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Force transmission through adherens junctions (AJs) is crucial for multicellular organization, wound healing and tissue regeneration. Recent studies shed light on the molecular mechanisms of mechanotransduction at the AJs. However, the canonical model fails to explain force transmission when essential proteins of the mechanotransduction module are mutated or missing. Here, we demonstrate that, in absence of α-catenin, β-catenin can directly and functionally interact with vinculin in its open conformation, bearing physiological forces. Furthermore, we found that β-catenin can prevent vinculin autoinhibition in the presence of α-catenin by occupying vinculin´s head-tail interaction site, thus preserving force transmission capability. Taken together, our findings suggest a multi-step force transmission process at AJs, where α-catenin and β-catenin can alternatively and cooperatively interact with vinculin. This can explain the graded responses needed to maintain tissue mechanical homeostasis and, importantly, unveils a force-bearing mechanism involving β-catenin and extended vinculin that can potentially explain the underlying process enabling collective invasion of metastatic cells lacking α-catenin. Adherens junctions mediate force transmission and connect the cytoskeleton of adjacent cells. This study demonstrates that force transmission can be achieved through direct and functional β-catenin/vinculin interaction in the absence of α-catenin.

Motivation Quantitative descriptions of multi-cellular structures from optical microscopy imaging are prime to understand the variety of three-dimensional (3D) shapes in living organisms. Experimental models of vertebrates, invertebrates and plants, such as zebrafish, killifish, Drosophila or Marchantia , mainly comprise multilayer tissues, and even if microscopes can reach the needed depth, their geometry hinders the selection and subsequent analysis of the optical volumes of interest. Computational tools to “peel” tissues by removing specific layers and reducing 3D volume into planar images, can critically improve visualization and analysis. Results We developed VolumePeeler, a versatile FIJI plugin for virtual 3D “peeling” of image stacks. The plugin implements spherical and spline surface projections. We applied VolumePeeler to perform peeling in 3D images of spherical embryos, as well as non-spherical tissue layers. The produced images improve the 3D volume visualization and enable analysis and quantification of geometrically challenging microscopy datasets. Availability ImageJ/FIJI software, source code, examples, and tutorials are openly available in https://cimt.uchile.cl/mcerda

Cells are exposed and respond to various mechanical forces and physical cues stemming from their environment. This interaction has been seen to differentially regulate various cellular processes for maintenance of homeostasis, of which autophagy represents one of the major players. In addition, autophagy has been suggested to regulate mechanical functions of the cells including their interaction with the environment. In this minireview, we summarize the state of the art of the fascinating interplay between autophagy and the mechanotransduction machinery associated with cell adhesions, that we name ¨Mechanoautophagy¨

The advent of micro and nanotechnologies, such as microfabrication, have impacted scientific research and contributed to meaningful real-world applications, to a degree seen during historic technological revolutions. Some key areas benefitting from the invention and advancement of microfabrication platforms are those of biological and biomedical sciences. Modern therapeutic approaches, involving point-of-care, precision or personalized medicine, are transitioning from the experimental phase to becoming the standard of care. At the same time, biological research benefits from the contribution of microfluidics at every level from single cell to tissue engineering and organoids studies. The aim of this commentary is to describe, through proven examples, the interdisciplinary process used to develop novel biological technologies and to emphasize the role of technical knowledge in empowering researchers who are specialized in a niche area to look beyond and innovate.

