Biomechanical signaling within the developing zebrafish heart attunes endocardial growth to myocardial chamber dimensions

Intra-organ communication guides morphogenetic processes that are essential for an organ to carry out complex physiological functions. In the heart, the growth of the myocardium is tightly coupled to that of the endocardium, a specialized endothelial tissue that lines its interior. Several molecular pathways have been implicated in the communication between these tissues including secreted factors, components of the extracellular matrix, or proteins involved in cell-cell communication. Yet, it is unknown how the growth of the endocardium is coordinated with that of the myocardium. Here, we show that an increased expansion of the myocardial atrial chamber volume generates higher junctional forces within endocardial cells. This leads to biomechanical signaling involving VE-cadherin, triggering nuclear localization of the Hippo pathway transcriptional regulator Yap1 and endocardial proliferation. Our work suggests that the growth of the endocardium results from myocardial chamber volume expansion and ends when the tension on the tissue is relaxed

Gastruloids - a minimalistic model to study complex developmental metabolism.

Metabolic networks are well placed to orchestrate the coordination of multiple cellular processes associated with embryonic development such as cell growth, proliferation, differentiation and cell movement. Here, we discuss the advantages that gastruloids, aggregates of mammalian embryonic stem cells that self-assemble a rudimentary body plan, have for uncovering the instructive role of metabolic pathways play in directing developmental processes. We emphasise the importance of using such reductionist systems to link specific pathways to defined events of early mammalian development and their utility for obtaining enough material for metabolomic studies. Finally, we review the ways in which the basic gastruloid protocol can be adapted to obtain specific models of embryonic cell types, tissues and regions. Together, we propose that gastruloids are an ideal system to rapidly uncover new mechanistic links between developmental signalling pathways and metabolic networks, which can then inform precise in vivo studies to confirm their function in the embryo

Gastruloids — a minimalistic model to study complex developmental metabolism

Peer reviewed: TrueAcknowledgements: For the purpose of open access, the author has applied a Creative Commons Attribution (CC BY) licence to any Author Accepted Manuscript version arising from this submission.Publication status: PublishedMetabolic networks are well placed to orchestrate the coordination of multiple cellular processes associated with embryonic development such as cell growth, proliferation, differentiation and cell movement. Here, we discuss the advantages that gastruloids, aggregates of mammalian embryonic stem cells that self-assemble a rudimentary body plan, have for uncovering the instructive role of metabolic pathways play in directing developmental processes. We emphasise the importance of using such reductionist systems to link specific pathways to defined events of early mammalian development and their utility for obtaining enough material for metabolomic studies. Finally, we review the ways in which the basic gastruloid protocol can be adapted to obtain specific models of embryonic cell types, tissues and regions. Together, we propose that gastruloids are an ideal system to rapidly uncover new mechanistic links between developmental signalling pathways and metabolic networks, which can then inform precise in vivo studies to confirm their function in the embryo.</jats:p

Axis Specification in Zebrafish Is Robust to Cell Mixing and Reveals a Regulation of Pattern Formation by Morphogenesis.

A fundamental question in developmental biology is how the early embryo establishes the spatial coordinate system that is later important for the organization of the embryonic body plan. Although we know a lot about the signaling and gene-regulatory networks required for this process, much less is understood about how these can operate to pattern tissues in the context of the extensive cell movements that drive gastrulation. In zebrafish, germ layer specification depends on the inheritance of maternal mRNAs [1-3], cortical rotation to generate a dorsal pole of β-catenin activity [4-8], and the release of Nodal signals from the yolk syncytial layer (YSL) [9-12]. To determine whether germ layer specification is robust to altered cell-to-cell positioning, we separated embryonic cells from the yolk and allowed them to develop as spherical aggregates. These aggregates break symmetry autonomously to form elongated structures with an anterior-posterior pattern. Both forced reaggregation and endogenous cell mixing reveals how robust early axis specification is to spatial disruption of maternal pre-patterning. During these movements, a pole of Nodal signaling emerges that is required for explant elongation via the planar cell polarity (PCP) pathway. Blocking of PCP-dependent elongation disrupts the shaping of opposing poles of BMP and Wnt/TCF activity and the anterior-posterior patterning of neural tissue. These results lead us to suggest that embryo elongation plays a causal role in timing the exposure of cells to changes in BMP and Wnt signal activity during zebrafish gastrulation. VIDEO ABSTRACT

Intraflagellar Transport Complex B Proteins Regulate the Hippo Effector Yap1 during Cardiogenesis.

