Comparative Morphogenesis between Plants and Animals: the role of Mechanical Signals / Morphogenèse comparée des plantes et des animaux: le rôle des signaux mécaniques

Séminaire organisé par Olivier Hamant (INRAE, Reproduction et Développement des Plantes, Lyon, France) du 17 au 22 février 2020

Ce séminaire a fait l’objet de la publication suivante : Olivier Hamant, Bruno Moulia, Stéphane Douady, Fluctuations shape plants through proprioception, Science, 3 Apr 2021, Vol. 372, Issue 6540.

Résumé

Dans Forme et croissance (1917), D’Arcy Thompson souligne les interactions inévitables entre la physique et la biologie. Toutefois, ce concept fondateur est resté dans l’ombre de la biologie moléculaire pendant de nombreuses années. Au cours des deux dernières décennies, la communauté scientifique a fait des avancées significatives pour rapprocher la mécanique de la génétique moléculaire et du développement chez les plantes et les animaux. Bien que partager les connaissances chez ces deux règnes soit essentiel pour extraire les principes communs de la morphogenèse, cette analyse comparative reste à poursuivre. C’est ce que ce séminaire espère stimuler.

Summary

In On growth and forms (1917), D’Arcy Thompson stresses the inevitable interactions between physics and biology. Yet, this concept remained in the shadow of molecular biology for many years. In the past two decades, the scientific community has made major steps to bridge mechanics with molecular genetics and development in both plants and animals. Although insights from both kingdoms are essential to extract the key common principles behind morphogenesis, this comparative analysis remains to be pursued. This is what this seminar is hoping to trigger. 

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Report

Olivier Hamant (RDP, Lyon, France) discussed the role of mechanical signals in plant development. Multicellular organisms exhibit reproducible shapes, yet at the cell level, growth can be extremely heterogeneous and variable. What are the buffering mechanisms that filter such heterogeneitity and variability? Here he took the example of plant organs where final shape only depends on cell division and cell elongation. His team and others showed that shape – and growth – derived forces act as signals that orient microtubules and cellulose microfibrils in the cell walls. This response channels key biological features, such as cell shape or cell division plane orientation. Such mechanical feedback contributes to organ shape reproducibility. Surprisingly, the response of microtubule to stress in the wild type is not optimal, but suboptimal. Notably, phenotypic variability can also emerge from a too strong response to mechanical stress. Looking for molecular regulators of developmental robustness and transcriptional noise, his team identified interactions with mechanotransduction players. Altogether, this work reveals the mechanical complexity behind the robustness of organ shapes, and puts forward the question of suboptimality in biology.

Dagmar Iber (ETH, Basel) discussed how the stereotypic positions of branch points in the mouse lung and kidney epithelial tree can arise from interactions between morphogens and their receptors. She also showed that the 3D packing of cells in epithelia can be accounted to the minimization of the lateral contact surface energy. 

Virgile Viasnoff (MBI Singapore) presented his latest results on lumenogenesis. This mechanism, common to plant and animals, consists in creating, gaps, and lumens between cells to allow the formation of ducts. The creation of tubes occurs following several scenarios that are worth comparing. In his talk, he presented the work of his group in understanding how small canaliculi are formed inside liver. These canaliculi serve as collecting bile secretion. Their morphogenesis is very poorly understood although deficient formation leads to bile flow defects and consequently diseases (Cholestasis). He adopted a deconstrutivist approach to understand how an osmotically driven growth of canaliculi can anisotropically lead to the development of a tube. He showed how single hepatocytes (the main cellular constituents of the liver) can be induced to form spherical hemi-lumen by contact with an inert material, demonstrating that lumenogenesis initiation does not require the concomitant development of lumen in the neighboring cells. He then showed how the spatial geometry of extra cellular matrix provides a guidance tool to favor the development of tubulogenesis in the direction of lowest intra-cellular tension. Lastly he showed the existence of special protrusions crossing the lumen cavity, bridging, suturing, the developing lumen to putatively minimize its extension in the lateral direction, and consequently favoring its extension along the tubular axis.

Stuart Newman (New York Medical college, USA) discussed the physico-genetics of development and the predictability of morphological evolution. The notion that biological forms are limited and predictable defies commonly accepted models of evolution, which favor the idea that morphology is molded by selection for adaption and arrived at purely opportunistically. In contrast, in sciences other than biology, it is recognized that a fixed range of forms is inherent to every type of matter, and that variability, where it exists, is only expressed within that range. This presentation argues that animal tissues have liquid-like and viscoelastic properties that make their repertoire of morphological motifs, such as multilayering, lumens, segments and appendages physically inherent and readily realizable. Developing plant tissues, in contrast, are more complex materials with no clear nonliving counterparts, making their physical modeling less straightforward. The case of the vertebrate limb skeleton was presented in detail as an illustration of the interplay between genetic change and the transformation of material processes in the evolution of animal development.

