Search bioRxiv⌕ Search

Biology subjects

Ikenouchi, J.

Publications and source records attributed to Ikenouchi, J..

9 recordsLinked to original sources

Environment-specific mechanosensing preserves a common bleb-based migratory program across diverse embryonic environments

Cell migration frequently requires cells to traverse tissue environments with distinct physical and biochemical properties. How migrating cells preserve a common migratory program while adapting to these heterogeneous environments remains poorly understood. Here, we show that chick primordial germ cells (PGCs) preserve a common bleb-based migratory mode throughout embryogenesis despite migrating through mechanically distinct tissues. PGCs formed membrane blebs during both intravascular crawling and migration through the dorsal mesentery. However, nuclear envelope (NE) unfolding and activation of the NE-cPLA2 pathway occurred specifically during migration through the mechanically confined dorsal mesentery, where this pathway was required for bleb formation and efficient migration. In contrast, bleb formation during vascular crawling occurred independently of the NE-cPLA2 pathway, demonstrating that distinct molecular mechanisms can generate the same migratory behavior in different tissue environments. Together, these findings suggest that diverse environmental cues converge on a conserved bleb-forming machinery. We propose a hierarchical model in which migrating cells preserve a common migratory program by flexibly switching the upstream molecular mechanisms that initiate bleb formation according to the tissue environment.

developmental biology↗

Bleb-based extravasation: conserved morphodynamics, divergent calcium control

Extravasation, the exit of cells from blood vessels, is essential in both development and disease, from germ cell migration to cancer metastasis. We show that all extravasating cells examined, avian primordial germ cells and diverse human cancer lines, form Ca{superscript 2}-dependent membrane blebs during transendothelial migration. Yet the source of Ca{superscript 2} diverges by lineage. PGCs and HT-1080 fibrosarcoma cells rely on store-operated Ca{superscript 2} entry (SOCE), whereas epithelial cancer cells such as PC-3 and MDA-MB-231 use IPR-mediated Ca{superscript 2} release from the endoplasmic reticulum. These findings establish bleb-based extravasation as a conserved morphodynamic strategy powered by distinct regulatory modules. Evolutionarily, they map onto an ancestral ER-release pathway and a metazoan-derived SOCE pathway. Conceptually, cancer blebbing emerges as a composite strategy that reactivates both ancient survival programs and developmental toolkits to maximize invasive success. This unified framework highlights Ca{superscript 2} supply as a critical bottleneck of vascular escape, offering new angles for targeting metastatic dissemination.

cell biology↗

A steady state pool of calcium-dependent actin is maintained by Homer and controls epithelial mechanosensation

Epithelial cells are inherently contractile and in homeostasis, tissue integrity is maintained by balancing the uneven contractile forces in neighboring cells at the cell-cell interface. By contrast, epithelial cells can utilize an imbalance in contractile force to communicate various information to induce tissue-wide response as in wound healing. Contractility is generated and processed at the apical junctional complex (AJC) by the dynamic behavior of the actin cytoskeleton. Calcium signaling can pattern cellular responses based on its reach and amplitude and the actin cytoskeleton is supported by its wide ranging effects on actin regulators. Calcium transients regulate various cell behaviors associated with actin remodeling, such as in damage response and developmental morphogenesis. Here we report that calcium maintains an adaptive pool of AJC-associated actin that is sensitive to tension and encoded by calcium dynamics. For this, the recently identified epithelial polarity module Homer-MUPP1/PatJ is required. Homer regulates calcium signaling in various tissue contexts through interaction with numerous components of the endoplasmic reticulum (ER) and plasma membrane (PM) calcium signal toolkit. Knockout of either Homer or MUPP1/PatJ attenuated tension-induced calcium response and severely disrupted wound healing migration, which is dependent on guidance input through AJC tension. We also show that Homer is integral to early embryonic neurodevelopment as its suppression causes failure of neural tube closure. Our findings highlight the critical role of localized calcium dynamics on AJC actin remodeling and cellular behavior, elucidating the means of tissue coordination through intercellular tension. Significance statementThis study uncovers a novel mechanism by which localized calcium dynamics at the apical junctional complex maintain an adaptive, tension sensitive pool of actin, regulated by the epithelial polarity scaffolds Homer and MUPP1/PatJ. Importantly, this mechanism operates without perturbing epithelial polarity, indicating a specific means to modulate tissue mechanics. By linking mechanical forces to localized calcium amplification, this module enables precise mechanosensation, coordinating collective behaviors such as epithelial wound healing and neural tube closure in Xenopus. These findings redefine our understanding of intercellular tension sensing in epithelial tissues and highlight the Homer-calcium signaling axis as a key driver of tissue morphogenesis and homeostasis, with far reaching implications for developmental biology, regenerative medicine, and neural tube defect pathogenesis.

cell biology↗

Wounding induces preexisting multinucleated cells to survive and thrive as leader cells

