bims-ginsta Biomed News
on Genome instability
Issue of 2024–06–02
twenty papers selected by
Jinrong Hu, National University of Singapore
  1. Force-activated zyxin assemblies coordinate actin nucleation and crosslinking to orchestrate stress fiber repair.
  2. Genome folding principles uncovered in condensin-depleted mitotic chromosomes.
  3. Tissue-specific landscape of protein aggregation and quality control in an aging vertebrate.
  4. The sex of organ geometry.
  5. CDK7 kinase activity promotes RNA polymerase II promoter escape by facilitating initiation factor release.
  6. Measuring the forces that shape early human embryos.
  7. Aging atlas reveals cell-type-specific effects of pro-longevity strategies.
  8. Cells dispose of cytoplasmic mitochondrial DNA by nucleoid-phagy.
  9. An alternative cell cycle coordinates multiciliated cell differentiation.
  10. SGLT2 inhibition eliminates senescent cells and alleviates pathological aging.
  11. Axonal endoplasmic reticulum tubules control local translation via P180/RRBP1-mediated ribosome interactions.
  12. MYC phase separation selectively modulates the transcriptome.
  13. TCAF1 promotes TRPV2-mediated Ca2+ release in response to cytosolic DNA to protect stressed replication forks.
  14. Compartmentalized Cytoplasmic Flows Direct Protein Transport to the Cell's Leading Edge.
  15. Asymmetric stem cell division maintains the genetic heterogeneity of tissue cells.
  16. Studying cell membrane tension and mechanosensation during mechanical stimulation.
  17. Molecular counting of myosin force generators in growing filopodia.
  18. SMAD4 promotes somatic-germline contact during murine oocyte growth.
  19. The evolutionarily ancient FOXA transcription factors shape the murine gut microbiome via control of epithelial glycosylation.
  20. Dissecting cell membrane tension dynamics and its effect on Piezo1-mediated cellular mechanosensitivity using force-controlled nanopipettes.
  1. bioRxiv. 2024 May 18. pii: 2024.05.17.594765. [Epub ahead of print]
    Force-activated zyxin assemblies coordinate actin nucleation and crosslinking to orchestrate stress fiber repair.
    Donovan Y Z Phua, Xiaoyu Sun, Gregory M Alushin.
      As the cytoskeleton sustains cell and tissue forces, it incurs physical damage that must be repaired to maintain mechanical homeostasis. The LIM-domain protein zyxin detects force-induced ruptures in actin-myosin stress fibers, coordinating downstream repair factors to restore stress fiber integrity through unclear mechanisms. Here, we reconstitute stress fiber repair with purified proteins, uncovering detailed links between zyxin's force-regulated binding interactions and cytoskeletal dynamics. In addition to binding individual tensed actin filaments (F-actin), zyxin's LIM domains form force-dependent assemblies that bridge broken filament fragments. Zyxin assemblies engage repair factors through multi-valent interactions, coordinating nucleation of new F-actin by VASP and its crosslinking into aligned bundles by ɑ-actinin. Through these combined activities, stress fiber repair initiates within the cores of micron-scale damage sites in cells, explaining how these F-actin depleted regions are rapidly restored. Thus, zyxin's force-dependent organization of actin repair machinery inherently operates at the network scale to maintain cytoskeletal integrity.
    Keywords:  Actin cytoskeleton; LIM domain; mechanosensing; mechanotranduction; stress fiber; zyxin
    DOI:  https://doi.org/10.1101/2024.05.17.594765
  2. Nat Genet. 2024 May 27.
    Genome folding principles uncovered in condensin-depleted mitotic chromosomes.
    Han Zhao, Yinzhi Lin, En Lin, Fuhai Liu, Lirong Shu, Dannan Jing, Baiyue Wang, Manzhu Wang, Fengnian Shan, Lin Zhang, Jessica C Lam, Susannah C Midla, Belinda M Giardine, Cheryl A Keller, Ross C Hardison, Gerd A Blobel, Haoyue Zhang.
