Should I shrink or should I grow: cell size changes in tissue morphogenesis.

Cells change shape, move, divide, and die to sculpt tissues. Common to all these cell behaviours are cell size changes, which have recently emerged as key contributors to tissue morphogenesis. Cells can change their mass-the number of macromolecules they contain-or their volume-the space they encompass. Changes in cell mass and volume occur through different molecular mechanisms and at different timescales, slow for changes in mass and rapid for changes in volume. Therefore, changes in cell mass and cell volume, which are often linked, contribute to the development and shaping of tissues in different ways. Here, we review the molecular mechanisms by which cells can control and alter their size, and we discuss how changes in cell mass and volume contribute to tissue morphogenesis. The role that cell size control plays in developing embryos is only starting to be elucidated. Research on the signals that control cell size will illuminate our understanding of the cellular and molecular mechanisms that drive tissue morphogenesis.

Integrin-based adhesions promote cell-cell junction and cytoskeletal remodelling to drive embryonic wound healing.

Embryos repair wounds rapidly, with no inflammation or scarring. Embryonic wound healing is driven by the collective movement of the cells around the lesion. The cells adjacent to the wound polarize the cytoskeletal protein actin and the molecular motor non-muscle myosin II, which accumulate at the wound edge forming a supracellular cable around the wound. Adherens junction proteins, including E-cadherin, are internalized from the wound edge and localize to former tricellular junctions at the wound margin, in a process necessary for cytoskeletal polarity. We found that the cells adjacent to wounds in the Drosophila embryonic epidermis polarized Talin, a core component of cell-extracellular matrix (ECM) adhesions, which preferentially accumulated at the wound edge. Integrin knockdown and inhibition of integrin binding delayed wound closure and reduced actin polarization and dynamics around the wound. Additionally, disrupting integrins caused a defect in E-cadherin reinforcement at tricellular junctions along the wound edge, suggesting crosstalk between integrin-based and cadherin-based adhesions. Our results show that cell-ECM adhesion contributes to embryonic wound repair and reveal an interplay between cell-cell and cell-ECM adhesion in the collective cell movements that drive rapid wound healing.

Cortical tension promotes Kibra degradation via Par-1.

The Hippo pathway is an evolutionarily conserved regulator of tissue growth. Multiple Hippo signaling components are regulated via proteolytic degradation. However, how these degradation mechanisms are themselves modulated remains unexplored. Kibra is a key upstream pathway activator that promotes its own ubiquitin-mediated degradation upon assembling a Hippo signaling complex. Here, we demonstrate that Hippo complex-dependent Kibra degradation is modulated by cortical tension. Using classical genetic, osmotic, and pharmacological manipulations of myosin activity and cortical tension, we show that increasing cortical tension leads to Kibra degradation, whereas decreasing cortical tension increases Kibra abundance. Our study also implicates Par-1 in regulating Kib abundance downstream of cortical tension. We demonstrate that Par-1 promotes ubiquitin-mediated Kib degradation in a Hippo complex-dependent manner and is required for tension-induced Kib degradation. Collectively, our results reveal a previously unknown molecular mechanism by which cortical tension affects Hippo signaling and provide novel insights into the role of mechanical forces in growth control.

Myosin waves and a mechanical asymmetry guide the oscillatory migration of Drosophila cardiac progenitors.

Heart development begins with the formation of a tube as cardiac progenitors migrate from opposite sides of the embryo. Abnormal cardiac progenitor movements cause congenital heart defects. However, the mechanisms of cell migration during early heart development remain poorly understood. Using quantitative microscopy, we found that in Drosophila embryos, cardiac progenitors (cardioblasts) migrated through a sequence of forward and backward steps. Cardioblast steps were associated with oscillatory non-muscle myosin II waves that induced periodic shape changes and were necessary for timely heart tube formation. Mathematical modeling predicted that forward cardioblast migration required a stiff boundary at the trailing edge. Consistent with this, we found a supracellular actin cable at the trailing edge of the cardioblasts that limited the amplitude of the backward steps, thus biasing the direction of cell movement. Our results indicate that periodic shape changes coupled with a polarized actin cable produce asymmetrical forces that promote cardioblast migration.

Rap1 coordinates cell-cell adhesion and cytoskeletal reorganization to drive collective cell migration in vivo.

