Triangles bridge the scales: Quantifying cellular contributions to tissue deformation

In this article, we propose a general framework to study the dynamics and topology of cellular networks that capture the geometry of cell packings in two-dimensional tissues. Such epithelia undergo large-scale deformation during morphogenesis of a multicellular organism. Large-scale deformations emerge from many individual cellular events such as cell shape changes, cell rearrangements, cell divisions, and cell extrusions. Using a triangle-based representation of cellular network geometry, we obtain an exact decomposition of large-scale material deformation. Interestingly, our approach reveals contributions of correlations between cellular rotations and elongation as well as cellular growth and elongation to tissue deformation. Using this Triangle Method, we discuss tissue remodeling in the developing pupal wing of the fly Drosophila melanogaster.Comment: 26 pages, 18 figure

Active dynamics of tissue shear flow

We present a hydrodynamic theory to describe shear flows in developing epithelial tissues. We introduce hydrodynamic fields corresponding to state properties of constituent cells as well as a contribution to overall tissue shear flow due to rearrangements in cell network topology. We then construct a generic linear constitutive equation for the shear rate due to topological rearrangements and we investigate a novel rheological behaviour resulting from memory effects in the tissue. We identify two distinct active cellular processes: generation of active stress in the tissue, and actively driven topological rearrangements. We find that these two active processes can produce distinct cellular and tissue shape changes, depending on boundary conditions applied on the tissue. Our findings have consequences for the understanding of tissue morphogenesis during development

Dancing disclinations in confined active nematics

The spontaneous emergence of collective flows is a generic property of active fluids and often leads to chaotic flow patterns characterised by swirls, jets, and topological disclinations in their orientation field. However, the ability to achieve structured flows and ordered disclinations is of particular importance in the design and control of active systems. By confining an active nematic fluid within a channel, we find a regular motion of disclinations, in conjunction with a well defined and dynamic vortex lattice. As pairs of moving disclinations travel through the channel, they continually exchange partners producing a dynamic ordered state, reminiscent of Ceilidh dancing. We anticipate that this biomimetic ability to self-assemble organised topological disclinations and dynamically structured flow fields in engineered geometries will pave the road towards establishing new active topological microfluidic devices

Gene Expression Changes in the Motor Cortex Mediating Motor Skill Learning

The primary motor cortex (M1) supports motor skill learning, yet little is known about the genes that contribute to motor cortical plasticity. Such knowledge could identify candidate molecules whose targeting might enable a new understanding of motor cortical functions, and provide new drug targets for the treatment of diseases which impair motor function, such as ischemic stroke. Here, we assess changes in the motor-cortical transcriptome across different stages of motor skill acquisition. Adult rats were trained on a gradually acquired appetitive reach and grasp task that required different strategies for successful pellet retrieval, or a sham version of the task in which the rats received pellet reward without needing to develop the reach and grasp skill. Tissue was harvested from the forelimb motor-cortical area either before training commenced, prior to the initial rise in task performance, or at peak performance. Differential classes of gene expression were observed at the time point immediately preceding motor task improvement. Functional clustering revealed that gene expression changes were related to the synapse, development, intracellular signaling, and the fibroblast growth factor (FGF) family, with many modulated genes known to regulate synaptic plasticity, synaptogenesis, and cytoskeletal dynamics. The modulated expression of synaptic genes likely reflects ongoing network reorganization from commencement of training till the point of task improvement, suggesting that motor performance improves only after sufficient modifications in the cortical circuitry have accumulated. The regulated FGF-related genes may together contribute to M1 remodeling through their roles in synaptic growth and maturation.McGovern Institute for Brain Research at MITNational Institutes of Health (U.S.) ((NIH grant 1-RC1-NS068103-01)National Institutes of Health (U.S.) (NIH grant R01-MH084966)Roberto Rocca Education Program (Fellowship)Massachusetts Institute of Technology. Undergraduate Research Opportunities Program (Fellowship)Italy. Ministero dell'istruzione, dell'università e della ricerca (MIUR grant RBIN04H5AS)Italy. Ministero dell'istruzione, dell'università e della ricerca (MIUR grant RBLA03FLJC)Italy. Ministero dell'istruzione, dell'università e della ricerca (FIRB n. RBAP10L8TY

