Apical constriction facilitates epithelial sheet bending and invagination during morphogenesis1 , 2 . 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 3-7. However, it is unclear whether other force-generating mechanisms can drive this process. Here, we use real-time imaging and quantitative image analysis of Drosophila gastrulation to show that apical constriction of ventral furrow cells is pulsed. Repeated constrictions, which are asynchronous between neighboring 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 inward. The transcription factors Twist and Snail differentially regulate pulsed constriction. Expression of snail initiates actin-myosin network contractions while 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 sub-cellular ratchet to incrementally reduce apical area.During Drosophila gastrulation, apical constriction of ventral cells facilitates the formation of a ventral furrow and the subsequent internalization of the presumptive mesoderm. Although myosin is known to localize to the apical cortex of constricting ventral furrow cells [8][9][10][11] , how myosin produces force to drive constriction is not known. Understanding this mechanism requires a quantitative analysis of cell and cytoskeletal dynamics. Therefore, we developed methods to visualize and quantify apical cell shape using Spider-GFP, a GFP-tagged transmembrane protein that outlines individual cells (Fig. 1a, 1b , Supplemental Figure 1 , Video 1) 12 . Ventral cells constricted to ~50 % of their initial apical area before the onset of invagination and continued to constrict during invagination (Fig. 1c, 1e). Although the average apical area steadily decreased at a rate of ~5 μm 2 /min, individual cells exhibited transient pulses of rapid constriction that exceeded 10-15 μm 2 /min (Fig. 1d, 1f, 1g, and Video 2). During the initial 2 minutes of constriction, weak constriction pulses were often interrupted by periods of cell stretching. However, at 2 minutes constriction pulses increased in magnitude and cell shape

Transcription factor Twist promotes cell junctions to link individual cells into a contractile network responsible for the apical constriction pulses during epithelial morphogenesis.

The brain’s visual cortex processes information concerning form, pattern, and motion within functional maps that reflect the layout of neuronal circuits. We analyzed functional maps of orientation preference in the ferret, tree shrew, and galago—three species separated since the basal radiation of placental mammals more than 65 million years ago—and found a common organizing principle. A symmetry-based class of models for the self-organization of cortical networks predicts all essential features of the layout of these neuronal circuits, but only if suppressive long-range interactions dominate development. We show mathematically that orientation-selective long-range connectivity can mediate the required interactions. Our results suggest that self-organization has canalized the evolution of the neuronal circuitry underlying orientation preference maps into a single common design.

The principles governing the functional organization and development of long-range network interactions in the neocortex remain poorly understood. Using in vivo wide-field and 2-photon calcium imaging of spontaneous activity patterns in mature ferret visual cortex, we find widespread modular correlation patterns that accurately predict the local structure of visually-evoked orientation columns several millimeters away. Longitudinal imaging demonstrates that long-range spontaneous correlations are present early in cortical development prior to the elaboration of horizontal connections, and predict mature network structure. Silencing feed-forward drive through retinal or thalamic blockade does not eliminate early long-range correlated activity, suggesting a cortical origin. Circuit models containing only local, but heterogeneous, connections are sufficient to generate long-range correlated activity by confining activity patterns to a low-dimensional subspace via multi-synaptic short-range interactions. These results suggest that local connections in early cortical circuits can generate structured long-range network correlations that guide the formation of visually-evoked distributed functional networks.

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 constriction1. 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 prior to 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 neighboring 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 toward the initiating cells, thereby facilitating tissue deformation. Our data thus establish a direct link between modification of epithelial polarity and initiation of epithelial folding.

During Drosophila gastrulation, the ventral mesodermal cells constrict their apices, undergo a series of coordinated cell-shape changes to form a ventral furrow (VF) and are subsequently internalized. Although it has been well documented that apical constriction is necessary for VF formation, the mechanism by which apical constriction transmits forces throughout the bulk tissue of the cell remains poorly understood. In this work, we develop a computational vertex model to investigate the role of the passive mechanical properties of the cellular blastoderm during gastrulation. We introduce to our knowledge novel data that confirm that the volume of apically constricting cells is conserved throughout the entire course of invagination. We show that maintenance of this constant volume is sufficient to generate invagination as a passive response to apical constriction when it is combined with region-specific elasticities in the membranes surrounding individual cells. We find that the specific sequence of cell-shape changes during VF formation is critically controlled by the stiffness of the lateral and basal membrane surfaces. In particular, our model demonstrates that a transition in basal rigidity is sufficient to drive VF formation along the same sequence of cell-shape change that we observed in the actual embryo, with no active force generation required other than apical constriction.

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.two-photon imaging | 4D reconstruction | segmentation

Networks of epithelial and endothelial tubes are essential for the function of organs such as the lung, kidney, and vascular system. The sizes and shapes of these tubes are highly regulated to match their individual functions. Defects in tube size can cause debilitating diseases such as polycystic kidney disease (PKD) and ischemia1,2. It is therefore critical to understand how tube dimensions are regulated. Here we identify the tyrosine kinase Src as an instructive regulator of epithelial tube length in the Drosophila tracheal system. Loss-of-function Src42 mutations shorten tracheal tubes while Src42 over-expression elongates them. Surprisingly, Src42 acts distinctly from known tube size pathways and regulates both the amount of apical surface growth and, with the conserved formin dDaam, the direction of growth. Quantitative 3-D image analysis reveals that Src42 and dDaam mutant tracheal cells expand more in the circumferential than the axial dimension, resulting in tubes that are shorter in length – but larger in diameter – than WT tubes. Thus, Src42 and dDaam control tube dimensions by regulating the direction of anisotropic growth, a mechanism that has not previously been described.