Collective cell movements contribute to tissue development and repair and spread metastatic disease. In epithelia, cohesive cell movements require reorganization of adherens junctions and the actomyosin cytoskeleton. However, the mechanisms that coordinate cell-cell adhesion and cytoskeletal remodeling during collective cell migration in vivo are unclear. We investigated the mechanisms of collective cell migration during epidermal wound healing in Drosophila embryos. Upon wounding, the cells adjacent to the wound internalize cell-cell adhesion molecules and polarize actin and the motor protein non-muscle myosin II to form a supracellular cable around the wound that coordinates cell movements. The cable anchors at former tricellular junctions (TCJs) along the wound edge, and TCJs are reinforced during wound closure. We found that the small GTPase Rap1 was necessary and sufficient for rapid wound repair. Rap1 promoted myosin polarization to the wound edge and E-cadherin accumulation at TCJs. Using embryos expressing a mutant form of the Rap1 effector Canoe/Afadin that cannot bind Rap1, we found that Rap1 signals through Canoe for adherens junction remodeling, but not for actomyosin cable assembly. Instead, Rap1 was necessary and sufficient for RhoA/Rho1 activation at the wound edge. The RhoGEF Ephexin localized to the wound edge in a Rap1-dependent manner, and Ephexin was necessary for myosin polarization and rapid wound repair, but not for E-cadherin redistribution. Together, our data show that Rap1 coordinates the molecular rearrangements that drive embryonic wound healing, promoting actomyosin cable assembly through Ephexin-Rho1, and E-cadherin redistribution through Canoe, thus enabling rapid collective cell migration in vivo.

Contractile and expansive actin networks in Drosophila: Developmental cell biology controlled by network polarization and higher-order interactions.

Actin networks are central to shaping and moving cells during animal development. Various spatial cues activate conserved signal transduction pathways to polarize actin network assembly at sub-cellular locations and to elicit specific physical changes. Actomyosin networks contract and Arp2/3 networks expand, and to affect whole cells and tissues they do so within higher-order systems. At the scale of tissues, actomyosin networks of epithelial cells can be coupled via adherens junctions to form supracellular networks. Arp2/3 networks typically integrate with distinct actin assemblies, forming expansive composites which act in conjunction with contractile actomyosin networks for whole-cell effects. This review explores these concepts using examples from Drosophila development. First, we discuss the polarized assembly of supracellular actomyosin cables which constrict and reshape epithelial tissues during embryonic wound healing, germ band extension, and mesoderm invagination, but which also form physical borders between tissue compartments at parasegment boundaries and during dorsal closure. Second, we review how locally induced Arp2/3 networks act in opposition to actomyosin structures during myoblast cell-cell fusion and cortical compartmentalization of the syncytial embryo, and how Arp2/3 and actomyosin networks also cooperate for the single cell migration of hemocytes and the collective migration of border cells. Overall, these examples show how the polarized deployment and higher-order interactions of actin networks organize developmental cell biology.

The α-Catenin mechanosensing M region is required for cell adhesion during tissue morphogenesis.

α-Catenin couples the cadherin-catenin complex to the actin cytoskeleton. The mechanosensitive α-Catenin M region undergoes conformational changes upon application of force to recruit interaction partners. Here, we took advantage of the tension landscape in the Drosophila embryo to define three different states of α-Catenin mechanosensing in support of cell adhesion. Low-, medium-, and high-tension contacts showed a corresponding recruitment of Vinculin and Ajuba, which was dependent on the α-Catenin M region. In contrast, the Afadin homolog Canoe acts in parallel to α-Catenin at bicellular low- and medium-tension junctions but requires an interaction with α-Catenin for its tension-sensitive enrichment at high-tension tricellular junctions. Individual M region domains make complex contributions to cell adhesion through their impact on interaction partner recruitment, and redundancies with the function of Canoe. Our data argue that α-Catenin and its interaction partners are part of a cooperative and partially redundant mechanoresponsive network that supports AJs remodeling during morphogenesis.

Septate junction proteins are required for cell shape changes, actomyosin reorganization and cell adhesion during dorsal closure in Drosophila.