Cilia and the intraflagellar transport (IFT) proteins involved in ciliogenesis are associated with congenital heart diseases (CHDs). However, the molecular links between cilia, IFT proteins, and cardiogenesis are yet to be established. Using a combination of biochemistry, genetics, and live-imaging methods, we show that IFT complex B proteins (Ift88, Ift54, and Ift20) modulate the Hippo pathway effector YAP1 in zebrafish and mouse. We demonstrate that this interaction is key to restrict the formation of the proepicardium and the myocardium. In cellulo experiments suggest that IFT88 and IFT20 interact with YAP1 in the cytoplasm and functionally modulate its activity, identifying a molecular link between cilia-related proteins and the Hippo pathway. Taken together, our results highlight a noncanonical role for IFT complex B proteins during cardiogenesis and shed light on a mechanism of action for ciliary proteins in YAP1 regulation.This project has received funding from the European Union’s Horizon 2020 Research and Innovation Programme under the Marie Sklodowska-Curie Grant Agreement No. 708312 (M.P.) and from the European Research Council (ERC) under the European Union’s Horizon 2020 Research and Innovation Programme: GA No. 682938 (J.V.). This work was supported by FRM (DEQ20140329553), by ANR (ANR-15-CE13-0015–liveheart, ANR- SNF310030E-164245-forcinregeneration), and by the Grant ANR-10-LABX-0030-INRT, a French State fund managed by the Agence Nationale de la Recherche under the frame program Investissements d’Avenir labeled ANR-10-IDEX-0002-02. B.D.’s team was supported by ANR-12-CHEX-005 and CNRS. S.M.M.’s team was supported by core funding from the Institut Imagine, Institut Pasteur, Inserm, Universite´ Paris Descartes, and a grant from the AFM-Te´ le´ thon Trampoline 18727). T.L. was funded by the ED515 (1691/2014). L.O.L. is supported by the European Commission (H2020-MSCA-ITN-2016 European Industrial Doctorate 4DHeart 722427).S

Selective utilization of glucose metabolism guides mammalian gastrulation.

Acknowledgements: The authors thank K. Sumigray for support with confocal microscopy; Z. Smith and Smith laboratory members for support with microinjection and preparation of pseudopregnant mice for tetraploid complementation assays; Yale Genome Editing Center for support in tetraploid complementation experiments; T. Finkelstein for support with mouse experiments and illustrations shown in Extended Data Fig. 1c; members of the Sozen laboratory for discussions throughout the project; and Z. Smith for helpful feedback on the manuscript. J.B. is supported by the NIH NRSA F31 pre-doctoral fellowship. A.H. is a NYSCF-Druckenmiller Fellow and is supported by The New York Stem Cell Foundation. V.G. is supported by NIH grants number 1R01AR063663, 1R01AR067755 and DP1AG066590. B. Steventon is supported by a Wellcome Trust Discovery Award (225360_Z_22_Z). Work in the Steventon laboratory was funded by MRC Research grant MR/ V009192/1 jointly held by C.D. and B. Steventon. The Sozen laboratory is funded by NIH Early Innovators Award (DP2HD112040), the Richard and Susan Smith Family Foundation, Reprogrants and American Society of Reproductive Medicine (ASRM). The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.The prevailing dogma for morphological patterning in developing organisms argues that the combined inputs of transcription factor networks and signalling morphogens alone generate spatially and temporally distinct expression patterns. However, metabolism has also emerged as a critical developmental regulator1-10, independent of its functions in energy production and growth. The mechanistic role of nutrient utilization in instructing cellular programmes to shape the in vivo developing mammalian embryo remains unknown. Here we reveal two spatially resolved, cell-type- and stage-specific waves of glucose metabolism during mammalian gastrulation by using single-cell-resolution quantitative imaging of developing mouse embryos, stem cell models and embryo-derived tissue explants. We identify that the first spatiotemporal wave of glucose metabolism occurs through the hexosamine biosynthetic pathway to drive fate acquisition in the epiblast, and the second wave uses glycolysis to guide mesoderm migration and lateral expansion. Furthermore, we demonstrate that glucose exerts its influence on these developmental processes through cellular signalling pathways, with distinct mechanisms connecting glucose with the ERK activity in each wave. Our findings underscore that-in synergy with genetic mechanisms and morphogenic gradients-compartmentalized cellular metabolism is integral in guiding cell fate and specialized functions during development. This study challenges the view of the generic and housekeeping nature of cellular metabolism, offering valuable insights into its roles in various developmental contexts

Selective utilization of glucose metabolism guides mammalian gastrulation.

