Cell Extrusion Research Articles

Specific mitotic events drive left-right organizer development.

Cell proliferation is crucial for tissue development. Here, we investigate its role in the Left-Right Organizer (LRO), which establishes the left-right (LR) axis. In zebrafish, we mapped mitotic events in Kupffer's Vesicle (KV) and identified an anteriorly enriched, FGF-dependent mitotic pattern. Laser ablation of mitotic cells and pericentrin-null mutants, both reducing mitotic events, resulted in smaller lumens, confirming that cell division is essential for KV development. Pericentrin-null mutants also exhibited defects in leftward cardiac jogging, indicative of KV dysfunction. Using a KV-specific fluorescent microtubule marker, we found that the KV rosette is a transient, centrally organized cluster interconnected by cytokinetic bridges and containing microtubule bundles. This structure emerges after the first four divisions and precedes lumen formation. Mitotic events during KV rounding coincide with rosette formation, spindle rotation, and cell extrusion, likely driven by increased packing. Eliminating the first four mitotic events disrupted rosette formation and prevented normal KV rounding. These findings demonstrate that mitotic events are critical for KV development, with cell division timing shaping KV architecture and function.

KRASG12D cells override homeostatic cell elimination mechanisms in adult pancreas via Wnt5a and cell dormancy.

PaperClip The adult pancreas protects against cancer by actively expelling genetically mutated cells. Pancreatic cancer starts with cells carrying KRAS mutations; however, it is not clear how some KRAS mutant cells override cell elimination mechanisms to survive in tissues. Methods We used an in vivo mouse model of sporadic tumorigenesis to induce Kras and/or Tp53 mutations in low numbers of cells in the adult pancreas. We monitored mutant cell fate over time using quantitative fluorescence imaging. Gene signatures of non-eliminated mutant cell populations were identified using bulk RNA sequencing. Differential gene expression was overlapped with publicly available datasets. Key molecular pathways were validated in murine pancreas using immunofluorescence and functionally tested using inhibitor studies in vivo and epithelial co-culture systems in vitro. Results While most genetically mutant cells are eliminated from the adult pancreas, a population of KRASG12D- or p53R172H-expressing cells are stably retained. We identify Wnt5a signalling, cell dormancy and stemness as key features of surviving KrasG12D cells in vivo. Wnt5a specifically inhibits apical extrusion of RasV12 cells by promoting stable E-cadherin-based cell-cell adhesions at RasV12: normal cell-cell boundaries in vitro. In the pancreas, Wnt signalling, E-cadherin and β-catenin are increased at cell-cell contacts between non-eliminated KrasG12D cells and normal neighbours. Active Wnt signalling is a general mechanism required to promote KrasG12D and p53R172H cell retention and cell survival in vivo. Conclusions RAS mutant cells activate Wnt5a and cell dormancy to avoid cell expulsion and to survive in the adult pancreas.

Cell membrane-derived nanovesicles as extracellular vesicle-mimetics in wound healing.

Cell membrane-derived nanovesicles as extracellular vesicle-mimetics in wound healing.

Microalgae-Derived Microparticles Improve Immunomodulation via Combined Glycolysis and MAPK Activation.

Natural polysaccharides possess potent immunostimulatory properties, but their poor solubility impedes efficiency of cellular delivery. This study focuses on extraction of microparticles (MPs) fromEuglena gracilis, a microalgae species characterized by abundant intracellular β-1,3-glucan and flexible cell membrane. We introduce anE. gracilis-derived MP (MPEG) system as a natural carrier for solubilized β-glucan. The MPEG system enhances β-glucan's solubility and loading efficiency through sequential sonication and cell extrusion. In vitro studies reveal that MPEG utilizes multiple endocytosis pathways, including phagocytosis, clathrin-mediated, and lipid raft-mediated routes, for effective β-glucan delivery into cells. Upon cellular internalization, MPEG components trigger dual activation of the MAPK signaling pathway and glycolysis in macrophages, leading to enhanced production of pro-inflammatory cytokines and lactic acid, ultimately strengthening innate immune responses. This MPEG system offers a promising approach to harnessing the immunostimulatory properties of natural polysaccharides while overcoming their solubility limitations, opening new avenues for targeted cellular delivery in immunomodulation therapies.