Septate junctions (SJs) serve as occluding barriers in invertebrate epithelia. In Drosophila, at least 30 genes are required for the formation or maintenance of SJs. Interestingly, loss-of-function mutations in core SJ components are embryonic lethal, with defects in developmental events such as head involution and dorsal closure (DC) that occur prior to the formation of a mature SJ, indicating a role for these proteins in mid-embryogenesis independent of their occluding function. To understand this novel function in development, we examined loss-of-function mutations in three core SJ proteins during the process of DC. DC occurs during mid-embryogenesis to seal a dorsal gap in the epidermis following germ band retraction. Closure is driven by contraction of the extraembryonic amnioserosa cells that temporarily cover the dorsal surface and by cell shape changes (elongation) of lateral epidermal cells that bring the contralateral sheets together at the dorsal midline. Using live imaging and examination of fixed tissues, we show that early events in DC occur normally in SJ mutant embryos, but during later closure, coracle, Macroglobulin complement-related and Neurexin-IV mutant embryos exhibit slower rates of closure and display aberrant cells shapes in the dorsolateral epidermis, including dorsoventral length and apical surface area. SJ mutant embryos also show mild defects in actomyosin structures along the leading edge, but laser cutting experiments suggest similar tension and viscoelastic properties in SJ mutant versus wild type epidermis. In a high percentage of SJ mutant embryos, the epidermis tears free from the amnioserosa near the end of DC and live imaging and immunostaining reveal reduced levels of E-cadherin, suggesting that defective adhesion may be responsible for these tears. Supporting this notion, reducing E-cadherin by half significantly enhances the penetrance of DC defects in coracle mutant embryos.

Powering morphogenesis: multiscale challenges at the interface of cell adhesion and the cytoskeleton.

Among the defining features of the animal kingdom is the ability of cells to change shape and move. This underlies embryonic and postembryonic development, tissue homeostasis, regeneration, and wound healing. Cell shape change and motility require linkage of the cell's force-generating machinery to the plasma membrane at cell-cell and cell-extracellular matrix junctions. Connections of the actomyosin cytoskeleton to cell-cell adherens junctions need to be both resilient and dynamic, preventing tissue disruption during the dramatic events of embryonic morphogenesis. In the past decade, new insights radically altered the earlier simple paradigm that suggested simple linear linkage via the cadherin-catenin complex as the molecular mechanism of junction-cytoskeleton interaction. In this Perspective we provide a brief overview of our current state of knowledge and then focus on selected examples highlighting what we view as the major unanswered questions in our field and the approaches that offer exciting new insights at multiple scales from atomic structure to tissue mechanics.

Crumbs complex-directed apical membrane dynamics in epithelial cell ingression.

Epithelial cells often leave their tissue context and ingress to form new cell types or acquire migratory ability to move to distant sites during development and tumor progression. Cells lose their apical membrane and epithelial adherens junctions during ingression. However, how factors that organize apical-basal polarity contribute to ingression is unknown. Here, we show that the dynamic regulation of the apical Crumbs polarity complex is crucial for normal neural stem cell ingression. Crumbs endocytosis and recycling allow ingression to occur in a normal timeframe. During early ingression, Crumbs and its complex partner the RhoGEF Cysts support myosin and apical constriction to ensure robust ingression dynamics. During late ingression, the E3-ubiquitin ligase Neuralized facilitates the disassembly of the Crumbs complex and the rapid endocytic removal of the apical cell domain. Our findings reveal a mechanism integrating cell fate, apical polarity, endocytosis, vesicle trafficking, and actomyosin contractility to promote cell ingression, a fundamental morphogenetic process observed in animal development and cancer.

PyJAMAS: open-source, multimodal segmentation and analysis of microscopy images.

SUMMARY: Our increasing ability to resolve fine details using light microscopy is matched by an increasing need to quantify images in order to detect and measure phenotypes. Despite their central role in cell biology, many image analysis tools require a financial investment, are released as proprietary software, or are implemented in languages not friendly for beginners, and thus are used as black boxes. To overcome these limitations, we have developed PyJAMAS, an open-source tool for image processing and analysis written in Python. PyJAMAS provides a variety of segmentation tools, including watershed and machine learning-based methods; takes advantage of Jupyter notebooks for the display and reproducibility of data analyses; and can be used through a cross-platform graphical user interface or as part of Python scripts via a comprehensive application programming interface. AVAILABILITY AND IMPLEMENTATION: PyJAMAS is open-source and available at https://bitbucket.org/rfg_lab/pyjamas. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

p38-mediated cell growth and survival drive rapid embryonic wound repair.

Embryos repair wounds rapidly, with no inflammation or scarring, in a process that involves polarization of the actomyosin cytoskeleton. Actomyosin polarization results in the assembly of a contractile cable around the wound that drives wound closure. Here, we demonstrate that a contractile actomyosin cable is not sufficient for rapid wound repair in Drosophila embryos. We show that wounding causes activation of the serine/threonine kinase p38 mitogen-activated protein kinase (MAPK) in the cells adjacent to the wound. p38 activation reduces the levels of wound-induced reactive oxygen species in the cells around the wound, limiting wound size. In addition, p38 promotes an increase in volume in the cells around the wound, thus facilitating the collective cell movements that drive rapid wound healing. Our data indicate that p38 regulates cell volumes through the sodium-potassium-chloride cotransporter NKCC1. Our work reveals cell growth and cell survival as cell behaviors critical for embryonic wound repair.