      During mitosis, condensin activity is thought to interfere with interphase chromatin structures. To investigate genome folding principles in the absence of chromatin loop extrusion, we codepleted condensin I and condensin II, which triggered mitotic chromosome compartmentalization in ways similar to that in interphase. However, two distinct euchromatic compartments, indistinguishable in interphase, emerged upon condensin loss with different interaction preferences and dependencies on H3K27ac. Constitutive heterochromatin gradually self-aggregated and cocompartmentalized with facultative heterochromatin, contrasting with their separation during interphase. Notably, some cis-regulatory element contacts became apparent even in the absence of CTCF/cohesin-mediated structures. Heterochromatin protein 1 (HP1) proteins, which are thought to partition constitutive heterochromatin, were absent from mitotic chromosomes, suggesting, surprisingly, that constitutive heterochromatin can self-aggregate without HP1. Indeed, in cells traversing from M to G1 phase in the combined absence of HP1α, HP1β and HP1γ, constitutive heterochromatin compartments are normally re-established. In sum, condensin-deficient mitotic chromosomes illuminate forces of genome compartmentalization not identified in interphase cells.
    DOI:  https://doi.org/10.1038/s41588-024-01759-x
  3. Dev Cell. 2024 May 24. pii: S1534-5807(24)00266-1. [Epub ahead of print]
    Tissue-specific landscape of protein aggregation and quality control in an aging vertebrate.
    Yiwen R Chen, Itamar Harel, Param Priya Singh, Inbal Ziv, Eitan Moses, Uri Goshtchevsky, Ben E Machado, Anne Brunet, Daniel F Jarosz.
      Protein aggregation is a hallmark of age-related neurodegeneration. Yet, aggregation during normal aging and in tissues other than the brain is poorly understood. Here, we leverage the African turquoise killifish to systematically profile protein aggregates in seven tissues of an aging vertebrate. Age-dependent aggregation is strikingly tissue specific and not simply driven by protein expression differences. Experimental interrogation in killifish and yeast, combined with machine learning, indicates that this specificity is linked to protein-autonomous biophysical features and tissue-selective alterations in protein quality control. Co-aggregation of protein quality control machinery during aging may further reduce proteostasis capacity, exacerbating aggregate burden. A segmental progeria model with accelerated aging in specific tissues exhibits selectively increased aggregation in these same tissues. Intriguingly, many age-related protein aggregates arise in wild-type proteins that, when mutated, drive human diseases. Our data chart a comprehensive landscape of protein aggregation during vertebrate aging and identify strong, tissue-specific associations with dysfunction and disease.
    Keywords:  aggregates; aging; multi-tissue; protein homeostasis; proteomics; systems biology
    DOI:  https://doi.org/10.1016/j.devcel.2024.04.014
  4. Nature. 2024 May 29.
    The sex of organ geometry.
    Laura Blackie, Pedro Gaspar, Salem Mosleh, Oleh Lushchak, Lingjin Kong, Yuhong Jin, Agata P Zielinska, Boxuan Cao, Alessandro Mineo, Bryon Silva, Tomotsune Ameku, Shu En Lim, Yanlan Mao, Lucía Prieto-Godino, Todd Schoborg, Marta Varela, L Mahadevan, Irene Miguel-Aliaga.
