Search

The final size and shape of organs results from volume expansion by growth and shape changes by contractility. Complex morphologies arise from differences in growth rate between tissues. We address here how differential growth drives epithelial thickening and doming during the morphogenesis of the growing Drosophila wing imaginal disc. We report that 3D morphology results from elastic deformation due to differential growth between the epithelial cell layer and its enveloping extracellular matrix (ECM). Furthermore, the ECM envelope exhibits differential growth anisotropy (i.e. anisotropic expansion in 3D), growing in-plane on one side, but out of plane on the other side. The elasticity, anisotropy and morphogenesis is fully captured by a mechanical bilayer model. Moreover, differential expression of the Matrix metalloproteinase MMP2 controls growth anisotropy of the two ECM layers. This study shows that the ECM is a controllable mechanical constraint whose intrinsic growth anisotropy directs tissue morphogenesis in a developing organ.

The size and proportions of animals are tightly controlled during development. How this is achieved remains poorly understood. The control of organ size entails coupling of cellular growth and cell division on one hand, and the measure of organ size on the other. In this review we focus on three layers of growth control consisting of genetic patterning, notably chemical gradients, mechanics and energetics which are complemented by a systemic control unit that modulates growth in response to the nutritional conditions and coordinates growth between different organs so as to maintain proportions. Growth factors, often present as concentration dependent chemical gradients, are positive inducers of cellular growth that may be considered as deterministic cues, hence acting as organ-intrinsic controllers of growth. However, the exponential growth dynamics in many developing tissues necessitate more stringent growth control in the form of negative feedbacks. Feedbacks endow biological systems with the capacity to quickly respond to perturbations and to correct the growth trajectory to avoid overgrowth. We propose to integrate chemical, mechanical and energetic control over cellular growth in a framework that emphasizes the self-organizing properties of organ-autonomous growth control in conjunction with systemic organ non-autonomous feedback on growth.

Summary Interfaces between cells with distinct genetic identities elicit signals to organize local cell behaviors driving tissue morphogenesis. The Drosophila embryonic axis extension requires planar polarized enrichment of myosin-II powering oriented cell intercalations. Myosin-II levels are quantitatively controlled by GPCR signaling, whereas myosin-II polarity requires patterned expression of several Toll receptors. How Toll receptors polarize myosin-II and how this involves GPCRs remain unknown. Here, we report that differential expression of a single Toll receptor, Toll-8, polarizes myosin-II through binding to the adhesion GPCR Cirl/latrophilin. Asymmetric expression of Cirl is sufficient to enrich myosin-II, and Cirl localization is asymmetric at Toll-8 expression boundaries. Exploring the process dynamically, we reveal that Toll-8 and Cirl exhibit mutually dependent planar polarity in response to quantitative differences in Toll-8 expression between neighboring cells. Collectively, we propose that the cell surface protein complex Toll-8/Cirl self-organizes to generate local asymmetric interfaces essential for planar polarization of contractility., Graphical abstract, Highlights • Asymmetric expression of a single Toll receptor leads to Myo-II polarization • The adhesion GPCR Cirl binds to Toll-8 mediating Toll-8-induced Myo-II polarization • Toll-8 boundaries generate a Cirl interfacial asymmetry that can polarize Myo-II • Differences in Toll-8 levels lead to interdependent Toll-8 and Cirl planar polarity, Lavalou, Mao et al. report that Toll-8 controls myosin-II planar polarity in Drosophila embryos and wing discs via a physical interaction with the GPCR Cirl/latrophilin. They show that Toll-8 expression boundaries generate a Cirl interfacial asymmetry and propose that it is a potential signal leading to myosin-II polarization.

The Drosophila tracheal system consists of an interconnected network of monolayered epithelial tubes that ensures oxygen transport in the larval and adult body. During tracheal dorsal branch (DB) development, individual DBs elongate as a cluster of cells, led by tip cells at the front and trailing cells in the rear. Branch elongation is accompanied by extensive cell intercalation and cell lengthening of the trailing stalk cells. Although cell intercalation is governed by Myosin II (MyoII)-dependent forces during tissue elongation in the Drosophila embryo that lead to germ-band extension, it remained unclear whether MyoII plays a similar active role during tracheal branch elongation and intercalation. Here, we have used a nanobody-based approach to selectively knock down MyoII in tracheal cells. Our data show that, despite the depletion of MyoII function, tip cell migration and stalk cell intercalation (SCI) proceed at a normal rate. This confirms a model in which DB elongation and SCI in the trachea occur as a consequence of tip cell migration, which produces the necessary forces for the branching process.