Myosin cables control the timing of tissue internalization in the Drosophila embryo.

Compartment boundaries prevent cell mixing during animal development. In the early Drosophila embryo, the mesectoderm is a group of glial precursors that separate ectoderm and mesoderm, forming the ventral midline. Mesectoderm cells undergo one round of oriented divisions during axis elongation and are eventually internalized 6 h later. Using spinning disk confocal microscopy and image analysis, we found that after dividing, mesectoderm cells reversed their planar polarity. The polarity factor Bazooka was redistributed to mesectoderm-mesectoderm cell interfaces, and the molecular motor non-muscle Myosin II and its upstream activator Rho-kinase (Rok) accumulated at mesectoderm-ectoderm (ME) interfaces, forming supracellular cables flanking the mesectoderm on either side of the tissue. Laser ablation revealed the presence of increased tension at ME cables, where Myosin was stabilized, as shown by fluorescence recovery after photobleaching. We used laser nanosurgery to reduce tension at the ME boundary, and we found that Myosin fluorescence decreased rapidly, suggesting a role for tension in ME boundary maintenance. Mathematical modelling predicted that increased tension at the ME boundary was necessary to prevent the premature establishment of contacts between the two ectodermal sheets on opposite sides of the mesectoderm, thus controlling the timing of mesectoderm internalization. We validated the model in vivo: Myosin inhibition disrupted the linearity of the ME boundary and resulted in early internalization of the mesectoderm. Our results suggest that the redistribution of Rok polarizes Myosin and Bazooka within the mesectoderm to establish tissue boundaries, and that ME boundaries control the timely internalization of the mesectoderm as embryos develop.

Pak1 and PP2A antagonize aPKC function to support cortical tension induced by the Crumbs-Yurt complex.

The Drosophila polarity protein Crumbs is essential for the establishment and growth of the apical domain in epithelial cells. The protein Yurt limits the ability of Crumbs to promote apical membrane growth, thereby defining proper apical/lateral membrane ratio that is crucial for forming and maintaining complex epithelial structures such as tubes or acini. Here, we show that Yurt also increases Myosin-dependent cortical tension downstream of Crumbs. Yurt overexpression thus induces apical constriction in epithelial cells. The kinase aPKC phosphorylates Yurt, thereby dislodging the latter from the apical domain and releasing apical tension. In contrast, the kinase Pak1 promotes Yurt dephosphorylation through activation of the phosphatase PP2A. The Pak1-PP2A module thus opposes aPKC function and supports Yurt-induced apical constriction. Hence, the complex interplay between Yurt, aPKC, Pak1, and PP2A contributes to the functional plasticity of Crumbs. Overall, our data increase our understanding of how proteins sustaining epithelial cell polarization and Myosin-dependent cell contractility interact with one another to control epithelial tissue architecture.

Schwannoma del nervio vago / Vagus-nerve schwannoma

Introducción:los schwannomas, también llamados neurilemomas o neurinomas, son tumores de origen neuroectodérmico, encapsulados y benignos, que generalmente aparecen como una masa laterocervical asintomática de crecimiento lento. Son pocos frecuentes en el área de cabeza y cuello.Caso clínico:mujer de 39 años con una historia de un año de evolución de una masa cervical derecha de crecimiento lento, pulsátil, dolorosa y móvil en el lado derecho del cuello. La imagen de la angiotomografía computarizada preoperatoria demostró que la masa involucraba la bifurcación carotídea, compatible con un paraganglioma carotídeo. La disección cuidadosa a través de una cervicotomía longitudinal mostró que la masa surgía de un único fascículo del nervio vago. La masa se envió a anatomía patológica con el resultado de schwannoma. En el posoperatorio la paciente presentó una función sensorial y motora del vago normal, con un leve síndrome de Horner.Discusión:los tumores nerviosos del cuello surgen a partir de grandes troncos nerviosos, los IX, X, XI y XII pares craneales, el simpático cervical, el plexo cervical y el plexo braquial. El nervio vago es un lugar infrecuente de aparición, con muy pocos casos reportados. Son lesiones de diagnóstico preoperatorio difícil, cuyo tratamiento de elección es la exéresis quirúrgica; en ocasiones, es difícil conservar el nervio de origen.(AU)