      Organs have a distinctive yet often overlooked spatial arrangement in the body1-5. We propose that there is a logic to the shape of an organ and its proximity to its neighbours. Here, by using volumetric scans of many Drosophila melanogaster flies, we develop methods to quantify three-dimensional features of organ shape, position and interindividual variability. We find that both the shapes of organs and their relative arrangement are consistent yet differ between the sexes, and identify unexpected interorgan adjacencies and left-right organ asymmetries. Focusing on the intestine, which traverses the entire body, we investigate how sex differences in three-dimensional organ geometry arise. The configuration of the adult intestine is only partially determined by physical constraints imposed by adjacent organs; its sex-specific shape is actively maintained by mechanochemical crosstalk between gut muscles and vascular-like trachea. Indeed, sex-biased expression of a muscle-derived fibroblast growth factor-like ligand renders trachea sexually dimorphic. In turn, tracheal branches hold gut loops together into a male or female shape, with physiological consequences. Interorgan geometry represents a previously unrecognized level of biological complexity which might enable or confine communication across organs and could help explain sex or species differences in organ function.
    DOI:  https://doi.org/10.1038/s41586-024-07463-4
  5. Mol Cell. 2024 May 27. pii: S1097-2765(24)00400-3. [Epub ahead of print]
    CDK7 kinase activity promotes RNA polymerase II promoter escape by facilitating initiation factor release.
    Taras Velychko, Eusra Mohammad, Ivan Ferrer-Vicens, Iwan Parfentev, Marcel Werner, Cecilia Studniarek, Björn Schwalb, Henning Urlaub, Shona Murphy, Patrick Cramer, Michael Lidschreiber.
      Cyclin-dependent kinase 7 (CDK7), part of the general transcription factor TFIIH, promotes gene transcription by phosphorylating the C-terminal domain of RNA polymerase II (RNA Pol II). Here, we combine rapid CDK7 kinase inhibition with multi-omics analysis to unravel the direct functions of CDK7 in human cells. CDK7 inhibition causes RNA Pol II retention at promoters, leading to decreased RNA Pol II initiation and immediate global downregulation of transcript synthesis. Elongation, termination, and recruitment of co-transcriptional factors are not directly affected. Although RNA Pol II, initiation factors, and Mediator accumulate at promoters, RNA Pol II complexes can also proceed into gene bodies without promoter-proximal pausing while retaining initiation factors and Mediator. Further downstream, RNA Pol II phosphorylation increases and initiation factors and Mediator are released, allowing recruitment of elongation factors and an increase in RNA Pol II elongation velocity. Collectively, CDK7 kinase activity promotes the release of initiation factors and Mediator from RNA Pol II, facilitating RNA Pol II escape from the promoter.
    Keywords:  CDK7; CTD; Mediator; RNA polymerase II; analog sensitive; gene regulation; pre-initiation complex; promoter escape; transcription; transcription initiation
    DOI:  https://doi.org/10.1016/j.molcel.2024.05.007
  6. Nature. 2024 May 30.
    Measuring the forces that shape early human embryos.
      
    Keywords:  Biological techniques; Developmental biology
    DOI:  https://doi.org/10.1038/d41586-024-01556-w
  7. Nat Aging. 2024 May 30.
    Aging atlas reveals cell-type-specific effects of pro-longevity strategies.
    Shihong Max Gao, Yanyan Qi, Qinghao Zhang, Youchen Guan, Yi-Tang Lee, Lang Ding, Lihua Wang, Aaron S Mohammed, Hongjie Li, Yusi Fu, Meng C Wang.
      Organismal aging involves functional declines in both somatic and reproductive tissues. Multiple strategies have been discovered to extend lifespan across species. However, how age-related molecular changes differ among various tissues and how those lifespan-extending strategies slow tissue aging in distinct manners remain unclear. Here we generated the transcriptomic Cell Atlas of Worm Aging (CAWA, http://mengwanglab.org/atlas ) of wild-type and long-lived strains. We discovered cell-specific, age-related molecular and functional signatures across all somatic and germ cell types. We developed transcriptomic aging clocks for different tissues and quantitatively determined how three different pro-longevity strategies slow tissue aging distinctively. Furthermore, through genome-wide profiling of alternative polyadenylation (APA) events in different tissues, we discovered cell-type-specific APA changes during aging and revealed how these changes are differentially affected by the pro-longevity strategies. Together, this study offers fundamental molecular insights into both somatic and reproductive aging and provides a valuable resource for in-depth understanding of the diversity of pro-longevity mechanisms.