Developmental biology research would benefit greatly from tools that enable protein function to be regulated, both systematically and in a precise spatial and temporal manner, in vivo. In recent years, functionalized protein binders have emerged as versatile tools that can be used to target and manipulate proteins. Such protein binders can be based on various scaffolds, such as nanobodies, designed ankyrin repeat proteins (DARPins) and monobodies, and can be used to block or perturb protein function in living cells. In this Primer, we provide an overview of the protein binders that are currently available and highlight recent progress made in applying protein binder-based tools in developmental and synthetic biology.

It is assumed that the description of the exchange processes between heterogeneous natural surfaces and the atmosphere within turbulence closure models is mainly limited by a realistic three-dimensional (3D) representation of the vegetation architecture. Within this contribution we present a method to record the 3D vegetation structure and to use this information to derive model parameters that are suitable for numerical flow models. A mixed conifer forest stand around a clearing was scanned and represented by a dense 3D point cloud applying a terrestrial laser scanner. Thus, the plant area density (PAD) with a resolution of one cubic meter was provided for analysis and for numerical simulations. Multi-level high-frequency wind velocity measurements were recorded simultaneously by 27 ultrasonic anemometers on 4 towers for a period of one year. The relationship between wind speed, Reynolds stress and PAD was investigated and a parametrization of the drag coefficient CD$C_D$ by the PAD is suggested. The derived 3D vegetation model and a simpler model (based on classical forest assessments of the site) were applied in a boundary layer model (BLM) and in large-eddy simulations (LES). The spatial development of the turbulent flow over the clearing is further demonstrated by the results of a wind tunnel experiment. The project showed, that the simulation results were improved significantly by the usage of realistic vegetation models. 3D simulations are necessary to depict the influence of heterogeneous canopies on the turbulent flow. Whereas we found limits for the mapping of the vegetation structure within the wind tunnel, there is a considerable potential for numerical simulations. The field measurements and the LES gave new insight into the turbulent flow in the vicinity and across the clearing. The results show that the zones of intensive turbulence development can not be restricted to the locations found in previous studies with more idealized canopies.

Investigating the role of protein localization is crucial to understand protein function in cells or tissues. However, in many cases the role of different subcellular fractions of given proteins along the apical-basal axis of polarized cells has not been investigatedin vivo, partially due to lack of suitable tools. Here, we present the GrabFP system, a nanobody-based toolbox to modify the localization and the dispersal of GFP-tagged proteins along the apical-basal axis of polarized cells. We show that the GrabFP system is an effective and easy-to-implement tool to mislocalize cytosolic and transmembrane GFP-tagged proteins and thereby functionally investigate protein localization along the apical-basal axis. We use the GrabFP system as a tool to study the extracellular dispersal of the Decapentaplegic (Dpp) protein and show that the Dpp gradient forming in the lateral plane of theDrosophilawing disc epithelium is essential for patterning of the wing imaginal disc.

TheDrosophilatracheal system consists of an interconnected network of monolayered epithelial tubes that ensures oxygen transport in the larval and adult body. During tracheal dorsal branch (DB) development, individual DBs elongate as a cluster of cells, led by tip cells at the front and trailing cells in the rear. Branch elongation is accompanied by extensive cell intercalation and cell lengthening of the trailing stalk cells. While cell intercalation is governed by Myosin II (MyoII)-dependent forces during tissue elongation in theDrosophilaembryo leading to germ-band extension, it remained unclear whether MyoII plays a similar active role during tracheal branch elongation and intercalation. Here, we use a nanobody-based approach to selectively knock-down MyoII in tracheal cells. Our data shows that despite the depletion of MyoII function, tip cells migration and stalk cell intercalation (SCI) proceeds at a normal rate. Therefore, our data confirms a model in which DB elongation and SCI in the trachea occurs as a consequence of tip cell migration, which produces the necessary forces for the branching process.Summary statementBranch elongation duringDrosophilatracheal development mechanistically resembles MyoII-independent collective cell migration; tensile forces resulting from tip cell migration are reduced by cell elongation and passive stalk cell intercalation.AbbreviationsDBDorsal branchDCDorsal closureE-CadE-CadherinGBEGerm-band extensionMRLCMyosin regulatory light chainMyoIIMyosin IISCIstalk cell intercalationSqhSpaghetti squashSxllSex lethalTCTip cellTrTracheomere

