gradients with temporal regulation during the maternal-to-zygotic transition.
Assuntos
Deleção Cromossômica , Proteínas de Drosophila/fisiologia , Drosophila melanogaster/embriologia , Regulação da Expressão Gênica no Desenvolvimento , RNA Mensageiro/metabolismo , Proteínas de Ligação a RNA/fisiologia , Animais , Sequência de Bases , Regulação para Baixo , Proteínas de Drosophila/biossíntese , Drosophila melanogaster/genética , Feminino , Perfilação da Expressão Gênica , Ativação Transcricional/fisiologia , Zigoto/fisiologiaRESUMO
Optogenetic perturbations, live imaging, and time-resolved ChIP-seq assays in Drosophila embryos were used to dissect the ERK-dependent control of the HMG-box repressor Capicua (Cic), which plays critical roles in development and is deregulated in human spinocerebellar ataxia and cancers. We established that Cic target genes are activated before significant downregulation of nuclear localization of Cic and demonstrated that their activation is preceded by fast dissociation of Cic from the regulatory DNA. We discovered that both Cic-DNA binding and repression are rapidly reinstated in the absence of ERK activation, revealing that inductive signaling must be sufficiently sustained to ensure robust transcriptional response. Our work provides a quantitative framework for the mechanistic analysis of dynamics and control of transcriptional repression in development.
Assuntos
Proteínas de Drosophila/metabolismo , Proteínas HMGB/metabolismo , Sistema de Sinalização das MAP Quinases , Proteínas Repressoras/metabolismo , Transporte Ativo do Núcleo Celular , Animais , Núcleo Celular/metabolismo , Proteínas de Drosophila/genética , Drosophila melanogaster , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Proteínas HMGB/genética , Ligação Proteica , Proteínas Repressoras/genéticaRESUMO
The Bicoid morphogen evolved approximately 150 MYA from a Hox3 duplication and is only found in higher dipterans. A major difference between dipteran species, however, is the size of the embryo, which varies up to 5-fold. Although the expression of developmental factors scale with egg length, it remains unknown how this scaling is achieved. To test whether scaling is accounted for by the properties of Bicoid, we expressed eGFP fused to the coding region of bicoid from three dipteran species in transgenic Drosophila embryos using the Drosophila bicoid cis-regulatory and mRNA localization sequences. In such embryos, we find that Lucilia sericata and Calliphora vicina Bicoid produce gradients very similar to the endogenous Drosophila gradient and much shorter than what they would have produced in their own respective species. The common shape of the Drosophila, Lucilia and Calliphora Bicoid gradients appears to be a conserved feature of the Bicoid protein. Surprisingly, despite their similar distributions, we find that Bicoid from Lucilia and Calliphora do not rescue Drosophila bicoid mutants, suggesting that that Bicoid proteins have evolved species-specific functional amino acid differences. We also found that maternal expression and anteriorly localization of proteins other than Bcd does not necessarily give rise to a gradient; eGFP produced a uniform protein distribution. However, a shallow gradient was observed using eGFP-NLS, suggesting nuclear localization may be necessary but not sufficient for gradient formation.
Assuntos
Dípteros/embriologia , Drosophila melanogaster/embriologia , Proteínas de Homeodomínio/fisiologia , Transativadores/fisiologia , Animais , Padronização Corporal/fisiologia , Proteínas de Drosophila , Drosophila melanogaster/genética , Embrião não Mamífero/citologia , Embrião não Mamífero/fisiologia , Proteínas de Homeodomínio/química , Especificidade da Espécie , Transativadores/químicaRESUMO
In a recent paper, Lee and Goldstein develop an explant assay that recapitulates key aspects of gastrulation in C elegans and permits classical embryological manipulations. The resulting detailed analysis of cell behavior will ultimately extend to broader issues, such as, whether morphogenesis can be described as the sum of single-cell events or if unique phenomena emerge at the multicellular level.