Cell extrusion drives neural crest cell delamination

Neural crest cells (NCC) comprise a heterogeneous population of cells with variable potency that contribute to nearly every tissue and organ throughout the body. Considered unique to vertebrates, NCC are transiently generated within the dorsolateral region of the neural plate or neural tube during neurulation. Their delamination and migration are crucial for embryo development as NCC differentiation is influenced by their final resting locations. Previous work in avian and aquatic species revealed that NCC delaminate via an epithelial-mesenchymal transition (EMT), which transforms these progenitor cells from static polarized epithelial cells into migratory mesenchymal cells with fluid front and back polarity. However, the cellular and molecular mechanisms facilitating NCC delamination in mammals are poorly understood. Through time-lapse imaging of NCC delamination in mouse embryos, we identified a subset of cells that exit the neuroepithelium as isolated round cells, which then halt for a short period prior to acquiring the mesenchymal migratory morphology classically associated with delaminating NCC. High-magnification imaging and protein localization analyses of the cytoskeleton, together with measurements of pressure and tension of delaminating NCC and neighboring neuroepithelial cells, revealed that round NCC are extruded from the neuroepithelium prior to completion of EMT. Furthermore, cranial NCC are extruded through activation of the mechanosensitive ion channel, PIEZO1. Our results support a model in which cell density, pressure, and tension in the neuroepithelium result in activation of the live cell extrusion pathway and delamination of a subpopulation of NCC in parallel with EMT, which has implications for cell delamination in development and disease.

Ultrastructural changes in epithelial cells on different stages of sinantropic muscoid dipterans fed with spores of Brevibacillus laterosporus

Ultrastructural changes in epithelial cells on different stages of sinantropic muscoid dipterans fed with spores of Brevibacillus laterosporus

Epithelial-to-mesenchymal transition and live cell extrusion contribute to measles virus release from human airway epithelia.

Measles virus (MeV) is a highly contagious respiratory virus transmitted via aerosols. To understand how MeV exits the airways of an infected host, we use unpassaged primary cultures of human airway epithelial cells (HAE). MeV typically remains cell-associated in HAE and forms foci of infection, termed infectious centers, by directly spreading cell-to-cell. We previously described the phenomenon in which infectious centers detach en masse from HAE and remain viable. Here, we investigate the mechanism of this cellular detachment. Via immunostaining, we observed loss of tight junction and cell adhesion proteins within infectious centers. These morphological changes indicate activation of epithelial-to-mesenchymal transition (EMT). EMT can contribute to wound healing in respiratory epithelia by mobilizing nearby cells. Inhibiting TGF-β, and thus EMT, reduced infectious center detachment. Compared with uninfected cells, MeV-infected cells also expressed increased levels of sphingosine kinase 1 (SK1), a regulator of live cell extrusion. Live cell extrusion encourages cells to detach from respiratory epithelia by contracting the actomyosin of neighboring cells. Inhibition or induction of live cell extrusion impacted infectious center detachment rates. Thus, these two related pathways contributed to infectious center detachment in HAE. Detached infectious centers contained high titers of virus that may be protected from the environment, allowing the virus to live on surfaces longer and infect more hosts.IMPORTANCEMeasles virus (MeV) is an extremely contagious respiratory pathogen that continues to cause large, disruptive outbreaks each year. Here, we examine mechanisms of detachment of MeV-infected cells. MeV spreads cell-to-cell in human airway epithelial cells (HAE) to form groups of infected cells, termed "infectious centers". We reported that infectious centers ultimately detach from HAE as a unit, carrying high titers of virus. Viral particles within cells may be more protected from environmental conditions, such as ultraviolet radiation and desiccation. We identified two host pathways, epithelial-to-mesenchymal transition and live cell extrusion, that contribute to infectious center detachment. Perturbing these pathways altered the kinetics of infectious center detachment. These pathways influence one another and contribute to epithelial wound healing, suggesting that infectious center detachment may be a usurped consequence of the host's response to infection that benefits MeV by increasing its transmissibility between hosts.

