@phdthesis{9623, abstract = {Cytoplasmic reorganizations are essential for morphogenesis. In large cells like oocytes, these reorganizations become crucial in patterning the oocyte for later stages of embryonic development. Ascidians oocytes reorganize their cytoplasm (ooplasm) in a spectacular manner. Ooplasmic reorganization is initiated at fertilization with the contraction of the actomyosin cortex along the animal-vegetal axis of the oocyte, driving the accumulation of cortical endoplasmic reticulum (cER), maternal mRNAs associated to it and a mitochondria-rich subcortical layer – the myoplasm – in a region of the vegetal pole termed contraction pole (CP). Here we have used the species Phallusia mammillata to investigate the changes in cell shape that accompany these reorganizations and the mechanochemical mechanisms underlining CP formation. We report that the length of the animal-vegetal (AV) axis oscillates upon fertilization: it first undergoes a cycle of fast elongation-lengthening followed by a slow expansion of mainly the vegetal pole (VP) of the cell. We show that the fast oscillation corresponds to a dynamic polarization of the actin cortex as a result of a fertilization-induced increase in cortical tension in the oocyte that triggers a rupture of the cortex at the animal pole and the establishment of vegetal-directed cortical flows. These flows are responsible for the vegetal accumulation of actin causing the VP to flatten. We find that the slow expansion of the VP, leading to CP formation, correlates with a relaxation of the vegetal cortex and that the myoplasm plays a role in the expansion. We show that the myoplasm is a solid-like layer that buckles under compression forces arising from the contracting actin cortex at the VP. Straightening of the myoplasm when actin flows stops, facilitates the expansion of the VP and the CP. Altogether, our results present a previously unrecognized role for the myoplasm in ascidian ooplasmic segregation. }, author = {Caballero Mancebo, Silvia}, isbn = {978-3-99078-012-1}, issn = {2663-337X}, pages = {111}, publisher = {Institute of Science and Technology Austria}, title = {{Fertilization-induced deformations are controlled by the actin cortex and a mitochondria-rich subcortical layer in ascidian oocytes}}, doi = {10.15479/at:ista:9623}, year = {2021}, } @article{9006, abstract = {Cytoplasm is a gel-like crowded environment composed of various macromolecules, organelles, cytoskeletal networks, and cytosol. The structure of the cytoplasm is highly organized and heterogeneous due to the crowding of its constituents and their effective compartmentalization. In such an environment, the diffusive dynamics of the molecules are restricted, an effect that is further amplified by clustering and anchoring of molecules. Despite the crowded nature of the cytoplasm at the microscopic scale, large-scale reorganization of the cytoplasm is essential for important cellular functions, such as cell division and polarization. How such mesoscale reorganization of the cytoplasm is achieved, especially for large cells such as oocytes or syncytial tissues that can span hundreds of micrometers in size, is only beginning to be understood. In this review, we will discuss recent advances in elucidating the molecular, cellular, and biophysical mechanisms by which the cytoskeleton drives cytoplasmic reorganization across different scales, structures, and species.}, author = {Shamipour, Shayan and Caballero Mancebo, Silvia and Heisenberg, Carl-Philipp J}, issn = {18781551}, journal = {Developmental Cell}, number = {2}, pages = {P213--226}, publisher = {Elsevier}, title = {{Cytoplasm's got moves}}, doi = {10.1016/j.devcel.2020.12.002}, volume = {56}, year = {2021}, } @phdthesis{9397, abstract = {Accumulation of interstitial fluid (IF) between embryonic cells is a common phenomenon in vertebrate embryogenesis. Unlike other model systems, where these accumulations coalesce into a large central cavity – the blastocoel, in zebrafish, IF is more uniformly distributed between the deep cells (DC) before the onset of gastrulation. This is likely due to the presence of a large extraembryonic structure – the yolk cell (YC) at the position where the blastocoel typically forms in other model organisms. IF has long been speculated to play a role in tissue morphogenesis during embryogenesis, but direct evidence supporting such function is still sparse. Here we show that the relocalization of IF to the interface between the YC and DC/epiblast is critical for axial mesendoderm (ME) cell protrusion formation and migration along this interface, a