In systematic searches for embryonic lethal mutants of Drosophila melanogaster we have identified 15 loci which when mutated alter the segmental pattern of the larva. These loci probably represent the majority of such genes in Drosophila. The phenotypes of the mutant embryos indicate that the process of segmentation involves at least three levels of spatial organization: the entire egg as developmental unit, a repeat unit with the length of two segments, and the individual segment.

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

The reproducibility and precision of biological patterning is limited by the accuracy with which concentration profiles of morphogen molecules can be established and read out by their targets. We consider four measures of precision for the Bicoid morphogen in the Drosophila embryo: the concentration differences that distinguish neighboring cells, the limits set by the random arrival of Bicoid molecules at their targets (which depends on absolute concentration), the noise in readout of Bicoid by the activation of Hunchback, and the reproducibility of Bicoid concentration at corresponding positions in multiple embryos. We show, through a combination of different experiments, that all of these quantities are approximately 10%. This agreement among different measures of accuracy indicates that the embryo is not faced with noisy input signals and readout mechanisms; rather, the system exerts precise control over absolute concentrations and responds reliably to small concentration differences, approaching the limits set by basic physical principles.

Patterning in multicellular organisms results from spatial gradients in morphogen concentration, but the dynamics of these gradients remain largely unexplored. We characterize, through in vivo optical imaging, the development and stability of the Bicoid morphogen gradient in Drosophila embryos that express a Bicoid-eGFP fusion protein. The gradient is established rapidly (approximately 1 hr after fertilization), with nuclear Bicoid concentration rising and falling during mitosis. Interphase levels result from a rapid equilibrium between Bicoid uptake and removal. Initial interphase concentration in nuclei in successive cycles is constant (+/-10%), demonstrating a form of gradient stability, but it subsequently decays by approximately 30%. Both direct photobleaching measurements and indirect estimates of Bicoid-eGFP diffusion constants (D < or = 1 microm(2)/s) provide a consistent picture of Bicoid transport on short ( approximately min) time scales but challenge traditional models of long-range gradient formation. A new model is presented emphasizing the possible role of nuclear dynamics in shaping and scaling the gradient.