A 3D Computational Study on the Formation and Progression of Tumor Cells in Diffuse Gastric Cancer.

Hereditary diffuse gastric cancer is characterized by an increased risk of diffuse gastric cancer and lobular breast cancer, and is caused by pathogenic germline variants of E-cadherin and -E-catenin, which are key regulators of cell-cell adhesion. However, how the loss of cell-cell adhesion promotes cell dissemination remains to be fully understood. Therefore, a three-dimensional computer model was developed to describe the initial steps of diffuse gastric cancer development. In this model, we have implemented a cellular Potts approach that contemplates cell adhesion to other cells and to the extracellular matrix, cell extrusion from the gastric epithelia, and subsequent proliferation. We demonstrate that early disease features are determined by decreased adhesion of mutant cells to their normal epithelial neighbors, with concomitant increased attachment to matrix components. Importantly, our simulation shows how mechanical pressure and uncontrolled proliferation of mutant cells lead to modifications in cell shape and in gastric gland morphology. In conclusion, this work underscores the potential of computational models to elucidate the role of cellular and noncellular components in gastric cancer that may be relevant targets in therapeutic interventions.

Live Imaging Microscopy of Human Retina Organoids: Photoreceptor Pathology.

Neurodegenerative diseases involve many different (sub)cellular pathologic processes like mitochondria damage, which are still incompletely understood and present future targets for therapy development. Live imaging microscopy of dynamic pathologic changes at single cell resolution might advance studies of pathomechanisms and biomarkers. Organoid technologies may complement and facilitate animal and patient studies: We recently reported a novel pathomechanism of photoreceptor degeneration via cell extrusion, identified and validated by live imaging in human retina organoids. Here, we describe the related workflow for live imaging organoid studies. As another use-case, we describe mitochondrial alterations in photoreceptors acutely damaged by the ionophore CCCP. Our data further support a CCCP-induced experimental model for photoreceptor pathology, which can be monitored by JC-1 live dye-assisted live imaging. Thus, we demonstrate that live imaging advances studies of early subcellular pathologic events in human organoids. Such approaches might facilitate future therapeutic target identification and development for early intervention.

ERK activation waves coordinate mechanical cell competition leading to collective elimination via extrusion of bacterially infected cells.

Epithelial cells respond to infection with the intracellular bacterial pathogen Listeria monocytogenes by altering their mechanics to promote collective infected cell extrusion (CICE) and limit infection spread across cell monolayers. However, the underlying biochemical pathways remain elusive. Here, using invitro (epithelial monolayers) and invivo (zebrafish larvae) models of infection with L. monocytogenes or Shigella flexneri, we explored the role of extracellular-signal-regulated kinase (ERK) activity waves in coordinating the mechanical battle between infected and surrounder uninfected cells that leads to CICE. We discovered that when ERK waves are suppressed, cells fail to exhibit alterations in cell shape and kinematics associated with CICE and behave more like quiescent uninfected monolayers. In particular, uninfected cells surrounding infection foci are unable to polarize, reinforce their monolayer stresses, and promote CICE. Our findings reveal that crosstalk between ERK waves and cell mechanics is key to collective elimination of large domains of infected cells.

Epithelial Polarity Loss and Multilayer Formation: Insights Into Tumor Growth and Regulatory Mechanisms.