    DOI:  https://doi.org/10.1038/s43587-024-00631-1
  8. Nat Cell Biol. 2024 May 29.
    Cells dispose of cytoplasmic mitochondrial DNA by nucleoid-phagy.
      
    DOI:  https://doi.org/10.1038/s41556-024-01421-y
  9. Nature. 2024 May 29.
    An alternative cell cycle coordinates multiciliated cell differentiation.
    Semil P Choksi, Lauren E Byrnes, Mia J Konjikusic, Benedict W H Tsai, Rachel Deleon, Quanlong Lu, Christopher J Westlake, Jeremy F Reiter.
      The canonical mitotic cell cycle coordinates DNA replication, centriole duplication and cytokinesis to generate two cells from one1. Some cells, such as mammalian trophoblast giant cells, use cell cycle variants like the endocycle to bypass mitosis2. Differentiating multiciliated cells, found in the mammalian airway, brain ventricles and reproductive tract, are post-mitotic but generate hundreds of centrioles, each of which matures into a basal body and nucleates a motile cilium3,4. Several cell cycle regulators have previously been implicated in specific steps of multiciliated cell differentiation5,6. Here we show that differentiating multiciliated cells integrate cell cycle regulators into a new alternative cell cycle, which we refer to as the multiciliation cycle. The multiciliation cycle redeploys many canonical cell cycle regulators, including cyclin-dependent kinases (CDKs) and their cognate cyclins. For example, cyclin D1, CDK4 and CDK6, which are regulators of mitotic G1-to-S progression, are required to initiate multiciliated cell differentiation. The multiciliation cycle amplifies some aspects of the canonical cell cycle, such as centriole synthesis, and blocks others, such as DNA replication. E2F7, a transcriptional regulator of canonical S-to-G2 progression, is expressed at high levels during the multiciliation cycle. In the multiciliation cycle, E2F7 directly dampens the expression of genes encoding DNA replication machinery and terminates the S phase-like gene expression program. Loss of E2F7 causes aberrant acquisition of DNA synthesis in multiciliated cells and dysregulation of multiciliation cycle progression, which disrupts centriole maturation and ciliogenesis. We conclude that multiciliated cells use an alternative cell cycle that orchestrates differentiation instead of controlling proliferation.
    DOI:  https://doi.org/10.1038/s41586-024-07476-z
  10. Nat Aging. 2024 May 30.
    SGLT2 inhibition eliminates senescent cells and alleviates pathological aging.
    Goro Katsuumi, Ippei Shimizu, Masayoshi Suda, Yohko Yoshida, Takaaki Furihata, Yusuke Joki, Chieh-Lun Hsiao, Liang Jiaqi, Shinya Fujiki, Manabu Abe, Masataka Sugimoto, Tomoyoshi Soga, Tohru Minamino.
      It has been reported that accumulation of senescent cells in various tissues contributes to pathological aging and that elimination of senescent cells (senolysis) improves age-associated pathologies. Here, we demonstrate that inhibition of sodium-glucose co-transporter 2 (SGLT2) enhances clearance of senescent cells, thereby ameliorating age-associated phenotypic changes. In a mouse model of dietary obesity, short-term treatment with the SGLT2 inhibitor canagliflozin reduced the senescence load in visceral adipose tissue and improved adipose tissue inflammation and metabolic dysfunction, but normalization of plasma glucose by insulin treatment had no effect on senescent cells. Canagliflozin extended the lifespan of mice with premature aging even when treatment was started in middle age. Metabolomic analyses revealed that short-term treatment with canagliflozin upregulated 5-aminoimidazole-4-carboxamide-1-β-D-ribofuranoside, enhancing immune-mediated clearance of senescent cells by downregulating expression of programmed cell death-ligand 1. These findings suggest that inhibition of SGLT2 has an indirect senolytic effect by enhancing endogenous immunosurveillance of senescent cells.