The role of protein localization along the apical-basal axis of polarized cells is difficult to investigate in vivo, partially due to lack of suitable tools. Here, we present the GrabFP system, a collection of four nanobody-based GFP-traps that localize to defined positions along the apical-basal axis. We show that the localization preference of the GrabFP traps can impose a novel localization on GFP-tagged target proteins and results in their controlled mislocalization. These new tools were used to mislocalize transmembrane and cytoplasmic GFP fusion proteins in the Drosophila wing disc epithelium and to investigate the effect of protein mislocalization. Furthermore, we used the GrabFP system as a tool to study the extracellular dispersal of the Decapentaplegic (Dpp) protein and show that the Dpp gradient forming in the lateral plane of the Drosophila wing disc epithelium is essential for patterning of the wing imaginal disc. DOI: http://dx.doi.org/10.7554/eLife.22549.001

Applications of flow models to tall plant canopies are limited, amongst other factors, by the lack of detailed information on vegetation structure. A method is presented to record 3D vegetation structure and make this information applicable to the derivation of turbulence parameters suitable for flow models. The relationship between wind speed, drag coefficient (CD) and plant area density (PAD) was experimentally investigated in a mixed conifer forest in the lower part of the Eastern Ore Mountains. Essential information was gathered by collecting multi-level high-frequency wind velocity measurements and a dense 3D representation of the forest was obtained from terrestrial laser scanner data. Wind speed dependence or streamlining was observed for most of the wind directions. Edge effects, i.e. the influence of the here not regarded pressure gradient and the advective terms of the momentum equation, are assumed to cause this heterogeneity. Contrary to the hypothetic shelter effect, which would reduce the drag on sheltered plant parts, the calculated profiles of drag coefficients revealed an increasing CD with PAD (i.e. a dependence on canopy and plant structure).

Protein-protein interactions are crucial for almost all biological processes. Studying such interactions in their native environment is critical but not easy to perform. Recently developed genetically encoded protein binders were shown to function inside living cells. These molecules offer a new, direct way to assess protein function, distribution and dynamics in vivo. A widely used protein binder scaffold are the so-called nanobodies, which are derived from the variable domain of camelid heavy-chain antibodies. Another commonly used scaffold, the DARPins, is based on Ankyrin repeats. In this review, we highlight how these binders can be functionalized in order to study proteins in vivo during the development of multicellular organisms. It is to be anticipated that many more applications for such synthetic protein binders will be developed in the near future.

Bone morphogenetic proteins (BMPs) act as morphogens to control patterning and growth in a variety of developing tissues in different species. How BMP morphogen gradients are established and interpreted in the target tissues has been extensively studied in Drosophila melanogaster. In Drosophila, Decapentaplegic (Dpp), a homologue of vertebrate BMP2/4, acts as a morphogen to control dorsal-ventral patterning of the early embryo and anterior-posterior patterning and growth of the wing imaginal disc. Despite intensive efforts over the last twenty years, how the Dpp morphogen gradient in the wing imaginal disc forms remains controversial, while gradient formation in the early embryo is well understood. In this review, we first focus on the current models of Dpp morphogen gradient formation in these two tissues, and then discuss new strategies using genome engineering and nanobodies to tackle open questions.

Drosophila Decapentaplegic (Dpp) has served as a paradigm to study morphogen-dependent growth control. However, the role of a Dpp gradient in tissue growth remains highly controversial. Two fundamentally different models have been proposed: the 'temporal rule' model suggests that all cells of the wing imaginal disc divide upon a 50% increase in Dpp signalling, whereas the 'growth equalization model' suggests that Dpp is only essential for proliferation control of the central cells. Here, to discriminate between these two models, we generated and used morphotrap, a membrane-tethered anti-green fluorescent protein (GFP) nanobody, which enables immobilization of enhanced (e)GFP::Dpp on the cell surface, thereby abolishing Dpp gradient formation. We find that in the absence of Dpp spreading, wing disc patterning is lost; however, lateral cells still divide at normal rates. These data are consistent with the growth equalization model, but do not fit a global temporal rule model in the wing imaginal disc.

The Drosophila tracheal system consists of an interconnected network of monolayered epithelial tubes that ensures oxygen transport in the larval and adult body. During tracheal dorsal branch (DB) development, individual DBs elongate as a cluster of cells, led by tip cells at the front and trailing cells in the rear. Branch elongation is accompanied by extensive cell intercalation and cell lengthening of the trailing stalk cells. While cell intercalation is governed by Myosin II (MyoII)-dependent forces during tissue elongation in the Drosophila embryo leading to germ-band extension, it remained unclear whether MyoII plays a similar active role during tracheal branch elongation and intercalation. Here, we use a nanobody-based approach to selectively knock-down MyoII in tracheal cells. Our data shows that despite the depletion of MyoII function, tip cells migration and stalk cell intercalation (SCI) proceeds at a normal rate. Therefore, our data confirms a model in which DB elongation and SCI in the trachea occurs as a consequence of tip cell migration, which produces the necessary forces for the branching process.