Epithelial tissues serve as critical barriers in metazoan organisms, maintaining structural integrity and facilitating essential physiological functions. Epithelial cell polarity regulates mechanical properties, signaling, and transport, ensuring tissue organization and homeostasis. However, the barrier function is challenged by cell turnover during development and maintenance. To preserve tissue integrity while removing dying or unwanted cells, epithelial tissues employ cell extrusion. This process removes both dead and live cells from the epithelial layer, typically causing detached cells to undergo apoptosis. Transformed cells, however, often resist apoptosis, leading to multilayered structures and early carcinogenesis. Malignant cells may invade neighboring tissues. Loss of cell polarity can lead to multilayer formation, cell extrusion, and invasion. Recent studies indicate that multilayer formation in epithelial cells with polarity loss involves a mixture of wild-type and mutant cells, leading to apical or basal accumulation. The directionality of accumulation is regulated by mutations in polarity complex genes. This phenomenon, distinct from traditional apical or basal extrusion, exhibits similarities to the endophytic or exophytic growth observed in human tumors. This review explores the regulation and implications of these phenomena for tissue biology and disease pathology.

Comparative Analysis of Single and Tapering FSH Doses on Follicular and Oocyte Yield in Sahiwal Cows

The present study was conducted at Embryo Transfer & In vitro Fertilization (ET & IVF) laboratory, College of Veterinary Science, Korutla, Telangana. This study assessed the effects of single and multiple doses of follicle stimulating hormone (FSH) stimulation protocols on ovarian follicular development, oocyte retrieval, oocyte quality, in-vitro maturation (IVM) in Sahiwal cows. Thirty cows, irrespective of their estrous cycle stage, were categorized into three groups: Group 1 (single dose FSH, n = 10), Group 2 (tapered FSH doses, n = 10) and Group 3 (unstimulated controls, n = 10). Ovum pickup (OPU) was conducted following FSH stimulation in Groups 1 and 2, while Group 3 underwent OPU without hormonal stimulation. Retrieved oocytes were graded and subjected to IVM. Both FSH stimulation protocols significantly increased the follicular count (Group 1: 20.51 ± 2.30; Group 2: 22.11 ± 2.21; Group 3: 5.61 ± 0.65) and the number of recovered oocytes per cow (Group 1: 11.81 ± 1.80; Group 2: 15.81 ± 2.45; Group 3: 2.81 ± 0.29). FSH stimulated groups exhibited greater number of oocytes with cumulus cell expansion and first polar body extrusion than the control group, although the rates of Cumulus cell expansion and extrusion in percentages did not differ significantly. In conclusion, both single dose and tapering doses FSH stimulation protocols effectively promoted follicular development and improved oocyte recovery in Sahiwal cows.

PIEZO1 Affects Cell Growth and Migration via Microfilament-Mediated YAP trans-Latitudinal Regulation.

Environmental mechanical forces, such as cell membrane stress, cell extrusion, and stretch, have been proven to affect cell growth and migration. Piezo1, a mechanosensitive channel protein, responds directly to endogenous or exogenous mechanical stimuli. Here, we explored the Piezo1 distribution and microfilament morphological changes induced by mechanical forces in the tumor and normal cells. In addition, Piezo1 activation in tumor cells resulted in the nuclear accumulation of YAP, whereas nuclear export of YAP and microfilament depolymerization occurred with the prolonged activation, while a removal stimulation restored the YAP distribution and microfilament polymerization. Combining the morphological changes of the microfilament under Piezo1 activation and the function of YAP in regulating cell growth and development, we suggest that Piezo1 senses changes in environmental mechanical forces and regulates YAP distribution through the microfilament cytoskeleton network, which in turn affects the growth and migration more obviously in tumor cells rather than normal cells. Our results are essential for understanding the trans-latitudinal transmission of mechanical forces and exploring the role of environmental mechanical forces in tumor therapy.