    DOI:  https://doi.org/10.1038/s43587-024-00642-y
  11. Dev Cell. 2024 May 27. pii: S1534-5807(24)00322-8. [Epub ahead of print]
    Axonal endoplasmic reticulum tubules control local translation via P180/RRBP1-mediated ribosome interactions.
    Max Koppers, Nazmiye Özkan, Ha H Nguyen, Daphne Jurriens, Janine McCaughey, Dan T M Nguyen, Chun Hei Li, Riccardo Stucchi, Maarten Altelaar, Harold D MacGillavry, Lukas C Kapitein, Casper C Hoogenraad, Ginny G Farías.
      Local mRNA translation in axons is critical for the spatiotemporal regulation of the axonal proteome. A wide variety of mRNAs are localized and translated in axons; however, how protein synthesis is regulated at specific subcellular sites in axons remains unclear. Here, we establish that the axonal endoplasmic reticulum (ER) supports axonal translation in developing rat hippocampal cultured neurons. Axonal ER tubule disruption impairs local translation and ribosome distribution. Using nanoscale resolution imaging, we find that ribosomes make frequent contacts with axonal ER tubules in a translation-dependent manner and are influenced by specific extrinsic cues. We identify P180/RRBP1 as an axonally distributed ribosome receptor that regulates local translation and binds to mRNAs enriched for axonal membrane proteins. Importantly, the impairment of axonal ER-ribosome interactions causes defects in axon morphology. Our results establish a role for the axonal ER in dynamically localizing mRNA translation, which is important for proper neuron development.
    Keywords:  ER shape; ER-based translation; P180/RRBP1; axonal ER; axonal protein synthesis; endoplasmic reticulum; local translation; mRNA localization; neuron development; ribosomes
    DOI:  https://doi.org/10.1016/j.devcel.2024.05.005
  12. Nat Struct Mol Biol. 2024 May 29.
    MYC phase separation selectively modulates the transcriptome.
    Junjiao Yang, Chan-I Chung, Jessica Koach, Hongjiang Liu, Ambuja Navalkar, Hao He, Zhimin Ma, Qian Zhao, Xiaoyu Yang, Liang He, Tanja Mittag, Yin Shen, William A Weiss, Xiaokun Shu.
      Dysregulation and enhanced expression of MYC transcription factors (TFs) including MYC and MYCN contribute to the majority of human cancers. For example, MYCN is amplified up to several hundredfold in high-risk neuroblastoma. The resulting overexpression of N-myc aberrantly activates genes that are not activated at low N-myc levels and drives cell proliferation. Whether increasing N-myc levels simply mediates binding to lower-affinity binding sites in the genome or fundamentally changes the activation process remains unclear. One such activation mechanism that could become important above threshold levels of N-myc is the formation of aberrant transcriptional condensates through phase separation. Phase separation has recently been linked to transcriptional regulation, but the extent to which it contributes to gene activation remains an open question. Here we characterized the phase behavior of N-myc and showed that it can form dynamic condensates that have transcriptional hallmarks. We tested the role of phase separation in N-myc-regulated transcription by using a chemogenetic tool that allowed us to compare non-phase-separated and phase-separated conditions at equivalent N-myc levels, both of which showed a strong impact on gene expression compared to no N-myc expression. Interestingly, we discovered that only a small percentage (<3%) of N-myc-regulated genes is further modulated by phase separation but that these events include the activation of key oncogenes and the repression of tumor suppressors. Indeed, phase separation increases cell proliferation, corroborating the biological effects of the transcriptional changes. However, our results also show that >97% of N-myc-regulated genes are not affected by N-myc phase separation, demonstrating that soluble complexes of TFs with the transcriptional machinery are sufficient to activate transcription.
