Journal of Cell Biology最新文献

Segmenting multidimensional microscopy data requires high accuracy across many images (e.g., time points or Z slices) and is thus a labor-intensive part of biological image processing pipelines. We present ReSCU-Nets, recurrent convolutional neural networks that use the segmentation results from the previous image in a sequence as a prompt to segment the current image. We demonstrate that ReSCU-Nets outperform state-of-the-art image segmentation models, including nnU-Net and the Segment Anything Model, in different segmentation tasks on time-lapse microscopy sequences. Furthermore, ReSCU-Nets enable human-in-the loop corrections that prevent propagation of segmentation errors throughout image sequences. Using ReSCU-Nets, we investigate the role of gap junctions during Drosophila embryonic wound healing. We show that pharmacological blocking of gap junctions slows down wound closure by disrupting cytoskeletal polarity and cell shape changes necessary to repair the wound. Our results demonstrate that ReSCU-Nets enable the analysis of the molecular and cellular dynamics of tissue morphogenesis from multidimensional microscopy data.

Trisomy 12 is the most common whole-chromosome abnormality in human pluripotent stem cells. Conventionally, this acquired aneuploidy is ascribed to a rare single-cell event followed by selective growth advantage. Instead, we show that trisomy 12 emerges simultaneously in a very high percentage of cells in critical transition passages. Mis-segregation and incorporation of chromosome 12 into micronuclei occur through bridging of the short p arms of chromosome 12. Subsequently, single, unreplicated chromosome 12 chromatids are observed in mitotic cells. Erosion of the subtelomeric regions of the 12p arms is found during the passages when chromosome 12 bridges become frequent and trisomy 12 increases. Trisomy 12 cells persist due to a slight growth advantage. Among the shortest telomeres in humans are those on the 12p arms, making them particularly vulnerable to damage and bridging during mitosis. These findings reveal a novel mechanism of whole-chromosome instability in human stem cells, with broad implications for understanding the genesis of aneuploidy across diverse biological systems.

Cell division commonly produces two daughter cells, but there are many exceptions where large cells produce multiple daughters. Multiple fission of some green algae and bacteria; cellularization during embryogenesis of plants and insects; and growth of Ichthyosporeans, Chytrids, and Apicomplexans all provide variations on this theme. In some yeast species, a large multinucleate mother cell grows multiple buds (daughters) simultaneously. Here, we address how mothers partition growth equally among their buds in the multi-budding yeast Aureobasidium pullulans. Bud growth is directed by actin cable networks that appear to be optimized for even partitioning despite complex cell geometries. Even partitioning does not rely on compensatory mechanisms to adjust bud volumes but rather stems directly from effective equalization of polarity sites. These results reveal how conserved cell polarity and cytoskeletal networks are adapted to build complex morphologies in fungi.

Macrophage phagocytosis is an essential immune response that eliminates pathogens, antibody-opsonized cancer cells, and debris. Macrophages can also trogocytose, or nibble, targets. Trogocytosis and phagocytosis are often activated by the same signal, including IgG antibodies. What makes a macrophage trogocytose instead of phagocytose is not clear. Using both CD47 antibodies and a Her2 chimeric antigen receptor (CAR) to induce phagocytosis, we found that macrophages preferentially trogocytose adherent target cells instead of phagocytose in both 2D cell monolayers and 3D cancer spheroid models. Disrupting target cell integrin using an RGD peptide or through CRISPR-Cas9 knockout of the αV integrin subunit in target cells increased macrophage phagocytosis. In contrast, increasing cell-cell adhesion by ectopically expressing E-cadherin in Raji B cell targets reduced phagocytosis. Finally, we examined phagocytosis of mitotic cells, a naturally occurring example of cells with reduced adhesion. Arresting target cells in mitosis significantly increased phagocytosis. Together, our data show that adhesion of target cells limits phagocytosis and promotes trogocytosis.

Phosphatidic acid (PA) regulates lipid homeostasis and vesicular trafficking, yet high-affinity tools to study PA in live cells are lacking. We identified the lipin-like sequence of Nir1 (PILS-Nir1) as a candidate PA biosensor based on structural analysis of Nir1's LNS2 domain. Using liposome-binding assays and pharmacological and genetic manipulations in HEK293A cells expressing fluorescent PILS-Nir1, we found that while PILS-Nir1 binds PA and PIP2in vitro, only PA is necessary and sufficient for membrane localization in cells. PILS-Nir1 displayed greater sensitivity to organelle-generated PA than Spo20-based probes, enabling visualization of modest PA production by PLD downstream of muscarinic receptors-previously undetectable with existing biosensors. Thus, PILS-Nir1 provides a versatile, sensitive tool for real-time PA dynamics in live cells.

Cell-cell fusion is an evolutionarily conserved process that is essential for many functions, including the formation of bone-resorbing multinucleated osteoclasts. Osteoclast multinucleation involves dynamic interactions between the actin cytoskeleton and the plasma membrane that are still poorly characterized. We found that moesin, a cytoskeletal linker protein member of the Ezrin, radixin, and moesin (ERM) protein family, plays a critical role in both osteoclast fusion and function. Moesin inhibition favors osteoclast multinucleation as well as HIV-1- and inflammation-induced cell fusion. Accordingly, moesin depletion decreases membrane-to-cortex attachment and enhances the formation of tunneling nanotubes, F-actin-based intercellular bridges triggering cell-cell fusion. In addition, moesin regulates the formation of the sealing zone, a key structure determining osteoclast bone resorption area, and thus controls bone degradation via a β3-integrin/RhoA/SLK pathway. Finally, moesin-deficient mice have reduced bone density and increased osteoclast abundance and activity. These findings provide a better understanding of cell-cell fusion and osteoclast biology, opening new opportunities to specifically target osteoclasts in bone disease therapy.

Clathrin-mediated endocytosis (CME) is a critical cellular process that regulates nutrient uptake, membrane composition, and signaling. Although replicative aging affects many cellular functions, its impact on CME remains largely unknown. We show that in budding yeast, older cells have slower assembly of early and coat CME modules, resulting in longer endocytic turnover and reduced Mup1 internalization. This change in CME dynamics is mother cell-specific, and not observed in daughters. Our data also show that perturbing vacuolar pH, a key driver of aging phenotypes, in young cells mimics aging-like CME dynamics, while maintaining an acidic vacuolar pH in aging cells preserves CME dynamics typical of young cells. We demonstrate that the vacuolar pH effect on CME is regulated through TORC1 via the effector kinase Npr1. Finally, we show that rescuing CME in aging cells improves mitochondrial health. These findings reveal that age-associated changes in cellular and vacuolar pH impair CME, and suggest CME as a potential driver of early cellular aging.

Budding yeasts present an especially challenging geometry for segregation of chromosomes, which must be delivered across the narrow mother-bud neck into the bud. Studies in the model yeast Saccharomyces cerevisiae have revealed an elaborate set of mechanisms that selectively orient one mitotic spindle pole toward the bud and then drive spindle elongation along the mother-bud axis, ensuring nuclear segregation between mother and bud. It is unclear how these pathways might be adapted to yield similar precision in more complex cell geometries. Here, we provide the first description of the dynamics of mitosis in a multinucleate, multibudding yeast, Aureobasidium pullulans, and identify many unexpected differences from uninucleate yeasts. Mitotic spindles do not orient along the mother-bud axis prior to anaphase, and accurate nuclear segregation often occurs after spindle disassembly. Cortical Num1-dynein forces pull highly mobile nuclei into buds, and once a nucleus enters a bud, it discourages others from entering, ensuring that most daughters inherit only one nucleus.