Morphofunctional Characteristics of the Mouse Small Intestine Mucosa After a sIngle Irradiation with Various Absorbed Doses

Radiation-induced intestinal damage is a typical complication of radiation exposure, it can be caused by various sources of radiation. Clinical treatment today consists of symptomatic supportive therapy, as there are no specific treatment strategies. The main target of radiation damage is the epithelial cells of the mucous membrane of the small intestine. The study of morphological changes and mechanisms of damage to epithelial cells is necessary when developing the model of radiation damage to the intestine for further assessment of the severity of pharmacological correction. The purpose of the work is to provide a morphological assessment to the state of the epithelial tissue in the mouse small intestine mucosa on the 9th day after a single exposure to X-ray radiation at various absorbed doses (6.5, 7 and 7.8 Gy). Materials and methods. The study was performed on sexually mature white outbred male mice weighing 18-22 g (n = 50). Acute radiation sickness was simulated using the RUM-17 X-ray therapy unit. The animals of the three experimental groups (n = 15 each) received a single X-ray irradiation with different absorbed doses of 6.5, 7 and 7.8 Gy, respectively. On the 9th day after irradiation, the material was fixed according to a routine technique for transmission electron microscopy. The analysis of semifine sections was carried out using a Scope A1c light microscope with an Axiocam ERc 5s camera and using the ZEN 2.3 morphometric program with further processing using Microsoft Office Excel 2013. Results. When exposed to radiation with different absorbed doses on the mucous membrane of the jejunum in mice, common morphological features were revealed: a decrease in the height and deformation of the mucous membrane villi; a decrease in the height of enterocytes from 32.03±2.21 µm in intact animals to 22.86±0.51 µm (at 7 Gy); in crypts, the height of epithelial cells had no significant changes; single dividing cells were located along the entire depth of the crypts; an increase in the length of the microvilli in the brush border on the apical surface of enterocytes from 0.89±0.01 µm to 1.46±0.03 µm (at 7.8 Gy) was revealed. Sublethal and lethal doses are characterized by a slight expansion of the basal parts of crypts with profiles of actively synthesizing goblet cells, in the absorbed dose of 6.5 Gy, similar changes are more pronounced. In all animals of the experimental groups, the loose connective tissue of the proper mucous plate is abundantly infiltrated by leukocytes. Epithelial cell extrusion zones are weakly expressed on the apical surfaces of the villi. A sharp decrease in the number of Paneth cells was revealed. Conclusions. On the 9th day after a single exposure to X-ray radiation at absorbed doses of 6.5, 7 and 7.8 Gy, reactive changes in the epithelial tissue of the mucous membrane of the small intestine have a non-specific character. The most pronounced of them were noted at an absorbed dose of 6.5 Gy. A decrease in the absorbing surface of the mucous membrane results in disturbances of its basic function.

Resolving the design principles that control postnatal vascular growth and scaling.

After birth, tissues grow continuously until reaching adult size, with each organ exhibiting unique cellular dynamics, growth patterns, and (stem or non-stem) cell sources. Using a suite of experimental and computational multiscale approaches, we found that aortic expansion is guided by specific biological principles and scales with the vertebral column rather than animal body weight. Expansion proceeds via two distinct waves of arterial cell proliferation along blood flow that are spatially stochastic, yet temporally coordinated. Each wave exhibits unique cell cycle kinetics and properties, with the first wave exhibiting cell cycle durations as fast as 6 hours. Single-cell RNA sequencing showed changes in fatty acid metabolism concomitant with an increase in cell size. Mathematical modeling and experiments indicated endothelial cell extrusion is essential for homeostatic aortic growth and balancing excess proliferation. In a genetic model of achondroplasia, the aorta achieves proper scaling through enhanced cell extrusion while maintaining normal proliferation dynamics. Collectively, these results provide a blueprint of the principles that orchestrate aortic growth which depends entirely on differentiated cell proliferation rather than resident stem cells.

Nuclei as mechanical bumpers during epithelial remodeling.