Myosin Light-Chain Kinase Is Necessary for Membrane Homeostasis in Cochlear Inner Hair Cells

The structural homeostasis of the cochlear hair cell membrane is critical for all aspects of sensory transduction, but the regulation of its maintenance is not well understood. In this report, we analyzed the cochlear hair cells of mice with specific deletion of myosin light chain kinase (MLCK) in inner hair cells. MLCK-deficient mice showed impaired hearing, with a 5- to 14-dB rise in the auditory brainstem response (ABR) thresholds to clicks and tones of different frequencies and a significant decrease in the amplitude of the ABR waves. The mutant inner hair cells produced several ball-like structures around the hair bundles in vivo, indicating impaired membrane stability. Inner hair cells isolated from the knockout mice consistently displayed less resistance to hypoosmotic solution and less membrane F-actin. Myosin light-chain phosphorylation was also reduced in the mutated inner hair cells. Our results suggest that MLCK is necessary for maintaining the membrane stability of inner hair cells

ZO-1 Stabilizes the Tight Junction Solute Barrier through Coupling to the Perijunctional Cytoskeleton

ZO-1 binds numerous transmembrane and cytoplasmic proteins and is required for assembly of both adherens and tight junctions, but its role in defining barrier properties of an established tight junction is unknown. We depleted ZO-1 in MDCK cells using siRNA methods and observed specific defects in the barrier for large solutes, even though flux through the small claudin pores was unaffected. This permeability increase was accompanied by morphological alterations and reorganization of apical actin and myosin. The permeability defect, and to a lesser extent morphological changes, could be rescued by reexpression of either full-length ZO-1 or an N-terminal construct containing the PDZ, SH3, and GUK domains. ZO-2 knockdown did not replicate either the permeability or morphological phenotypes seen in the ZO-1 knockdown, suggesting that ZO-1 and -2 are not functionally redundant for these functions. Wild-type and knockdown MDCK cells had differing physiological and morphological responses to pharmacologic interventions targeting myosin activity. Use of the ROCK inhibitor Y27632 or myosin inhibitor blebbistatin increased TER in wild-type cells, whereas ZO-1 knockdown monolayers were either unaffected or changed in the opposite direction; paracellular flux and myosin localization were also differentially affected. These studies are the first direct evidence that ZO-1 limits solute permeability in established tight junctions, perhaps by forming a stabilizing link between the barrier and perijunctional actomyosin

Author response

Understanding the mechanisms regulating development requires a quantitative characterization of cell divisions, rearrangements, cell size and shape changes, and apoptoses. We developed a multiscale formalism that relates the characterizations of each cell process to tissue growth and morphogenesis. Having validated the formalism on computer simulations, we quantified separately all morphogenetic events in the Drosophila dorsal thorax and wing pupal epithelia to obtain comprehensive statistical maps linking cell and tissue scale dynamics. While globally cell shape changes, rearrangements and divisions all significantly participate in tissue morphogenesis, locally, their relative participations display major variations in space and time. By blocking division we analyzed the impact of division on rearrangements, cell shape changes and tissue morphogenesis. Finally, by combining the formalism with mechanical stress measurement, we evidenced unexpected interplays between patterns of tissue elongation, cell division and stress. Our formalism provides a novel and rigorous approach to uncover mechanisms governing tissue development. DOI: http://dx.doi.org/10.7554/eLife.08519.00

Surdités hérédtaires (rôles de la myosine VIIa dans le développement de la cellule sensorielle auditive)

