Lysosomes are multifunctional organelles that play important roles in cellular recycling, signaling, and homeostasis, relying on precise trafficking and activation of lysosomal enzymes. While the Golgi apparatus plays a...Lysosomes are multifunctional organelles that play important roles in cellular recycling, signaling, and homeostasis, relying on precise trafficking and activation of lysosomal enzymes. While the Golgi apparatus plays a central role in lysosomal enzyme sorting, the mechanisms linking Golgi function to lysosomal activity remain incompletely understood. Here, we identify the Golgi-resident protein GRASP55, but not its paralog GRASP65, as necessary for lysosome function. Loss of GRASP55 expression leads to missorting and secretion of lysosomal enzymes, lysosomal dysfunction and bloating. GRASP55 deficiency also disrupts lysosomal mTORC1 signaling, reducing the phosphorylation of its lysosomal substrates TFEB/TFE3, while sparing its non-lysosomal targets. Mechanistically, GRASP55 binds and maintains the COPI adaptor GOLPH3 protein at the Golgi, thereby controlling the Golgi localization and stability of LYSET and GNPTAB that are required for mannose 6-phosphate (M6P) tagging of lysosomal enzymes. These findings reveal an essential role for GRASP55 in Golgi-lysosome communication and lysosomal enzyme trafficking and underscore the importance of Golgi-mediated protein sorting in lysosome function and lysosomal mTORC1 signaling.
Puzzle-shaped epidermal cells can reduce mechanical stress during organ growth and, as shown here, can also record tissue expansion history in their outlines. By combining mechanical simulations with time-lapse imaging,...Puzzle-shaped epidermal cells can reduce mechanical stress during organ growth and, as shown here, can also record tissue expansion history in their outlines. By combining mechanical simulations with time-lapse imaging, we find that transitions from directional to isotropic expansion induce new lobes along the previous growth axis, and that reversing the sequence of anisotropic and isotropic phases yields hybrid shapes that preserve the sequence of growth phases. In maize, model predictions closely match live imaging, and in Arabidopsis, final lobe patterns correlate more strongly with growth history than with cell size alone. Genetic or pharmacological perturbations that reduce lobing are associated with constrained leaf expansion or compensatory elongation, consistent with a mechanical role. A broad survey of living and fossil vascular plants indicates that the capacity to form puzzle-shaped cells is widespread and developmentally plastic, suggesting that single snapshots of leaves can reflect prior growth dynamics across species. Together, these findings show that puzzle cells transform cell geometry into a living record of how tissues grow.
The use of AI in grant writing to address non-scientific selection criteria along with growing application numbers and score inflation have turned many research funding schemes efficiently into lotteries. [Image: see tex...The use of AI in grant writing to address non-scientific selection criteria along with growing application numbers and score inflation have turned many research funding schemes efficiently into lotteries. [Image: see text]
Feng CH, Peltier L, Chouleur T
… +13 more, DiPonzio M, Aubry I, Poirier AJ, Cordova ZM, Shen Y, Tabariès S, Cao X, Chen G, Bikfalvi A, Vidal SM, Siegel PM, Laneuville P, Tremblay ML
Natural killer (NK) cells are promising candidates for allogeneic anti-cancer immunotherapy. However, their cytolytic activity is often suppressed by the tumor microenvironment. We demonstrate that genetic silencing or p...Natural killer (NK) cells are promising candidates for allogeneic anti-cancer immunotherapy. However, their cytolytic activity is often suppressed by the tumor microenvironment. We demonstrate that genetic silencing or pharmacological dual inhibition of protein tyrosine phosphatases PTPN1 and PTPN2 (PTPN1/N2) in NK cells significantly enhances anti-tumor cytolytic activity both in vitro and in vivo. This augmented NK cell activity is mediated by increased expression of early activation markers and the production of effector molecules such as granzyme B and interferon-gamma (IFN-γ). Notably, this elevated cell cytolytic response remains substantially resistant to the immunosuppressive effects of TGFβ-1, a cytokine known to dampen NK cell activity and commonly present in the tumor microenvironment. Mechanistically, targeting PTPN1/N2 in NK cells promotes JAK/STAT signaling pathways and sensitizes cells to IL-2 stimulation. Importantly, dual inhibition of PTPN1/N2 markedly enhances the cytolytic activity of cord blood NK cells against patient-derived glioblastoma cells, highlighting the potential of this approach for future therapeutic applications. These findings provide compelling evidence that dual targeting of PTPN1/N2 could significantly improve the efficacy of therapeutic "off-the-shelf" NK cell-based immunotherapy.