Epithelial wounds are repaired through collective cell migration, a process orchestrated by a small subset of leader cells at the wound edge1-3. How these functionally distinct cells arise from an apparently homogeneous population of epithelial cells remains unclear. Here, we show that injury to cultured epithelial sheets allows the survival of multinucleated cells that are otherwise eliminated under normal conditions. We reveal that multinucleated cells preexist prior to injury and extend protrusions toward the wound, eventually adopting leader-like behaviors. These findings identify multinucleated cells as a latent reservoir for leader cell emergence. Our work highlights the inherent heterogeneity of epithelial sheets and uncovers a previously unrecognized function of multinucleated cells during wound healing.

cell biology↗

Chemotherapy resistance due to epithelial-to-mesenchymal transition is caused by abnormal lipid metabolic balance

Invasive cancer is defined by the loss of epithelial cell traits resulting from the ectopic expression of epithelial-mesenchymal transition (EMT)-related transcription factors such as Snail. Although EMT is known to impart chemoresistance to cancer cells, the precise molecular mechanisms remain elusive. We found that Snail expression confers chemoresistance by upregulating the cholesterol efflux pump ABCA1 as a countermeasure to the excess of cytotoxic free cholesterol relative to its major interaction partner in cellular membranes, sphingomyelin. This imbalance is introduced by the transcriptional repression of enzymes involved in the biosynthesis of sphingomyelin by Snail. Inhibiting esterification of cholesterol, which renders it inert, selectively suppresses growth of a xenograft model of Snail-positive kidney cancer. Our findings offer a new perspective on lipid-targeting strategies for invasive cancer therapy.

cell biology↗

Rho-ROCK liberates sequestered claudin for rapid de novo tight junction formation

The epithelial cell sheet maintains its integrity as a barrier while undergoing turnover of constituent cells. To sustain the barrier continuously, its essential to preserve the old tight junctions (TJs) between cells being excluded from the sheet and their neighbors while simultaneously forming de novo TJs between newly adjacent cells. However, the molecular mechanisms involved in the formation of de novo TJs remain largely unknown. This study investigates two scenarios: the formation of de novo TJs during the removal of apoptotic cells from monolayer epithelial sheets and during the differentiation of the granular layer in stratified epidermis. We revealed that rapid claudin assembly is achieved by actively regulating the dissociation of the EpCAM/TROP2-claudin complex in both situations. Furthermore, we found that the Rho-ROCK pathway initiates the activation of matriptase, which cleaves EpCAM/TROP2, resulting in the supply of polymerizable claudin from the stockpiled EpCAM/TROP2-claudin complex at the plasma membrane to induce rapid de novo TJ formation.

cell biology↗

The yolk sac vasculature in early avian embryo provides a novel model for the analysis of cancer extravasation

Hematogenous metastasis, a key trait of cancer cells, involves a complex sequence of cell migration steps, including intravasation, circulation, arrest in capillary vessels, and extravasation. Among these steps, extravasation is challenging to image in amniotes like humans and mice due to its unpredictable timing and location, limiting our understanding of cellular and molecular mechanisms through imaging. Establishing a new cancer carrier model with high-resolution imaging capabilities in amniotes is crucial. In this study, we investigated the yolk sac vasculature (YSV) of early avian embryos (chickens and quail) as a new model for studying extravasation, offering excellent imaging capabilities. We examined the YSV structure and attempted fluorescent labeling to enhance visibility. We then injected mCherry-labeled HT-1080 cells into YSV and observed their behavior, revealing distinct morphologies and extravasation dynamics. Our findings suggest that the YSV model holds promise as a novel cancer carrier model for elucidating cellular and molecular mechanisms through imaging-based approaches.

cancer biology↗

The SOCE system is critical for membrane bleb formation to drive avian primordial germ cell migration

Amoeboid cell migration is driven by the specialized cell protrusion, membrane bleb. A recent in vitro analysis of bleb formation using a cancer cell line showed that store-operated calcium entry (SOCE)-mediated local elevation of Ca2+ concentration triggers bleb formation, but it remains unknown how commonly this system is utilized in the bleb formation and bleb-driven cell migration in vivo. We demonstrate with avian primordial germ cell (PGC) model that that chick PGCs use SOCE, triggered by stem cell factor (SCF), to induce bleb formation and that it is essential for in vivo migration including trans-endothelial and mesenchymal cell migration. Our discovery also provides insight into the correlation between cancer metastasis/invasion and SOCE-mediated bleb formation.

developmental biology↗

Cholesterol-mediated tight junction formation

Lipids have the ability to self-organize but the significance of this phenomenon in the assembly of membrane structures is unknown. We previously reported that cholesterol enrichment directs the formation of tight junctions (TJs), adhesion structures responsible for the epithelial barrier (Shigetomi et al., 2018). However, it is unclear how cholesterol accumulates and informs TJ formation. Cholesterol typically accumulates in the vicinity of apical cell junctions (Shigetomi et al., 2018). Surprisingly, cholesterol distribution is unaltered in an epithelial cell line that lacks TJs, generated by suppressing the expression of claudins, membrane proteins that determine the barrier properties of TJs. Assembly of claudin into hallmark strands is canonically thought to require binding to Zonula occludens (ZO) proteins but a claudin mutant that cannot bind to ZO proteins still form strands. These results suggest a novel mechanism for TJ formation that is dependent on cholesterol and reveal an unexpected role of ZO proteins as organizers of the cholesterol domain.

cell biology↗