    DOI:  https://doi.org/10.1038/s41594-024-01322-6
  13. Nat Commun. 2024 May 30. 15(1): 4609
    TCAF1 promotes TRPV2-mediated Ca2+ release in response to cytosolic DNA to protect stressed replication forks.
    Lingzhen Kong, Chen Cheng, Abigael Cheruiyot, Jiayi Yuan, Yichan Yang, Sydney Hwang, Daniel Foust, Ning Tsao, Emily Wilkerson, Nima Mosammaparast, Michael B Major, David W Piston, Shan Li, Zhongsheng You.
      The protection of the replication fork structure under stress conditions is essential for genome maintenance and cancer prevention. A key signaling pathway for fork protection involves TRPV2-mediated Ca2+ release from the ER, which is triggered after the generation of cytosolic DNA and the activation of cGAS/STING. This results in CaMKK2/AMPK activation and subsequent Exo1 phosphorylation, which prevent aberrant fork processing, thereby ensuring genome stability. However, it remains poorly understood how the TRPV2 channel is activated by the presence of cytosolic DNA. Here, through a genome-wide CRISPR-based screen, we identify TRPM8 channel-associated factor 1 (TCAF1) as a key factor promoting TRPV2-mediated Ca2+ release under replication stress or other conditions that activate cGAS/STING. Mechanistically, TCAF1 assists Ca2+ release by facilitating the dissociation of STING from TRPV2, thereby relieving TRPV2 repression. Consistent with this function, TCAF1 is required for fork protection, chromosomal stability, and cell survival after replication stress.
    DOI:  https://doi.org/10.1038/s41467-024-48988-6
  14. bioRxiv. 2024 May 14. pii: 2024.05.12.593794. [Epub ahead of print]
    Compartmentalized Cytoplasmic Flows Direct Protein Transport to the Cell's Leading Edge.
    Catherine G Galbraith, Brian P English, Ulrike Boehm, James A Galbraith.
      Inside the cell, proteins essential for signaling, morphogenesis, and migration navigate complex pathways, typically via vesicular trafficking or microtubule-driven mechanisms 1-3 . However, the process by which soluble cytoskeletal monomers maneuver through the cytoplasm's ever-changing environment to reach their destinations without using these pathways remains unknown. 4-6 Here, we show that actin cytoskeletal treadmilling leads to the formation of a semi-permeable actin-myosin barrier, creating a specialized compartment separated from the rest of the cell body that directs proteins toward the cell edge by advection, diffusion facilitated by fluid flow. Contraction at this barrier generates a molecularly non-specific fluid flow that transports actin, actin-binding proteins, adhesion proteins, and even inert proteins forward. The local curvature of the barrier specifically targets these proteins toward protruding edges of the leading edge, sites of new filament growth, effectively coordinating protein distribution with cellular dynamics. Outside this compartment, diffusion remains the primary mode of protein transport, contrasting sharply with the directed advection within. This discovery reveals a novel protein transport mechanism that redefines the front of the cell as a pseudo-organelle, actively orchestrating protein mobilization for cellular front activities such as protrusion and adhesion. By elucidating a new model of protein dynamics at the cellular front, this work contributes a critical piece to the puzzle of how cells adapt their internal structures for targeted and rapid response to extracellular cues. The findings challenge the current understanding of intracellular transport, suggesting that cells possess highly specialized and previously unrecognized organizational strategies for managing protein distribution efficiently, providing a new framework for understanding the cellular architecture's role in rapid response and adaptation to environmental changes.
    DOI:  https://doi.org/10.1101/2024.05.12.593794
  15. bioRxiv. 2024 May 16. pii: 2024.05.16.594576. [Epub ahead of print]
    Asymmetric stem cell division maintains the genetic heterogeneity of tissue cells.
    Muhammed Burak Bener, Boris M Slepchenko, Mayu Inaba.
