Elongation of the body axis is accompanied by the assembly of a polarized cytoarchitecture that provides the basis for directional cell behavior. We find that planar polarity in the Drosophila embryo is established through a sequential enrichment of actin-myosin cables and adherens junction proteins in complementary surface domains. F-actin accumulation at AP interfaces represents the first break in planar symmetry and occurs independently of proper junctional protein distribution at DV interfaces. Polarized cells engage in a novel program of locally coordinated behavior to generate multicellular rosette structures that form and resolve in a directional fashion. Actin-myosin structures align across multiple cells during rosette formation, and adherens junction proteins assemble in a stepwise fashion during rosette resolution. Patterning genes essential for axis elongation selectively affect the frequency and directionality of rosette formation. We propose that the generation of higher-order rosette structures links local cell interactions to global tissue reorganization during morphogenesis.

Summary Cell rearrangements shape the Drosophila embryo through spatially regulated changes in cell shape and adhesion. We show that Bazooka/Par-3 (Baz) is required for the planar polarized distribution of myosin II and adherens junction proteins and polarized intercalary behavior is disrupted in baz mutants. The myosin II activator Rho-kinase is asymmetrically enriched at anterior and posterior borders of intercalating cells in a pattern complementary to Baz. Loss of Rho-kinase results in expansion of the Baz domain and activated Rho-kinase is sufficient to exclude Baz from the cortex. The planar polarized distribution of Baz requires its C-terminal domain. Rho-kinase can phosphorylate this domain and inhibit its interaction with phosphoinositide membrane lipids, suggesting a mechanism by which Rho-kinase could regulate Baz association with the cell cortex. These results demonstrate that Rho-kinase plays an instructive role in planar polarity by targeting Baz/Par-3 and myosin II to complementary cortical domains.

Epithelial remodeling determines the structure of many organs in the body through changes in cell shape, polarity and behavior and is a major area of study in developmental biology. Accurate and high-throughput methods are necessary to systematically analyze epithelial organization and dynamics at single-cell resolution. We developed SEGGA, an easy-to-use software for automated image segmentation, cell tracking and quantitative analysis of cell shape, polarity and behavior in epithelial tissues. SEGGA is free, open source, and provides a full suite of tools that allow users with no prior computational expertise to independently perform all steps of automated image segmentation, semi-automated user-guided error correction, and data analysis. Here we use SEGGA to analyze changes in cell shape, cell interactions and planar polarity during convergent extension in the Drosophila embryo. These studies demonstrate that planar polarity is rapidly established in a spatiotemporally regulated pattern that is dynamically remodeled in response to changes in cell orientation. These findings reveal an unexpected plasticity that maintains coordinated planar polarity in actively moving populations through the continual realignment of cell polarity with the tissue axes.

We are using the maize leaf as an experimental system to ask how positional information establishes the proximal/distal axis.A mature maize leaf has three regions, the distal blade that functions in photosynthesis, the proximal sheath that wraps the stalk, and the ligule, marking a sharp boundary between blade and sheath. The recessive liguleless mutants remove the ligule, but the distinction of sheath and blade remains. When combined with a dominant mutant, Wavy auricle in blade, regions of the leaf are only sheath and leaves are very narrow. A similar phenotype is seen in the single mutant, Liguleless narrow (Lgn). Lgn heterozygotes have narrow leaves and no ligule at the margins.Lgn homozygotes leaves lack distinction of blade and sheath, and have no stem or reproductive parts. Lgn encodes a serine threonine kinase and the mutated version fails to autophosphorylate, suggesting that correct signaling from LGN is needed for plant architecture as well as proximal/distal patterning of the leaf. In contrast with these mutants, the dominant homeobox mutation, Knotted1, recreates proximal distal patterning due to misexpression in the blade. Normally, kn1 is expressed only in the meristem. We propose that kn1 establishes the proximal end of the leaf at its inception. The gain of function Kn1 phenotype is suppressed in the Lgn background. This result suggests that Lgn may provide or transmit a distal signal that interacts with that of kn1.As a cell divides in a developing embryo, the orientation of the mitotic spindle is a key decision that will affect the fates of the daughter cells and the embryo as a whole. However, what role the regulated orientation of mitotic spindles plays in the tissue shaping movements of early embryogenesis remains largely unknown. We are investigating mitotic spindle orientation in Xenopus laevis embryos during the early stages of gastrulation, as the embryo undergoes epiboly. Unlike blastula stages in Xenopus embryos, where spindle orientation follows the long axis of cells, at the onset of gastrulation mitotic spindles in the outer cell layer of the embryo consistently align parallel to the apical cortex, irrespective of cell shape. Live imaging of the mitotic spindles reveals that they take up their orientation during spindle assembly and maintain it throughout mitosis, despite undergoing rapid movements within the orientation plane. Furthermore, the spindles maintain a constant position along the apico-basal axis of the cell. We are now working to reveal the molecular mechanisms controlling mitotic spindle orientation and positioning during gastrulation. We report that depolymerising F-actin with Latrunculin B leads to a randomisation of spindle orientation and basally positioned spindles. Loss of Myosin-10 also affects spindle orientation but results in apically positioned spindles. These findings show that F-actin and Myosin-10 are both required to correctly position and orient the spindle and also reveal the presence of opposing forces on the spindle along the apico-basal axis. During...

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