Elizabeth Haswell (Washington University, St Louis, USA) described one well-established molecular mechanism for force sensing, the activation of mechanosensitive (MS) ion channels. The Mechanosensitive channel of Small conductance (MscS) from E. coli functions as a hypo-osmotic safety valve, opening in response to increased membrane tension and preventing cellular rupture. Genes predicted to encode MscS homologs are found in genomes from all three kingdoms of life. Her team has been characterizing the structure, function, and regulation of ten MscS-Like (MSL) proteins in the model plant Arabidopsis thaliana. Based on their modest homology to MscS and high topological diversity, she has proposed that MSLs might (1) sense and respond to sources of membrane tension other than environmental hypo-osmotic shock; (2) be regulated by mechanisms in addition to membrane tension; and (3) signal in ways that are separable from ion flux. She discussed recent, unpublished results that suggest that MSL channels serve to maintain mechanical homeostasis, or  “mechanostasis”, at the plasma membrane in response to a wide array of abiotic and biotic stresses.

Patrick Lemaire (CRBM, Montpellier, France) studies the embryos of ascidians, a group of small marine invertebrates some of which are seen as delicacies in France and Japan (sea squirts, sea pineapples). He chose this group of animals because the different species included in this group have very different genomes, but develop almost identically. To understand this apparent paradox, his team combines fluorescent imaging approaches in live embryos, genome sequencing, classical embryology and computer modelling. His hope is that this work will allow a better understanding of the, sometimes, tortuous paths of species evolution. The team’s major aims are: 1) to elucidate the evolution of the repertoire of developmental ascidian regulatory proteins; 2) to quantify ascidian morphogenesis and its evolution through advanced light-sheet microscopy and computational image analysis.

Gwyneth Ingram (RDP, Lyon, France) discussed the mechanical control of seed morphogenesis in Arabidopsis. Plant morphogenesis is a complex process that relies on chemical and mechanical interactions between cells and tissues. The growth of the developing seed of Arabidopsis is the product of the mechanical interaction between two genetically and physically distinct tissues: the endosperm and the testa (i.e. seed coat). It has been proposed that endosperm pressure, the driving force of seed growth, generates mechanical tension in the surrounding tissues, which drives cell wall stiffening in a specific mechanosensitive layer of the testa, thus restricting growth. Here, she tested the implications of such an incoherent mechanical feedforward loop, where endosperm pressure directly induces growth but indirectly represses it through its effect on the mechanical properties of the testa, in the control of seed growth and size. Through the development of mathematical models and a multiscale analysis of seed development, she showed that this mechanism can qualitatively account for the growth pattern observed both in wild type seeds but also in mutants showing altered seed mechanical properties. This work thus sheds new light in the control of organ size in plants.

Yusuke Toyama (MBI Singapore) has been investigating the mechanical role of apoptosis, or programmed cell death, which is the most common mechanism of eliminating damaged or unnecessary cells during embryonic development, tissue homeostasis, and certain pathological conditions. It is well known that when a cell undergoes apoptosis within a tissue, the apoptotic cell is expelled from its neighboring non-dying cells. Many labs, including his own, show that this mechanical process of cell extrusion is driven by the formation and contraction of the actomyosin cables in the dying and the neighboring cells, and/or by the lamellipodial crawling of the neighboring cells. However, how cell mechanics arises upon apoptotic cell extrusion and feedbacks to cellular and molecular function especially in the neighboring non-dying cells is largely elusive. During the presentation, he presented his latest results on how mechanical tension and biochemical natures are altered in the surrounding tissue as a consequence of apoptosis, and how these factors contribute to cell proliferation in the neighboring cells.