Acknowledgements: The authors thank K. Sumigray for support with confocal microscopy; Z. Smith and Smith laboratory members for support with microinjection and preparation of pseudopregnant mice for tetraploid complementation assays; Yale Genome Editing Center for support in tetraploid complementation experiments; T. Finkelstein for support with mouse experiments and illustrations shown in Extended Data Fig. 1c; members of the Sozen laboratory for discussions throughout the project; and Z. Smith for helpful feedback on the manuscript. J.B. is supported by the NIH NRSA F31 pre-doctoral fellowship. A.H. is a NYSCF-Druckenmiller Fellow and is supported by The New York Stem Cell Foundation. V.G. is supported by NIH grants number 1R01AR063663, 1R01AR067755 and DP1AG066590. B. Steventon is supported by a Wellcome Trust Discovery Award (225360_Z_22_Z). Work in the Steventon laboratory was funded by MRC Research grant MR/ V009192/1 jointly held by C.D. and B. Steventon. The Sozen laboratory is funded by NIH Early Innovators Award (DP2HD112040), the Richard and Susan Smith Family Foundation, Reprogrants and American Society of Reproductive Medicine (ASRM). The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.The prevailing dogma for morphological patterning in developing organisms argues that the combined inputs of transcription factor networks and signalling morphogens alone generate spatially and temporally distinct expression patterns. However, metabolism has also emerged as a critical developmental regulator1-10, independent of its functions in energy production and growth. The mechanistic role of nutrient utilization in instructing cellular programmes to shape the in vivo developing mammalian embryo remains unknown. Here we reveal two spatially resolved, cell-type- and stage-specific waves of glucose metabolism during mammalian gastrulation by using single-cell-resolution quantitative imaging of developing mouse embryos, stem cell models and embryo-derived tissue explants. We identify that the first spatiotemporal wave of glucose metabolism occurs through the hexosamine biosynthetic pathway to drive fate acquisition in the epiblast, and the second wave uses glycolysis to guide mesoderm migration and lateral expansion. Furthermore, we demonstrate that glucose exerts its influence on these developmental processes through cellular signalling pathways, with distinct mechanisms connecting glucose with the ERK activity in each wave. Our findings underscore that-in synergy with genetic mechanisms and morphogenic gradients-compartmentalized cellular metabolism is integral in guiding cell fate and specialized functions during development. This study challenges the view of the generic and housekeeping nature of cellular metabolism, offering valuable insights into its roles in various developmental contexts

Yap/Taz-TEAD activity links mechanical cues to progenitor cell behavior during zebrafish hindbrain segmentation

Cells perceive their microenvironment through chemical and physical cues. However, how the mechanical signals are interpreted during embryonic tissue deformation to result in specific cell behaviors is largely unknown. The Yap/Taz family of transcriptional co-activators has emerged as an important regulator of tissue growth and regeneration, responding to physical cues from the extracellular matrix, and to cell shape and actomyosin cytoskeletal changes. In this study, we demonstrate the role of Yap/Taz-TEAD activity as a sensor of mechanical signals in the regulation of the progenitor behavior of boundary cells during zebrafish hindbrain compartmentalization. Monitoring of in vivo Yap/Taz activity during hindbrain segmentation indicated that boundary cells responded to mechanical cues in a cell-autonomous manner through Yap/Taz-TEAD activity. Cell-lineage analysis revealed that Yap/Taz-TEAD boundary cells decreased their proliferative activity when Yap/Taz-TEAD activity ceased, which preceded changes in their cell fate from proliferating progenitors to differentiated neurons. Functional experiments demonstrated the pivotal role of Yap/Taz-TEAD signaling in maintaining progenitor features in the hindbrain boundary cell population.This work was supported by a La Marató-TV3 grant (345/C/2014) and by a Ministerio de Ciencia, Innovación y Universidades grant (BFU2015-67400-P and BFU2016-81887-REDT/AEI) to C.P.; and by a Unidad de Excelencia María de Maetzu grant (MDM-2014-0370) to the Department of Experimental and Health Sciences of the Pompeu Fabra University (DCEXS-UPF). A.V. was a recipient of a predoctoral fellowship from the Fundació La Caixa, and C.E.-P. holds a predoctoral fellowship from the Ministerio de Ciencia, Innovación y Universidades (FPU). J.T. was a recipient of a postdoctoral Beatriu de Pinos fellowship (AGAUR, Generalitat de Catalunya). C.P. is recipient of an Institució Catalana per la Recerca i Estudis Avançats Academia award (Generalitat de Catalunya)