      Within a given tissue, the stem cell niche provides the microenvironment for stem cells suitable for their self-renewal. Conceptually, the niche space constrains the size of a stem-cell pool, as the cells sharing the niche compete for its space. It has been suggested that either neutral- or non-neutral-competition of stem cells changes the clone dynamics of stem cells. Theoretically, if the rate of asymmetric division is high, the stem cell competition is limited, thus suppressing clonal expansion. However, the effects of asymmetric division on clone dynamics have never been experimentally tested. Here, using the Drosophila germline stem cell (GSC) system, as a simple model of the in-vivo niche, we examine the effect of division modes (asymmetric or symmetric) on clonal dynamics by combining experimental approaches with mathematical modeling. Our experimental data and computational model both suggest that the rate of asymmetric division is proportional to the time a stem cell clone takes to expand. Taken together, our data suggests that asymmetric division is essential for maintaining the genetic variation of stem cells and thus serves as a critical mechanism for safeguarding fertility over the animal age or preventing multiple disorders caused by the clonal expansion of stem cells.
    DOI:  https://doi.org/10.1101/2024.05.16.594576
  16. Nat Methods. 2024 May 28.
    Studying cell membrane tension and mechanosensation during mechanical stimulation.
      
    DOI:  https://doi.org/10.1038/s41592-024-02278-7
  17. bioRxiv. 2024 May 17. pii: 2024.05.14.593924. [Epub ahead of print]
    Molecular counting of myosin force generators in growing filopodia.
    Gillian N Fitz, Matthew J Tyska.
      Animal cells build actin-based surface protrusions to enable biological activities ranging from cell motility to mechanosensation to solute uptake. Long-standing models of protrusion growth suggest that actin filament polymerization provides the primary mechanical force for "pushing" the plasma membrane outward at the distal tip. Expanding on these actin-centric models, our recent studies used a chemically inducible system to establish that plasma membrane-bound myosin motors, which are abundant in protrusions and accumulate at the distal tips, can also power robust filopodial growth. How protrusion resident myosins coordinate with actin polymerization to drive elongation remains unclear, in part because the number of force generators and thus, the scale of their mechanical contributions remain undefined. To address this gap, we leveraged the SunTag system to count membrane-bound myosin motors in actively growing filopodia. Using this approach, we found that the number of myosins is log-normally distributed with a mean of 12.0 ± 2.5 motors [GeoMean ± GeoSD] per filopodium. Together with unitary force values and duty ratio estimates derived from biophysical studies for the motor used in these experiments, we calculate that a distal tip population of myosins could generate a time averaged force of ∼tens of pN to elongate filopodia. This range is comparable to the expected force production of actin polymerization in this system, a point that necessitates revision of popular physical models for protrusion growth.
    SIGNIFICANCE STATEMENT: This study describes the results of in-cell molecular counting experiments to define the number of myosin motors that are mechanically active in growing filopodia. This data should be used to constrain future physical models of the formation of actin-based protrusions.
    DOI:  https://doi.org/10.1101/2024.05.14.593924
  18. Elife. 2024 May 31. pii: RP91798. [Epub ahead of print]13
    SMAD4 promotes somatic-germline contact during murine oocyte growth.
    Sofia Granados-Aparici, Qin Yang, Hugh J Clarke.
      Development of the mammalian oocyte requires physical contact with the surrounding granulosa cells of the follicle, which provide it with essential nutrients and regulatory signals. This contact is achieved through specialized filopodia, termed transzonal projections (TZPs), that extend from the granulosa cells to the oocyte surface. Transforming growth factor (TGFβ) family ligands produced by the oocyte increase the number of TZPs, but how they do so is unknown. Using an inducible Cre recombinase strategy together with expression of green fluorescent protein to verify Cre activity in individual cells, we examined the effect of depleting the canonical TGFβ mediator, SMAD4, in mouse granulosa cells. We observed a 20-50% decrease in the total number of TZPs in SMAD4-depleted granulosa cell-oocyte complexes, and a 50% decrease in the number of newly generated TZPs when the granulosa cells were reaggregated with wild-type oocytes. Three-dimensional image analysis revealed that TZPs of SMAD4-depleted cells were longer than controls and more frequently oriented towards the oocyte. Strikingly, the transmembrane proteins, N-cadherin and Notch2, were reduced by 50% in SMAD4-depleted cells. SMAD4 may thus modulate a network of cell adhesion proteins that stabilize the attachment of TZPs to the oocyte, thereby amplifying signalling between the two cell types.