Emmanuel Farge (Institut Curie, Paris, France) described a feedback regulation on active biochemical and biomechanical patterning by animal tissue physical morphology in embryo and tumour development. From embryos to adults, the development of biomechanical shape is genetically regulated by the expression of biochemical patterning genes. His team has established that the expression of biochemical patterning genes is reversely regulated by the shape changes associated to embryonic and tumorigenic biomechanical development. He has discovered that genes specifying tissue differentiation are mechanosensitive. By combining genetic and pharmacological with mechanical and micro-magnetic tools, respectively preventing and rescuing endogenous morphogenetic movements with physiological forces in vivo, he found that the developmental gene Twist is mechanical stimulated by first endogenous morphogenetic movements of the embryo, in the tissues that will be internalized to form internal organs (anterior gut, mesodermal all other organs). And that this mechanically induced expression is vitally required for specification of these tissues to make these organs functional. His team found that the mechanical activation of a –-catenin (-cat) biochemical reaction at junctions by the first morphogenetic movements of embryogenesis, leading to its release to the cytoplasm and the DNA nucleus, is involved and conserved in earliest mesoderm differentiation in the vertebrate zebrafish in addition to the un-vertebrate Drosophila, two species having directly diverged from the last common animal ancestor in which mesoderm emerged. This allowed to propose mechanical activation of -cat signalling as having been involved in mesoderm differentiation and complex animals evolutionary emergence, a still opened question of animal evolution. His team also found mechanical cues triggering the Fog-dependent apical stabilisation of Myo-II leading to mesoderm invagination, and initiating endoderm invagination, i.e triggering gastrulation in Drosophila embryos. Extended to cancer biology, he found mechanical activation of -cat conserved in mice and involved in the mechanical activation of tumorigenic genes, in the healthy epithelium compressed by the neighbouring tumour, in response to tumour growth pressure in vivo.

Anja Geitmann (McGill University, Montreal, Canada) investigated morphogenesis and structure of the plant leaf epidermis. Pavement cells in the leaf epidermis of many plant species form intricate wavy patterns which are generated through a growth mechanism governed by spatio-temporal control of the material properties of the cell wall – the extracellular matrix enveloping all plant cells. She investigated how the cell wall regulates the morphogenetic process in these cells and which initial steps lead to the characteristic undulations in the cell circumference. Her team shows that non-uniform distribution of cellulose microfibrils and demethylated pectin correlate with spatial differences in cell wall stiffness but intervene at different developmental stages. Lobe initiation involves a modulation of cell wall stiffness through the local enrichment in demethylated pectin, whereas only the subsequent increase in lobe amplitude is mediated by the stress-induced deposition of aligned cellulose microfibrils. Finite element simulations lead her to propose that both steps are preceded by a turgor-driven mechanical buckling event that serves as the initial trigger for the multi-step morphogenetic process. The wavy cell shapes in the leaf epidermis have been hypothesized to improve the tensile strength of the tissue by increasing cell-cell contact. She puts this hypothesis to test by stretching strips of epidermis and observing the propagation of fractures depending on cell shape patterns. Her results indicate that the wavy interlocking cell patterns increase the tear resistance of the plant leaf epidermis; an ingenious defense strategy at the plants’ most exposed surface.

Malcom Bennett (University of Nottingham, UK) investigated how plants sense mechanical signals underground. Plant growth and development is often regulated by their environmental in the form of light, chemical and mechanical-based information. Soil presents plants with many mechanical challenges, particularly as new seedlings establishing themselves and exploring for key resources such as water and nutrients. Plants have developed a number of adaptive strategies to establish themselves. For example, germinating seedlings for a hook like structure to protect the shoot apical meristem. When pushing up through overlaying soil to reach the surface to photosynthesise. His team revealed that the overlaying soil provides a mechanical signal that triggers hook formation. Components of the plant microtubule and cell wall machinery are required to generate a gradient of the hormone auxin across hook cells, causing a differential growth response which leads to hook formation. He describes a mechanistic framework for controlling hook formation, involving several mechano-sensing and response components and signals triggering this important mechano-regulated adaptive response. Seedling roots must also penetrate soil, as they forage for key resources and provide the new plant with anchorage.  Roots can encounter compacted soil, that increases soil density, limiting the availability of water and nutrients. Soil compaction poses a serious challenge to crop cultivation world-wide, impacting root penetration. Modern agriculture practices have exacerbated soil compaction, largely due to deployment of heavier machinery and poor tillage practices, severely degrading ~ 65 million hectares of land globally. To improve penetration through compact soil, roots have been reported to undergo adaptive growth responses, including increasing radial expansion of root tips. However, the mechanistic basis underpinning these root adaptive responses remain unclear. The potential mechanisms explaining how roots sense and adapt to soil compaction remain to be explored.

Atef Asnacios (MSC, Paris, France) is interested in the physics of isolated living cells. In particular, he tries to bring out the role of mechanical phenomena within biological processes. For this purpose, he designed a microplates setup amenable to apply controlled forces on isolated cells, but also to measure forces generated by cell themselves. More particularly, his interest focuses on cell adhesion, geometry and force generation, mechanotransduction of animal and plant cells, and cellular interactions within the immune system.