key process in embryonic axis formation. We further demonstrate that axial ME cell migration and IF relocalization engage in a positive feedback loop, where axial ME migration triggers IF accumulation ahead of the advancing axial ME tissue by mechanically compressing the overlying epiblast cell layer. Upon compression, locally induced flow relocalizes the IF through the porous epiblast tissue resulting in an IF accumulation ahead of the leading axial ME. This IF accumulation, in turn, promotes cell protrusion formation and migration of the leading axial ME cells, thereby facilitating axial ME extension. Our findings reveal a central role of dynamic IF relocalization in orchestrating germ layer morphogenesis during gastrulation.}, author = {Huljev, Karla}, issn = {2663-337X}, pages = {101}, publisher = {Institute of Science and Technology Austria}, title = {{Coordinated spatiotemporal reorganization of interstitial fluid is required for axial mesendoderm migration in zebrafish gastrulation}}, doi = {10.15479/at:ista:9397}, year = {2021}, } @article{7888, abstract = {Embryonic stem cell cultures are thought to self-organize into embryoid bodies, able to undergo symmetry-breaking, germ layer specification and even morphogenesis. Yet, it is unclear how to reconcile this remarkable self-organization capacity with classical experiments demonstrating key roles for extrinsic biases by maternal factors and/or extraembryonic tissues in embryogenesis. Here, we show that zebrafish embryonic tissue explants, prepared prior to germ layer induction and lacking extraembryonic tissues, can specify all germ layers and form a seemingly complete mesendoderm anlage. Importantly, explant organization requires polarized inheritance of maternal factors from dorsal-marginal regions of the blastoderm. Moreover, induction of endoderm and head-mesoderm, which require peak Nodal-signaling levels, is highly variable in explants, reminiscent of embryos with reduced Nodal signals from the extraembryonic tissues. Together, these data suggest that zebrafish explants do not undergo bona fide self-organization, but rather display features of genetically encoded self-assembly, where intrinsic genetic programs control the emergence of order.}, author = {Schauer, Alexandra and Nunes Pinheiro, Diana C and Hauschild, Robert and Heisenberg, Carl-Philipp J}, issn = {2050-084X}, journal = {eLife}, publisher = {eLife Sciences Publications}, title = {{Zebrafish embryonic explants undergo genetically encoded self-assembly}}, doi = {10.7554/elife.55190}, volume = {9}, year = {2020}, } @article{8680, abstract = {Animal development entails the organization of specific cell types in space and time, and spatial patterns must form in a robust manner. In the zebrafish spinal cord, neural progenitors form stereotypic patterns despite noisy morphogen signaling and large-scale cellular rearrangements during morphogenesis and growth. By directly measuring adhesion forces and preferences for three types of endogenous neural progenitors, we provide evidence for the differential adhesion model in which differences in intercellular adhesion mediate cell sorting. Cell type–specific combinatorial expression of different classes of cadherins (N-cadherin, cadherin 11, and protocadherin 19) results in homotypic preference ex vivo and patterning robustness in vivo. Furthermore, the differential adhesion code is regulated by the sonic hedgehog morphogen gradient. We propose that robust patterning during tissue morphogenesis results from interplay between adhesion-based self-organization and morphogen-directed patterning.}, author = {Tsai, Tony Y.-C. and Sikora, Mateusz K and Xia, Peng and Colak-Champollion, Tugba and Knaut, Holger and Heisenberg, Carl-Philipp J and Megason, Sean G.}, issn = {1095-9203}, journal = {Science}, keywords = {Multidisciplinary}, number = {6512}, pages = {113--116}, publisher = {American Association for the Advancement of Science}, title = {{An adhesion code ensures robust pattern formation during tissue morphogenesis}}, doi = {10.1126/science.aba6637}, volume = {370}, year = {2020}, } @article{8957, abstract = {Global tissue tension anisotropy has been shown to trigger stereotypical cell division orientation by elongating mitotic cells along the main tension axis. Yet, how tissue tension elongates mitotic cells despite those cells undergoing mitotic rounding (MR) by globally upregulating cortical actomyosin tension remains unclear. We addressed this question by taking advantage of ascidian embryos, consisting of a small number of interphasic and mitotic blastomeres and displaying an invariant division pattern. We found that blastomeres undergo MR by locally relaxing cortical tension at their apex, thereby allowing extrinsic pulling forces from neighboring interphasic blastomeres to polarize their shape and thus division orientation. Consistently, interfering with extrinsic forces by reducing the contractility of interphasic blastomeres or disrupting the establishment of asynchronous mitotic domains leads to aberrant mitotic cell division orientations. Thus, apical relaxation during MR constitutes a key mechanism by which tissue tension anisotropy controls stereotypical cell division orientation.