Palmitic acid (PA) is significantly increased in the hypothalamus of mice, when fed chronically with a high-fat diet (HFD). PA impairs insulin signaling in hypothalamic neurons, by a mechanism dependent on autophagy, a process of lysosomal-mediated degradation of cytoplasmic material. In addition, previous work shows a crosstalk between autophagy and the primary cilium (hereafter cilium), an antenna-like structure on the cell surface that acts as a signaling platform for the cell. Ciliopathies, human diseases characterized by cilia dysfunction, manifest, type 2 diabetes, among other features, suggesting a role of the cilium in insulin signaling. Cilium depletion in hypothalamic pro-opiomelanocortin (POMC) neurons triggers obesity and insulin resistance in mice, the same phenotype as mice deficient in autophagy in POMC neurons. Here we investigated the effect of chronic consumption of HFD on cilia; and our results indicate that chronic feeding with HFD reduces the percentage of cilia in hypothalamic POMC neurons. This effect may be due to an increased amount of PA, as treatment with this saturated fatty acid in vitro reduces the percentage of ciliated cells and cilia length in hypothalamic neurons. Importantly, the same effect of cilia depletion was obtained following chemical and genetic inhibition of autophagy, indicating autophagy is required for ciliogenesis. We further demonstrate a role for the cilium in insulin sensitivity, as cilium loss in hypothalamic neuronal cells disrupts insulin signaling and insulin-dependent glucose uptake, an effect that correlates with the ciliary localization of the insulin receptor (IR). Consistently, increased percentage of ciliated hypothalamic neuronal cells promotes insulin signaling, even when cells are exposed to PA. Altogether, our results indicate that, in hypothalamic neurons, impairment of autophagy, either by PA exposure, chemical or genetic manipulation, cause cilia loss that impairs insulin sensitivity.

Cancer vasculogenic mimicry (VM) is the formation of vasculature structures in the absence of endothelial cells. We previously established an in vitro model that facilitates the formation of a lumen-containing and fluid-conducting tubular structures after 4 days of cancer cell growth on Matrigel. Herein, we mechanistically characterize this model in breast and ovarian cancer cell lines demonstrating distinct phases of VM formation and the dependence of specific extracellular matrix proteins. We report that VM occurs in four distinct stages. Firstly, alignment, migration then clustering delineate the area of the future tubular structure. Secondly, contraction of aligned structures followed by loss of attachment of some cells and cellular blebbing. Thirdly, a phase of mass proliferation followed by the raising of specific areas of the cancer cell mass above the Matrigel (bridge). Finally, the formation of a cell monolayer closes the tubular structure, forms a glycoprotein-rich luminal lining, then elevates the structure. Only later stages of VM require AKT and FAK signaling, as confirmed by chemical inhibition and phosphorylation analysis. We demonstrate that the lining of the tubular lumen is rich in laminin. Furthermore, the presence of Laminin 111 (but not collagen I) is sufficient in the extracellular matrix (Matrigel) for VM to occur and we confirm that integrin β1, but not integrin β3, is required and this protein changes location during the formation process. RNASeq analysis suggests that VM formation principally occurs through post-transcriptional regulation. As VM is associated with poor patient survival VM, an understanding of the mechanism of VM may bring to light novel biomarkers and anticancer targets.

Cadherin-mediated adhesions (also known as adherens junctions) are adhesive complexes that connect neighboring cells in a tissue. While the role of the actin cytoskeleton in withstanding tension at these sites of contact is well documented, little is known about the involvement of microtubules and the associated endoplasmic reticulum (ER) network in cadherin mechanotransduction. Therefore, we investigated how the organization of ER extensions in close proximity of cadherin-mediated adhesions can affect such complexes, and vice versa. Here, we show that the extension of the ER to cadherin-mediated adhesions is tension dependent and appears to be cadherin-type specific. Furthermore, the different structural organization of the ER/microtubule network seems to affect the localization of ER-bound PTP1B at cadherin-mediated adhesions. This phosphatase is involved in the modulation of vinculin, a molecular clutch which enables differential engagement of the cadherin-catenin layer with the actomyosin cytoskeleton in response to tension. This suggests a link between structural organization of the ER/microtubule network around cadherin-specific adhesions, to control the mechanotransduction of adherens junctions by modulation of vinculin conformational state.