The genome-wide chromosome conformation capture method, Hi-C, has greatly advanced our understanding of genome organization. However, its quantitative properties, including sensitivity, bias, and linearity, remain challe...The genome-wide chromosome conformation capture method, Hi-C, has greatly advanced our understanding of genome organization. However, its quantitative properties, including sensitivity, bias, and linearity, remain challenging to assess. Measuring these properties in vivo is difficult due to the heterogenous and dynamic nature of chromosomal interactions. Here, using Chemically Induced Chromosomal Interaction (CICI) method, we create stable intra- and inter-chromosomal interactions in G1-phase budding yeast across a broad range of contact frequencies. Hi-C analysis of these engineered cell populations demonstrates that static intra-chromosomal loops do not generate Topologically Associated Domains (TADs) and only promote 3D proximity within 10-60 kb flanking regions. At moderate sequencing depth, Hi-C is sensitive enough to detect interactions occurring in 5-10% of cells. It also shows no inherent bias toward intra- versus inter-chromosomal interactions. Furthermore, we observe a linear relationship between Hi-C signal intensity and contact frequency. These findings illuminate the intrinsic properties of the Hi-C assay and provide a robust framework for its calibration.
Although glycerol is a ubiquitous metabolite in mammalian systems, its cellular metabolic pathways and functions have not been fully elucidated. Here, we find that elevated extracellular glycerol modulates intracellular...Although glycerol is a ubiquitous metabolite in mammalian systems, its cellular metabolic pathways and functions have not been fully elucidated. Here, we find that elevated extracellular glycerol modulates intracellular metabolism and pro-inflammatory responses of macrophages. In pro-inflammatory macrophages stimulated with lipopolysaccharide, glycerol is taken up through glycerol channels including Aquaporin 3 (AQP3) and metabolized to glycerol-3-phosphate (G3P), which is then converted to dihydroxyacetone phosphate by glycerol-3-phosphate dehydrogenase 2 (GPD2). This glycerol-driven pathway enhances mitochondrial ATP production, potentially by supplying electrons to the electron transport chain (ETC) via GPD2, and by upregulating the transcription of genes encoding ETC complexes. In addition, glycerol supplementation elevates intracellular acetyl-CoA levels, promotes histone acetylation at the promoters of pro-inflammatory cytokine genes, and consequently increases cytokine gene expression, suggesting enhanced pro-inflammatory response. In vivo experiments, macrophage-specific AQP3 conditional knockout mice exhibit reduced weight gain and adipose tissue inflammation in a high-fat diet-induced obesity model. Our findings provide novel insights into the metabolic regulation and macrophage inflammation by extracellular glycerol.
The 5' untranslated region (5' UTR) of an mRNA is classically viewed as a regulatory region that controls the amount of protein production, but not the resulting protein sequence. Here, we demonstrate that 5' UTR length...The 5' untranslated region (5' UTR) of an mRNA is classically viewed as a regulatory region that controls the amount of protein production, but not the resulting protein sequence. Here, we demonstrate that 5' UTR length plays a direct role in alternative N-terminal protein isoform production by controlling start codon selection. We find that very short 5' UTRs enhance leaky ribosome scanning, thereby promoting the production of truncated alternative N-terminal protein isoforms. We also show that endogenous changes in 5' UTR length due to alternative transcription initiation can tune the relative abundance of alternative N-terminal isoforms from the same gene. In addition, we identify mutations in rare genetic diseases that alter 5' UTR length, including a deletion in the VHL 5' UTR in von Hippel-Lindau disease that shifts translation toward the shorter VHLp19 isoform. Together, our results implicate 5' UTR length as a determinant of alternative N-terminal isoform production and reveal an underappreciated mechanism by which noncoding changes can reshape the proteome.