Jacques Dumais (UAI, Vina Del Mar, Chile) presented the mechanisms of division plane selection in plant cells. Errera’s rule states that plant cell divide such that the new wall achieves a local area minimum while creating daughter cells of equal size. The probability of a specific minimum depends on its area as compared to other minima. However, the probabilistic Errera’s rule as yet to be connected to structural components of the cell. Here he simulates microtubule assemblies within 2D cells with stabilizing edges and find that microtubule arrays naturally align with the shortest axis of the cell, as long as one condition is fulfilled: the cell edges must be slightly stabilizing for microtubules, as opposed to edges that systematically induce catastrophes of the incident microtubules. Strikingly, the resulting alignments extracted from a variety of cell geometries are in good agreement with the prediction of the probabilistic Errera’s rule, with a β parameter that depends on the simulation parameters. The orientation process is found to be robust, and has been tested for varying boundary conditions and varying dynamical features such as treadmilling, microtubule severing or branch nucleation. Since it has been observed that the MT alignment direction coincides with the direction of the future cell wall, this study offers a bridge between empirical cell division rule and microtubule assembly.

Richard Smith (JIC, Norwich, UK) uses mathematical and computer simulation techniques to investigate questions in plant development. Working in close collaboration with experimental biologists, he develops cellular-level simulation models of hormone signalling and patterning in plant tissue. These models involve a biochemical aspect, genes, proteins, hormones, combined with growing, changing geometry as cells divide and tissues grow. His group is interested in the interaction between these two processes. How genes control physical properties of cells resulting in growth, and how the resulting change in geometry and physical forces feeds back on signalling and gene regulation. With this in mind, he is researching methods to quantify mechanical properties in plant tissues, to facilitate the construction of biophysically based simulation models of plant growth.

Muriel Grammont (LBMC, Lyon, France) discussed follicle morphogenesis in Drosophila. It is unknown how external forces and constraints impact morphogenesis in a neighboring tissue. To address this, her team used the Drosophilaovarian follicle, where a cluster of 15 nurse cells and a posteriorly located oocyte are surrounded by a layer of epithelial cells, which are themselves resting on a basement membrane. It is known that as the nurse cells grow, the overlying epithelial cells flatten in a wave that begins in the anterior and that this flattening depends on the TGFß signaling. On the one hand, she demonstrates that an anterior to posterior gradient of decreasing cytoplasmic pressure is present across the nurse cells and that this gradient acts through TGFß to control both the triggering and the progression of the wave of epithelial cell flattening. Her data indicate that intrinsic nurse cell growth is important to control proper nurse cell pressure. Finally, she reveals that nurse cell pressure and subsequent TGFß activity in the StC combine to increase follicle elongation in the anterior, which is crucial for allowing nurse cell growth and pressure control. On the other hand, the epithelial cells are surrounded by a basement membrane. She proves that BM softens around the flattening cells and that this softening depends on TGF pathway. She also demonstrates that interactions between BM constituents are necessary for cell flattening. Altogether, these results show that BM mechanical properties and the inner cytoplasmic pressure in the nurse cells have an important role in shaping neighboring epithelial cells.

Bruno Moulia (PIAF, Clermont-Ferrand, France) uses a bio-mechanical approach of the morphogenesis of plant, including works on leaf and stem display and on the process of mechanosensing and its relevance to adaptation to wind and gravity. This is due both to the conviction that solid and fluid mechanics are very helpful when studying the shaping of plant organs and the relationships between structures and functions, and to a personal equal inclination toward physics and plant sciences. Major findings from these works involve i) the biomechanical analysis of leaf rolling and unrolling in grasses and the importance of morpho-structural aspects in driving rolling phenotypic variability, ii) system analysis of mechanosensing in various organisms, and the demonstration that strainsensing rather than stress-sensing was involved in the process of mechanoperception in plants, iii) the recent validation of an integrative model of mechanosensitive control over tree growth using quantitative expression of primary mechanosensitive genes, and iv) the recent discovery of the role of strain-proprioception in the control of active tropic movement, as well v) a novel method for the spatio-temporal analysis of the regulation of quantitative expression of genes in growing tissues

Henrik Jönsson (SLCU, Cambridge, UK) develops computational morphodynamics models at the cellular level describing multicellular tissues such as the shoot apical meristem. The models are developed in close collaboration with experimental groups and describe the dynamics of gene regulatory networks, hormone transport and signalling, cell growth and division, and mechanical properties. Integral for the research is the iterative evaluation of the models and their parameters to new experimental data, mainly in the form of live microscopy data.