Introduction: Deciphering the biological and physical requirements for the outset of multicellularity is limited to few experimental models. The early embryonic development of annual killifish represents an almost unique opportunity to investigate de novo cellular aggregation in a vertebrate model. As an adaptation to seasonal drought, annual killifish employs a unique developmental pattern in which embryogenesis occurs only after undifferentiated embryonic cells have completed epiboly and dispersed in low density on the egg surface. Therefore, the first stage of embryogenesis requires the congregation of embryonic cells at one pole of the egg to form a single aggregate that later gives rise to the embryo proper. This unique process presents an opportunity to dissect the self-organizing principles involved in early organization of embryonic stem cells. Indeed, the physical and biological processes required to form the aggregate of embryonic cells are currently unknown. Methods: Here, we developed an in silico , agent-based biophysical model that allows testing how cell-specific and environmental properties could determine the aggregation dynamics of early Killifish embryogenesis. In a forward engineering approach, we then proceeded to test two hypotheses for cell aggregation (cell-autonomous and a simple taxis model) as a proof of concept of modeling feasibility. In a first approach (cell autonomous system), we considered how intrinsic biophysical properties of the cells such as motility, polarity, density, and the interplay between cell adhesion and contact inhibition of locomotion drive cell aggregation into self-organized clusters. Second, we included guidance of cell migration through a simple taxis mechanism to resemble the activity of an organizing center found in several developmental models. Results: Our numerical simulations showed that random migration combined with low cell-cell adhesion is sufficient to maintain cells in dispersion and that aggregation can indeed arise spontaneously under a limited set of conditions, but, without environmental guidance, the dynamics and resulting structures do not recapitulate in vivo observations. Discussion: Thus, an environmental guidance cue seems to be required for correct execution of early aggregation in early killifish development. However, the nature of this cue (e.g., chemical or mechanical) can only be determined experimentally. Our model provides a predictive tool that could be used to better characterize the process and, importantly, to design informed experimental strategies.

Multicellularity in animals requires dynamic maintenance of cell–cell contacts. Intercellularly ligated cadherins recruit numerous proteins to form supramolecular complexes that connect with the actin cytoskeleton and support force transmission. However, the molecular organization within such structures remains unknown. Here we mapped protein organization in cadherin-based adhesions by super-resolution microscopy, revealing a multi-compartment nanoscale architecture, with the plasma-membrane-proximal cadherin–catenin compartment segregated from the actin cytoskeletal compartment, bridged by an interface zone containing vinculin. Vinculin position is determined by α-catenin, and following activation, vinculin can extend ∼30 nm to bridge the cadherin–catenin and actin compartments, while modulating the nanoscale positions of the actin regulators zyxin and VASP. Vinculin conformational activation requires tension and tyrosine phosphorylation, regulated by Abl kinase and PTP1B phosphatase. Such modular architecture provides a structural framework for mechanical and biochemical signal integration by vinculin, which may differentially engage cadherin–catenin complexes with the actomyosin machinery to regulate cell adhesions. Bertocchi and colleagues describe the organization of cadherin-based adhesions using super-resolution microscopy. They find that α-catenin is important for vinculin localization and observe a conformational change in vinculin following its activation.

Adherens junctions connect the actin cytoskeleton of neighboring cells through transmembrane cadherin receptors and a network of adaptor proteins. The interactions between these adaptors and cadherin as well as the activity of actin regulators localized to adherens junctions are tightly controlled to facilitate cell junction assembly or disassembly in response to changes in external or internal forces and/or signaling. Phosphorylation of tyrosine, serine, or threonine residues acts as a switch on the majority of adherens junction proteins, turning “on” or “off” their interactions with other proteins and/or their enzymatic activity. Here, we provide an overview of the kinases and phosphatases regulating phosphorylation of adherens junction proteins and bring examples of phosphorylation events leading to the assembly or disassembly of adherens junctions, highlighting the important role of phosphorylation switches in regulating their dynamics.