The morphogenesis of developing tissues relies on extensive cellular rearrangements in shape, position, and identity. A key process in reshaping tissues is cell intercalation-driven elongation, where epithelial cells align and intercalate along a common axis. Typically, analyses focus on how peripheral cortical forces influence cell shape changes. Less attention is given to how inhomogeneities in internal structures, particularly the nucleus, impact cell shaping. Here, we examine how pulsed contractile and extension dynamics interact with the nucleus in elongating Drosophila embryos. Our data show that tightly packed nuclei in apical layers hinder tissue remodeling/oscillatory behaviors. We identify two mechanisms for resolving internuclear tensions: nuclear deformation and dispersion. Embryos with non-deformable nuclei use nuclear dispersion to maintain near-normal extensile rates, while those with non-dispersible nuclei due to microtubule inhibition exhibit disruptions in contractile behaviors. Disrupting both mechanisms leads to severe tissue extension defects and cell extrusion. These findings highlight the critical role of nuclear shape and positioning in topological remodeling of epithelia.

Abstract C014: Degradation of extracellular trap DNA sustains anti-tumor immune responses in breast cancer

Abstract Increased extracellular DNA (exDNA) levels in the blood are a hallmark of metastatic cancer, arising from tumor lysis, apoptosis, necrosis, and neutrophil extracellular traps (NETs) released by tumor-expanded neutrophils. NETs, web-like chromatin structures extruded by neutrophils to trap pathogens, are highly immunogenic due to their DNA and histone content and their persistent interaction with dendritic cells. Tumor cells can also release extracellular traps. In myeloid leukemia, blasts release nuclear content via traps to activate coagulation or sustain myeloproliferation. Extracellular traps have been found in NPM1 mutant AML patients, with mutant NPM co-localizing with histones along exDNA traps. Forcing trap extrusion in NPM mutant leukemia cells has been used to create dendritic cell-based vaccines that break tolerance against NPMc antigens. This suggests that NETs in the tumor microenvironment (TME) may promote tolerance, with breaking tolerance requiring consistent NET formation and dendritic cell interaction. NETs are also seen in solid tumors, such as triple-negative breast cancer (TNBC), where they aid in cancer cell migration, invasion, and awakening of circulating tumor cells. However, the capacity of tumor cells to directly extrude DNA traps is not fully understood. To investigate this, various human and murine breast cancer cell lines were seeded on poly-D-lysine coated slides, stained with DAPI and Sytoxgreen, and observed via confocal microscopy. Tumor cells extruded DNA traps within 4 hours without stimuli. Notably, trap extrusion correlated with cell line aggressiveness and was abolished by DNase, indicating dsDNA composition. The absence of traps in 24-hour cultures suggests DNA extrusion may help rare tumor cells survive and proliferate. In line, DNase treatment reduced tumor proliferation and increased apoptosis. To assess the significance of DNA traps in vivo, BALB/c mice were injected with the highly metastatic 4T1 clone 5 breast cancer model, with or without DNase treatment, and monitored starting 5 days post-injection and weekly until day 28. Histological analysis at the injection site 5 days post-injection showed that DNase treatment reduced both local trap formation and tumor cell proliferation, as indicated by BrdU incorporation. Moreover, immunohistochemistry revealed that in DNase-treated mice, initial tumor growth was followed by complete tumor regression, accompanied by local and systemic CD8 T cell activation. Correspondingly, although a few metastases were detectable at day 14 in DNase-treated mice, no metastases were observed by day 28. Although it remains unclear whether tumor-derived or immune cell-derived traps are most relevant in our model, the results demonstrate that their presence in the TME promotes tolerance despite the inherent immune adjuvant properties of the traps. This suggests that DNase treatment could be particularly effective in combating early tumor development or relapse when used as part of a combination therapy designed to sustain anti-tumor immune responses. Citation Format: Sabina Sangaletti, Paola Portararo, Laura Botti, Valeria Cancila, Claudio Tripodo, Mario P. Colombo, Claudia Chiodoni. Degradation of extracellular trap DNA sustains anti-tumor immune responses in breast cancer [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Tumor-body Interactions: The Roles of Micro- and Macroenvironment in Cancer; 2024 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(22_Suppl):Abstract nr C014.

Toxic effects of chlorpyrifos on biochemical composition, enzyme activity and gill surface ultrastructure of three species of small fishes from India.

