Geometric control of Myosin-II orientation during axis elongation

The actomyosin cytoskeleton is a crucial driver of morphogenesis. Yet how the behavior of large scale cytoskeletal patterns in deforming tissues emerges from the interplay of geometry, genetics, and mechanics remains incompletely understood. Convergent extension flow in D. melanogaster embryos provides the opportunity to establish a quantitative understanding of the dynamics of anisotropic non-muscle myosin II. Cell-scale analysis of protein localization in fixed embryos suggests that there are complex rules governing how the control of myosin anisotropy is regulated by gene expression patterns. However, technical limitations have impeded quantitative and dynamic studies of this process at the whole embryo level, leaving the role of geometry open. Here we combine in toto live imaging with quantitative analysis of molecular dynamics to characterize the distribution of myosin anisotropy and corresponding genetic patterning. We found pair rule gene expression continuously deformed, flowing with the tissue frame. In contrast, myosin anisotropy orientation remained nearly static, aligned with the stationary dorsal-ventral axis of the embryo. We propose myosin recruitment by a geometrically defined static source, potentially related to the embryo-scale epithelial tension, and account for transient deflections by the interplay of cytoskeletal turnover with junction reorientation by flow. With only one parameter, this model quantitatively accounts for the time course of myosin anisotropy orientation in wild-type, twist, and even-skipped embryos as well as embryos with perturbed egg geometry. Geometric patterning of the cytoskeleton suggests a simple physical strategy to ensure a robust flow and formation of shape.

Regulatory gene function handoff allows essential gene loss in mosquitoes

Regulatory genes are often multifunctional and constrained, which results in evolutionary conservation. It is difficult to understand how a regulatory gene could be lost from one species’ genome when it is essential for viability in closely related species. The gene paired is a classic Drosophila pair-rule gene, required for formation of alternate body segments in diverse insect species. Surprisingly, paired was lost in mosquitoes without disrupting body patterning. Here, we demonstrate that a paired family member, gooseberry, has acquired paired-like expression in the malaria mosquito Anopheles stephensi. Anopheles-gooseberry CRISPR-Cas9 knock-out mutants display pair-rule phenotypes and alteration of target gene expression similar to what is seen in Drosophila and beetle paired mutants. Thus, paired was functionally replaced by the related gene, gooseberry, in mosquitoes. Our findings document a rare example of a functional replacement of an essential regulatory gene and provide a mechanistic explanation of how such loss can occur.

bicoid RNA localization requires the trans-Golgi network

Background The formation of the bicoid (bcd) mRNA gradient is a crucial step for Bcd protein gradient formation in Drosophila. In the past, a microtubule (MT)-based cortical network had been shown to be indispensable for bcd mRNA transport to the posterior. Results We report the identification of a MT-binding protein CLASP/Chb as the first component associated with this cortical MT network. Since CLASPs in vertebrates were shown to serve as an acentriolar microtubule organization center (aMTOC) in concert with trans-Golgi proteins, we examined the effect of the Drosophila trans-Golgins on bcd localization and gradient formation. Using a genetic approach, we demonstrate that the Drosophila trans-Golgins dGCC88, dGolgin97 and dGCC185 indeed affect bcd mRNA localization during oocyte development. Consequently, the bcd mRNA is already mislocalized before the egg is fertilized. The expression domains of genes downstream of the hierarchy of bcd, e.g. of the gap gene empty spiracles or of the pair-rule gene even-skipped are changed, indicating an altered segmental anlagen, due to a faulty bcd gradient. Thus, at the end of embryogenesis, trans-Golgin mutants show bcd-like cuticle phenotypes. Conclusions Our data provides evidence that the Golgi as a cellular member of the secretory pathway exerts control on bcd localization which indicates that bcd gradient formation is probably more intricate than previously presumed.

Shifting roles of Drosophila pair-rule gene orthologs: segmental expression and function in the milkweed bug Oncopeltus fasciatus

The discovery of pair-rule genes (PRGs) in Drosophila revealed the existence of an underlying two-segment-wide prepattern directing embryogenesis. The milkweed bug Oncopeltus, a hemimetabolous insect, is a more representative arthropod: most of its segments form sequentially after gastrulation. Here we report the expression and function of orthologs of the complete set of nine Drosophila PRGs in Oncopeltus. Seven Of-PRG-orthologs are expressed in stripes in the primordia of every segment, rather than every-other segment, Of-runt is PR-like, and several are also expressed in the segment addition zone. RNAi-mediated knockdown of Of-odd-skipped, paired and sloppy-paired impacted all segments, with no indication of PR-like register. We confirm that Of-E75A is expressed in PR-like stripes, although it is not PR in Drosophila, demonstrating the existence of an underlying PR-like prepattern in Oncopeltus. These findings reveal that a switch occurred in regulatory circuits leading to segment formation: while several holometabolous insects are “Drosophila-like,” utilizing PRG-orthologs for PR-patterning, most Of-PRGs are expressed segmentally in Oncopeltus, a more basally-branching insect. Thus, an evolutionarily stable phenotype – segment formation – is directed by alternate regulatory pathways in diverse species.Summary StatementDespite the broad of conservation of segmentation in insects, the regulatory genes underlying this process in Drosophila have different roles in the hemipteran, Oncopeltus fasciatus.

Dynamic patterning by theDrosophilapair-rule network reconciles long-germ and short-germ segmentation

ABSTRACTDrosophilasegmentation is a well-established paradigm for developmental pattern formation. However, the later stages of segment patterning, regulated by the “pair-rule” genes, are still not well understood at the systems level. Building on established genetic interactions, I construct a logical model of theDrosophilapair-rule system that takes into account the demonstrated stage-specific architecture of the pair-rule gene network. Simulation of this model can accurately recapitulate the observed spatiotemporal expression of the pair-rule genes, but only when the system is provided with dynamic “gap” inputs. This result suggests that dynamic shifts of pair-rule stripes are essential for segment patterning in the trunk, and provides a functional role for observed posterior-to-anterior gap domain shifts that occur during cellularisation. The model also suggests revised patterning mechanisms for the parasegment boundaries, and accounts for theeven-skippednull mutant phenotype. Strikingly, a slightly modified version of the model is able to pattern segments in either simultaneous or sequential modes, depending only on initial conditions. This suggest that fundamentally similar mechanisms may underlie segmentation in short-germ and long-germ arthropods.