La myosine VIIa, dont le déficit est à l'origine du syndrome de Usher de type IB (USH1B), associe surdité profonde et rétinopathie pigmentaire. Son expression, au sein de l'oreille interne, est restreinte aux cellules sensorielles (cellules ciliées). La myosine VIIa et la myosine 1c ont été impliquées dans le processus d'adaptation du courant de transduction, dans les cellules ciliées de l oreille interne, en réponse à la stimulation sonore. J'ai entrepris la recherche de nouvelles molécules impliquées dans ce processus par un crible double-hybride chez la levure. J'ai identifié PHR1, une protéine transmembranaire à domaine Pleckstrin Homology, comme partenaire de liaison commun aux myosines 1c et VIIa. En supposant que PHR1 soit localisé dans les stéréocils, il pourrait ancrer les deux myosines à la membrane, et jouer lui aussi un rôle dans le processus d'adaptation. je me suis ensuite concentré sur le rôle de la myosine VIIa aux jonctions apicales entre les cellules ciliées et leurs cellules de soutien. Ces jonctions jouent notamment un rôle crucial dans le maintien du potentiel endocochléaire, qui contribue à la dépolarisation cellulaire en réponse à une stimulation sonore. Par une approche biochimique, j ai montré que la myosine VIIa se lie à shroom2, une protéine à domaine PDZ, qui se lie à ZO1 et à l actine. L'abondance de shroom2 aux jonctions serrées des cellules ciliées, continuellement soumises à des forces de cisaillement en réponse à la stimulation sonore, incite à penser que shroom2, en conjonction avec la myosine VIIa, est impliquée dans le renforcement des jonctions serrées.PARIS-BIUSJ-Thèses (751052125) / SudocPARIS-BIUSJ-Physique recherche (751052113) / SudocSudocFranceF

Shroom2, a myosin-VIIa- and actin-binding protein, directly interacts with ZO-1 at tight junctions.

International audienceDefects in myosin VIIa lead to developmental anomalies of the auditory and visual sensory cells. We sought proteins interacting with the myosin VIIa tail by using the yeast two-hybrid system. Here, we report on shroom2, a submembranous PDZ domain-containing protein that is associated with the tight junctions in multiple embryonic and adult epithelia. Shroom2 directly interacts with the C-terminal MyTH4-FERM domain of myosin VIIa and with F-actin. In addition, a shroom2 fragment containing the region of interaction with F-actin was able to protect actin filaments from cytochalasin-D-induced disruption in MDCK cells. Transfection experiments in MDCK and LE (L fibroblasts that express E-cadherin) cells led us to conclude that shroom2 is targeted to the cell-cell junctions in the presence of tight junctions only. In Ca(2+)-switch experiments on MDCK cells, ZO-1 (also known as TJP1) preceded GFP-tagged shroom2 at the differentiating tight junctions. ZO-1 directly interacts with the serine- and proline-rich region of shroom2 in vitro. Moreover, the two proteins colocalize in vivo at mature tight junctions, and could be coimmunoprecipitated from brain and cochlear extracts. We suggest that shroom2 and ZO-1 form a tight-junction-associated scaffolding complex, possibly linked to myosin VIIa, that bridges the junctional membrane to the underlying cytoskeleton, thereby contributing to the stabilization of these junctions

Extracting multiple surfaces from 3D microscopy images in complex biological tissues with the Zellige software tool

International audienceBackgroundEfficient tools allowing the extraction of 2D surfaces from 3D-microscopy data are essential for studies aiming to decipher the complex cellular choreography through which epithelium morphogenesis takes place during development. Most existing methods allow for the extraction of a single and smooth manifold of sufficiently high signal intensity and contrast, and usually fail when the surface of interest has a rough topography or when its localization is hampered by other surrounding structures of higher contrast. Multiple surface segmentation entails laborious manual annotations of the various surfaces separately.ResultsAs automating this task is critical in studies involving tissue-tissue or tissue-matrix interaction, we developed the Zellige software, which allows the extraction of a non-prescribed number of surfaces of varying inclination, contrast, and texture from a 3D image. The tool requires the adjustment of a small set of control parameters, for which we provide an intuitive interface implemented as a Fiji plugin.ConclusionsAs a proof of principle of the versatility of Zellige, we demonstrate its performance and robustness on synthetic images and on four different types of biological samples, covering a wide range of biological contexts