Bénédicte Sanson (University of Cambridge, UK) focuses her research on two fundamental and conserved morphogenetic phenomena, axis extension and compartmental boundary formation, for which her teams has evidence of an integration between the function of genes and the action of mechanical forces in the developing tissues. She studies these in a model organism, the Drosophila embryo, because this is one of the simplest (and cheapest) multicellular models that are genetically tractable. In addition, this embryo is very accessible to in vivo imaging, develops fast and is increasingly exploited as a paradigm for the mathematic modeling of morphogenesis. She analyses a window of development that encompasses both axis extension and compartmental boundary formation (Diagram). Axis extension starts shortly after gastrulation with the trunk ectoderm (the germ-band) elongating in the antero-posterior axis. Compartmental boundaries separating each parasegments form during germ-band extension. Her research is interdisciplinary, combining genetic, quantitative and in silico approaches to find novel and universal morphogenetic rules.

Miltos Tsiantis (Max Planck, Cologne, Germany) proposed a growth based framework for diversification of leaf form. How genes modify cell and tissue growth to create morphological diversity is a key question in biology. Arabidopsis thaliana and Cardamine hirsuta leaves, are respectively simple and divided to leaflets.  This presentation discussed how the combination of live imaging, computational modelling and genetics can help decompose these divergent shapes into their cell level constituent elements: growth amount, direction and differentiation. This approach showed that that leaf form depends on the interplay of a default organ-wide growth mode with local anisotropic growth foci generated by a self-organizing marginal patterning mechanism. Shape differences between the two species have two origins, each controlled by a different homeobox gene.  First, the SHOOTMERISTEMLESS genes causes differential distribution of global growth relative to marginal patterning, that fuelling leaflet emergence.  Second the REDUCED COMPLEXITY gene causes localized growth inhibition around emerging leaflets thus accentuating growth differences created by patterning. He demonstrated the predictivity of our findings by reconstructing key features of C. hirsuta leaf morphology in Arabidopsis and provided a growth-based framework for understanding evolution of leaf form.

 

Communications

• Atef ASNACIOS (Université de Paris, MSC, France) – Une comparaison mécanique des cellules animales et végétales   
• Malcom BENNETT (University of Nottingham CPIB, UK)  – Uncovering mechanical signaling mechanisms operating during the hidden half of plant development  
• Jacques DUMAIS (Universidad Adolfo Ibanez, Chili)  – The Role of Stress and Geometry in plant Cell Division        
• Emmanuel FARGE  (Institut Curie/ Inserm France)  – Signalisation Mécano-Biochimique: de l’Émergence du Mésoderme au cours de l’Évolution animale à l’Induction Mécanique de la Tumorigenèse.  
• Anja GEITMANN (Mac Gill University of Montreal, Canada) – Force, form and function       
• Muriel GRAMMONT (Ecole Normale Supérieure de Lyon, LBMC, France) –  External forces and constraints shape the Drosophila egg  
• Olivier HAMANT (RDP, Ecole Normale Supérieure de Lyon France) –  Signaux mécaniques et robustesse du développement       
• Elizabeth HASWELL (Washington University in St Louis, USA) –   Mechanosensitive Ion Channels in the Green Lineage       
• Dagmar IBER  (ETH Zurich, Suisse)  – From Networks to Function – Computational Models of Organogenesis   
• Gwyneth INGRAM (RDP, Ecole Normale Supérieure de Lyon, France) –  Seeds under pressure, mechanical interplay during seed development     
• Sten Henrik JONSSON (University of Cambridge, SLCU, UK) – From genes to shape in plant development  
• Patrick LEMAIRE  (Université de Montpellier, CRBM, France) –  Analyse de la mécanique et de la variabilité du développement embryonnaire de l’ascidie        
• Bruno MOULIA  (INRA Clermont-Ferrand, France)  –  Standing up to wind and gravity while growing up : the acclimative shaping of plants (including trees) by mechanical signals           
• Stuart NEWMAN (New York Medical College,USA) – Multicellular Matter and the Inherency of Biological Form 
• Bénédicte SANSON (University of Cambridge, UK) – Morphogenetic mechanisms in early Drosophila embryos  
• Richard SMITH (John Innes Centre, UK) –  MorphoRobotX: A versatile platform for experimental biophysics 
• Yusuke TOYAMA (Mechanobiology Institute, Singapore) –  Mechanical impact of apoptosis in tissue homeostasis       
• Miltos TSIANTIS (Max Planck Institute for plant breeding research, Germany) – The genetic basis for leaf development and diversity: from understanding to reconstructing    
• Virgile VIASNOFF  (CNRS, Singapore) –   From sphere to tube, the early life of bile canaliculi 
• Pamela J.HINES, Invited journalist, Science – AAAS (American Association for the Advancement of Science), USA

 

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