}, author = {Godard, Benoit G and Dumollard, Rémi and Munro, Edwin and Chenevert, Janet and Hebras, Céline and Mcdougall, Alex and Heisenberg, Carl-Philipp J}, issn = {18781551}, journal = {Developmental Cell}, number = {6}, pages = {695--706}, publisher = {Elsevier}, title = {{Apical relaxation during mitotic rounding promotes tension-oriented cell division}}, doi = {10.1016/j.devcel.2020.10.016}, volume = {55}, year = {2020}, } @inbook{7227, abstract = {Gastrulation entails specification and formation of three embryonic germ layers—ectoderm, mesoderm and endoderm—thereby establishing the basis for the future body plan. In zebrafish embryos, germ layer specification occurs during blastula and early gastrula stages (Ho & Kimmel, 1993), a period when the main morphogenetic movements underlying gastrulation are initiated. Hence, the signals driving progenitor cell fate specification, such as Nodal ligands from the TGF-β family, also play key roles in regulating germ layer progenitor cell segregation (Carmany-Rampey & Schier, 2001; David & Rosa, 2001; Feldman et al., 2000; Gritsman et al., 1999; Keller et al., 2008). In this review, we summarize and discuss the main signaling pathways involved in germ layer progenitor cell fate specification and segregation, specifically focusing on recent advances in understanding the interplay between mesoderm and endoderm specification and the internalization movements at the onset of zebrafish gastrulation.}, author = {Nunes Pinheiro, Diana C and Heisenberg, Carl-Philipp J}, booktitle = {Gastrulation: From Embryonic Pattern to Form}, issn = {00702153}, pages = {343--375}, publisher = {Elsevier}, title = {{Zebrafish gastrulation: Putting fate in motion}}, doi = {10.1016/bs.ctdb.2019.10.009}, volume = {136}, year = {2020}, } @inbook{7410, abstract = {Epiboly is a conserved gastrulation movement describing the thinning and spreading of a sheet or multi-layer of cells. The zebrafish embryo has emerged as a vital model system to address the cellular and molecular mechanisms that drive epiboly. In the zebrafish embryo, the blastoderm, consisting of a simple squamous epithelium (the enveloping layer) and an underlying mass of deep cells, as well as a yolk nuclear syncytium (the yolk syncytial layer) undergo epiboly to internalize the yolk cell during gastrulation. The major events during zebrafish epiboly are: expansion of the enveloping layer and the internal yolk syncytial layer, reduction and removal of the yolk membrane ahead of the advancing blastoderm margin and deep cell rearrangements between the enveloping layer and yolk syncytial layer to thin the blastoderm. Here, work addressing the cellular and molecular mechanisms as well as the sources of the mechanical forces that underlie these events is reviewed. The contribution of recent findings to the current model of epiboly as well as open questions and future prospects are also discussed.}, author = {Bruce, Ashley E.E. and Heisenberg, Carl-Philipp J}, booktitle = {Gastrulation: From Embryonic Pattern to Form}, editor = {Solnica-Krezel, Lilianna }, isbn = {9780128127988}, issn = {0070-2153}, pages = {319--341}, publisher = {Elsevier}, title = {{Mechanisms of zebrafish epiboly: A current view}}, doi = {10.1016/bs.ctdb.2019.07.001}, volume = {136}, year = {2020}, } @unpublished{9750, abstract = {Tension of the actomyosin cell cortex plays a key role in determining cell-cell contact growth and size. The level of cortical tension outside of the cell-cell contact, when pulling at the contact edge, scales with the total size to which a cell-cell contact can grow1,2. Here we show in zebrafish primary germ layer progenitor cells that this monotonic relationship only applies to a narrow range of cortical tension increase, and that above a critical threshold, contact size inversely scales with cortical tension. This switch from cortical tension increasing to decreasing progenitor cell-cell contact size is caused by cortical tension promoting E-cadherin anchoring to the actomyosin cytoskeleton, thereby increasing clustering and stability of E-cadherin at the contact. Once tension-mediated E-cadherin stabilization at the contact exceeds a critical threshold level, the rate by which the contact expands in response to pulling forces from the cortex sharply drops, leading to smaller contacts at physiologically relevant timescales of contact formation. Thus, the activity of cortical tension in expanding cell-cell contact size is limited by tension stabilizing E-cadherin-actin complexes at the contact.