Introduction:schwannomas, neurilemomas or neurinomas, are benign, encapsulated, neuroectodermal tumors that generally appear as a slow-growing, asymptomatic laterocervical mass. They are rare in the head and neck area.Clinical case:a 39-year-old woman with a one-year history of a slowly enlarging, pulsatile, painful and movable right cervical mass on the right side of the neck. Preoperative computed angiotomography image demonstrated that the mass involved the carotid bifurcation compatible with a carotid paraganglioma. Careful dissection through a longitudinal cervicotomy, showed that the mass arose from a single fascicle of the vagus nerve. The mass was sent to the pathologists and found to be a schwannoma. The patient had normal vagus sensory and motor function postoperatively, with a slight Horner syndrome.Discussion:nerve tumors of the neck arise from large nerve trunks, the IX, X, XI, and XII cranial nerves, the cervical sympathetic, the cervical plexus, and the brachial plexus. The vagus nerve is an infrequent place of appearance with very few reported cases. They are difficult preoperative diagnosis lesions; whose treatment of choice is surgical excision; sometimes it is difficult to preserve the nerve of origin.(AU)

Tratamiento híbrido de aneurisma micótico de la aorta visceral / Hybrid treatment of a mycotic aneurysm of the visceral aorta

Introducción: los aneurismas micóticos son una entidad grave y poco frecuente, cuyo tratamiento quirúrgico se ha basado históricamente en la resección del tejido infectado y en la reconstrucción arterial. En los últimos años, la importancia del uso de técnicas de reparación endovascular ha ido en aumento. Caso clínico: se presenta el caso de una mujer de 84 años con un aneurisma micótico de la aorta visceral tratado mediante un abordaje híbrido. Se revascularizan las arterias mesentérica superior y hepática y, posteriormente, se coloca una endoprótesis de aorta. Discusión: el empleo de dispositivos endovasculares para la reparación de los aneurismas micóticos genera controversia debido a que no se elimina el tejido infectado. Sin embargo, existen publicaciones que avalan su uso, ya que permiten el tratamiento, principalmente, de pacientes de alto riesgo quirúrgico sin aumentar el número de complicaciones.(AU)

Introduction: mycotic aneurysms are a serious and rare entity, whose surgical treatment has historically been based on resection of infected tissue and arterial reconstruction. In recent years, the importance of using endovascular repair techniques has increased. Case report: an 84-year-old woman with a mycotic aneurysm of the visceral aorta is presented, treated by a hybrid approach, revascularizing the superior mesenteric and hepatic arteries, and the placement of an aortic stent. Discussion: the use of endovascular devices for the repair of mycotic aneurysms generates controversy due to the maintenance of the infected tissue. However, there are publications that support its use since they allow the treatment mainly of patients at high surgical risk not increasing the rate of complications.(AU)

The recycling endosome protein Rab25 coordinates collective cell movements in the zebrafish surface epithelium.

In emerging epithelial tissues, cells undergo dramatic rearrangements to promote tissue shape changes. Dividing cells remain interconnected via transient cytokinetic bridges. Bridges are cleaved during abscission and currently, the consequences of disrupting abscission in developing epithelia are not well understood. We show that the Rab GTPase Rab25 localizes near cytokinetic midbodies and likely coordinates abscission through endomembrane trafficking in the epithelium of the zebrafish gastrula during epiboly. In maternal-zygotic Rab25a and Rab25b mutant embryos, morphogenic activity tears open persistent apical cytokinetic bridges that failed to undergo timely abscission. Cytokinesis defects result in anisotropic cell morphologies that are associated with a reduction of contractile actomyosin networks. This slows cell rearrangements and alters the viscoelastic responses of the tissue, all of which likely contribute to delayed epiboly. We present a model in which Rab25 trafficking coordinates cytokinetic bridge abscission and cortical actin density, impacting local cell shape changes and tissue-scale forces.

Multiscale In Vivo Imaging of Collective Cell Migration in Drosophila Embryos.

Coordinated cell movements drive embryonic development and tissue repair, and can also spread disease. Time-lapse microscopy is an integral part in the study of the cell biology of collective cell movements. Advances in imaging techniques enable monitoring dynamic cellular and molecular events in real time within living animals. Here, we demonstrate the use of spinning disk confocal microscopy to investigate coordinated cell movements and epithelial-to-mesenchymal-like transitions during embryonic wound closure in Drosophila. We describe image-based metrics to quantify the efficiency of collective cell migration. Finally, we show the application of super-resolution radial fluctuation microscopy to obtain multidimensional, super-resolution images of protrusive activity in collectively moving cells in vivo. Together, the methods presented here constitute a toolkit for the modern analysis of collective cell migration in living animals.