The effects of chlorpyrifos, a frequently detected organophosphate in aquatic ecosystems, on biochemical (protein and glycogen) contents and oxidative enzyme activities (catalase and lipid peroxidation) in liver, muscle and gill tissues of three freshwater fish Trichogaster fasciata, Mystus vittatus and Heteropneustes fossilis were evaluated after 21-day exposure to 1 and 10% of 96h LC50 of this pesticide, which were 1.63 and 16.3µg L-1; 5.87 and 58.7µg L-1 and 2.12 and 21.2µg L-1, respectively. On comparing with control, significant reductions in protein concentration were found in liver, muscle and gill of the three fishes treated with both higher as well as lower concentrations of the pesticide except in gill of M. vittatus and liver of H. fossilis treated with the lower concentrations. Glycogen content reductions were significant in the liver and muscle of the fishes, as well as gill tissue of T. fasciata treated with the two pesticide concentrations. Significant elevations of catalase activity were found in liver of the three fishes treated with the higher concentrations, in muscle tissues of both T. fasciata and M. vittatus treated with both the concentrations and in gills of the three fishes except H. fossilis treated with the lower concentration of the pesticide. Significant elevations of lipid peroxidation level were also found in liver of all the three fish species treated with the higher concentrations, in the muscle tissue of M. vittatus as well as in the gill of T. fasciata and H. fossilis treated with both the concentrations of the pesticide. Chlorpyrifos exposed gill ultrastructure of T. fasciata, M. vittatus and H. fossilis revealed concentration-dependent effects of the pesticide on gill surface ultrastructure which include distortion of primary and secondary lamellae, deterioration of pavement cell and microridge structures, extrusion of red blood cells (RBCs), secretion of mucous layer on filament, sloughing of primary lamellae and clumping of secondary lamellae. The present study parameters could serve as useful biomarkers for evaluating the risk of pesticide toxicity to fish. These findings also point out the possible health risks to the consumers of these fish captured from contaminated water bodies.

The IP3 receptor-KRAP complex at the desmosomes: A new player in the apoptotic process

The IP3 receptor-KRAP complex at the desmosomes: A new player in the apoptotic process

A sphingolipid rheostat controls apoptosis versus apical cell extrusion as alternative tumour-suppressive mechanisms

Evasion of cell death is a hallmark of cancer, and consequently the induction of cell death is a common strategy in cancer treatment. However, the molecular mechanisms regulating different types of cell death are poorly understood. We have formerly shown that in the epidermis of hypomorphic zebrafish hai1a mutant embryos, pre-neoplastic transformations of keratinocytes caused by unrestrained activity of the type II transmembrane serine protease Matriptase-1 heal spontaneously. This healing is driven by Matriptase-dependent increased sphingosine kinase (SphK) activity and sphingosine-1-phosphate (S1P)-mediated keratinocyte loss via apical cell extrusion. In contrast, amorphic hai1afr26 mutants with even higher Matriptase-1 and SphK activity die within a few days. Here we show that this lethality is not due to epidermal carcinogenesis, but to aberrant tp53-independent apoptosis of keratinocytes caused by increased levels of pro-apoptotic C16 ceramides, sphingolipid counterparts to S1P within the sphingolipid rheostat, which severely compromises the epidermal barrier. Mathematical modelling of sphingolipid rheostat homeostasis, combined with in vivo manipulations of components of the rheostat or the ceramide de novo synthesis pathway, indicate that this unexpected overproduction of ceramides is caused by a negative feedback loop sensing ceramide levels and controlling ceramide replenishment via de novo synthesis. Therefore, despite their initial decrease due to increased conversion to S1P, ceramides eventually reach cell death-inducing levels, making transformed pre-neoplastic keratinocytes die even before they are extruded, thereby abrogating the normally barrier-preserving mode of apical live cell extrusion. Our results offer an in vivo perspective of the dynamics of sphingolipid homeostasis and its relevance for epithelial cell survival versus cell death, linking apical cell extrusion and apoptosis. Implications for human carcinomas and their treatments are discussed.