    Keywords:  SMAD; developmental biology; follicle; granulosa; mouse; oocyte; transzonal projections
    DOI:  https://doi.org/10.7554/eLife.91798
  19. Dev Cell. 2024 May 29. pii: S1534-5807(24)00323-X. [Epub ahead of print]
    The evolutionarily ancient FOXA transcription factors shape the murine gut microbiome via control of epithelial glycosylation.
    Avital Swisa, Julia Kieckhaefer, Scott G Daniel, Hilana El-Mekkoussi, Hannah M Kolev, Mark Tigue, Chunsheng Jin, Charles-Antoine Assenmacher, Lenka Dohnalová, Christoph A Thaiss, Niclas G Karlsson, Kyle Bittinger, Klaus H Kaestner.
      Evolutionary adaptation of multicellular organisms to a closed gut created an internal microbiome differing from that of the environment. Although the composition of the gut microbiome is impacted by diet and disease state, we hypothesized that vertebrates promote colonization by commensal bacteria through shaping of the apical surface of the intestinal epithelium. Here, we determine that the evolutionarily ancient FOXA transcription factors control the composition of the gut microbiome by establishing favorable glycosylation on the colonic epithelial surface. FOXA proteins bind to regulatory elements of a network of glycosylation enzymes, which become deregulated when Foxa1 and Foxa2 are deleted from the intestinal epithelium. As a direct consequence, microbial composition shifts dramatically, and spontaneous inflammatory bowel disease ensues. Microbiome dysbiosis was quickly reversed upon fecal transplant into wild-type mice, establishing a dominant role for the host epithelium, in part mediated by FOXA factors, in controlling symbiosis in the vertebrate holobiont.
    Keywords:  FOXA; glycosylation; inflammatory bowel disease; intestinal epithelium; microbiome
    DOI:  https://doi.org/10.1016/j.devcel.2024.05.006
  20. Nat Methods. 2024 May 27.
    Dissecting cell membrane tension dynamics and its effect on Piezo1-mediated cellular mechanosensitivity using force-controlled nanopipettes.
    Ines Lüchtefeld, Igor V Pivkin, Lucia Gardini, Elaheh Zare-Eelanjegh, Christoph Gäbelein, Stephan J Ihle, Andreas M Reichmuth, Marco Capitanio, Boris Martinac, Tomaso Zambelli, Massimo Vassalli.
      The dynamics of cellular membrane tension and its role in mechanosensing, which is the ability of cells to respond to physical stimuli, remain incompletely understood, mainly due to the lack of appropriate tools. Here, we report a force-controlled nanopipette-based method that combines fluidic force microscopy with fluorescence imaging for precise manipulation of the cellular membrane tension while monitoring the impact on single-cell mechanosensitivity. The force-controlled nanopipette enables control of the indentation force imposed on the cell cortex as well as of the aspiration pressure applied to the plasma membrane. We show that this setup can be used to concurrently monitor the activation of Piezo1 mechanosensitive ion channels via calcium imaging. Moreover, the spatiotemporal behavior of the tension propagation is assessed with the fluorescent membrane tension probe Flipper-TR, and further dissected using molecular dynamics modeling. Finally, we demonstrate that aspiration and indentation act independently on the cellular mechanobiological machinery, that indentation induces a local pre-tension in the membrane, and that membrane tension stays confined by links to the cytoskeleton.
    DOI:  https://doi.org/10.1038/s41592-024-02277-8
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