}, author = {Slovakova, Jana and Sikora, Mateusz K and Caballero Mancebo, Silvia and Krens, Gabriel and Kaufmann, Walter and Huljev, Karla and Heisenberg, Carl-Philipp J}, booktitle = {bioRxiv}, pages = {41}, publisher = {Cold Spring Harbor Laboratory}, title = {{Tension-dependent stabilization of E-cadherin limits cell-cell contact expansion}}, doi = {10.1101/2020.11.20.391284}, year = {2020}, } @phdthesis{8350, abstract = {Cytoplasm is a gel-like crowded environment composed of tens of thousands of macromolecules, organelles, cytoskeletal networks and cytosol. The structure of the cytoplasm is thought to be highly organized and heterogeneous due to the crowding of its constituents and their effective compartmentalization. In such an environment, the diffusive dynamics of the molecules is very restricted, an effect that is further amplified by clustering and anchoring of molecules. Despite the jammed nature of the cytoplasm at the microscopic scale, large-scale reorganization of cytoplasm is essential for important cellular functions, such as nuclear positioning and cell division. How such mesoscale reorganization of the cytoplasm is achieved, especially for very large cells such as oocytes or syncytial tissues that can span hundreds of micrometers in size, has only begun to be understood. In this thesis, I focus on the recent advances in elucidating the molecular, cellular and biophysical principles underlying cytoplasmic organization across different scales, structures and species. First, I outline which of these principles have been identified by reductionist approaches, such as in vitro reconstitution assays, where boundary conditions and components can be modulated at ease. I then describe how the theoretical and experimental framework established in these reduced systems have been applied to their more complex in vivo counterparts, in particular oocytes and embryonic syncytial structures, and discuss how such complex biological systems can initiate symmetry breaking and establish patterning. Specifically, I examine an example of large-scale reorganizations taking place in zebrafish embryos, where extensive cytoplasmic streaming leads to the segregation of cytoplasm from yolk granules along the animal-vegetal axis of the embryo. Using biophysical experimentation and theory, I investigate the forces underlying this process, to show that this process does not rely on cortical actin reorganization, as previously thought, but instead on a cell-cycle-dependent bulk actin polymerization wave traveling from the animal to the vegetal pole of the embryo. This wave functions in segregation by both pulling cytoplasm animally and pushing yolk granules vegetally. Cytoplasm pulling is mediated by bulk actin network flows exerting friction forces on the cytoplasm, while yolk granule pushing is achieved by a mechanism closely resembling actin comet formation on yolk granules. This study defines a novel role of bulk actin polymerization waves in embryo polarization via cytoplasmic segregation. Lastly, I describe the cytoplasmic reorganizations taking place during zebrafish oocyte maturation, where the initial segregation of the cytoplasm and yolk granules occurs. Here, I demonstrate a previously uncharacterized wave of microtubule aster formation, traveling the oocyte along the animal-vegetal axis. Further research is required to determine the role of such microtubule structures in cytoplasmic reorganizations therein. Collectively, these studies provide further evidence for the coupling between cell cytoskeleton and cell cycle machinery, which can underlie a core self-organizing mechanism for orchestrating large-scale reorganizations in a cell-cycle-tunable manner, where the modulations of the force-generating machinery and cytoplasmic mechanics can be harbored to fulfill cellular functions.}, author = {Shamipour, Shayan}, issn = {2663-337X}, pages = {107}, publisher = {Institute of Science and Technology Austria}, title = {{Bulk actin dynamics drive phase segregation in zebrafish oocytes }}, doi = {10.15479/AT:ISTA:8350}, year = {2020}, }