Aneuploidy is one of the most common adaptive mechanisms to environmental selection in cells, yet its advantages over other genomic alterations remain unclear. We used budding yeast to determine if beneficial aneuploidy...Aneuploidy is one of the most common adaptive mechanisms to environmental selection in cells, yet its advantages over other genomic alterations remain unclear. We used budding yeast to determine if beneficial aneuploidy phenotypes are driven primarily by combinatorial expression changes of multiple genes. To determine the impact of complex aneuploidy on cellular fitness and drug resistance, we generated yeast collections with nearly every combination of two chromosome gains or losses. Additionally, we genetically dissected aneuploid chromosomes using partial deletions to identify chromosomal regions contributing to aneuploidy-driven drug resistance. Strong resistance phenotypes consistently came from combinations of chromosomes or chromosomal regions, with up to five regions on a chromosome contributing to resistance to a single drug. Gene ontology terms had limited predictive power in identifying the genes contributing to resistance phenotypes, as the combinatorial effects from different aneuploid regions act through multiple resistance pathways. The strongest phenotypes came from synergistic effects between copy number changes of different chromosomes or chromosomal regions, demonstrating how subtle gene expression changes of many genes combine to greatly impact cell survival.
N-WASP and WASP can induce actin polymerization via Arp2/3 and were reported to be crucial for homology-directed repair (HDR) of DNA double-strand breaks (DSB). The underlying mechanism was suggested to involve nuclear a...N-WASP and WASP can induce actin polymerization via Arp2/3 and were reported to be crucial for homology-directed repair (HDR) of DNA double-strand breaks (DSB). The underlying mechanism was suggested to involve nuclear actin polymerization, but the mechanistic details were debated. Unexpectedly, we show now that neither WASP nor N-WASP is required for HDR during CRISPR-mediated genome editing. Using knock-out and overexpression of N-WASP and WASP in U2OS cells, we did not detect alterations in total gene editing, HDR, or the ratio of HDR to non-homologous end joining (NHEJ) as assessed by different methods. Furthermore, we could not observe colocalization of HA-tagged WASP or N-WASP with DSBs. Finally, while the Arp2/3 inhibitor CK-666 and ARPC4 knockdown by siRNA reduced HDR efficiency in U2OS cells, this corresponded with a decreased transfection efficiency and a reduction of the HDR-proficient cell cycle phases S and G2/M. In summary, contrary to expectations, these data do not support a crucial role for N-WASP and WASP in DSB repair.
Head and neck squamous cell carcinoma is an aggressive malignancy with limited therapeutic options and poor outcomes. A key driver of tumor progression is the tumor microenvironment, particularly cancer-associated fibrob...Head and neck squamous cell carcinoma is an aggressive malignancy with limited therapeutic options and poor outcomes. A key driver of tumor progression is the tumor microenvironment, particularly cancer-associated fibroblasts, which remodel the extracellular matrix and secrete pro-tumorigenic signals. Emerging evidence suggests that vesicle trafficking pathways regulate these secretory functions, though their role in HNSCC remains unclear. Building on our discovery of Gαq as an autophagy regulator, we investigated how its absence reshapes fibroblast behavior and modulates crosstalk with HNSCC cells. We demonstrate that loss of Gαq reprograms murine fibroblasts into a CAF-like phenotype through deregulated intracellular trafficking and increased ceramide accumulation. Gαq-deficient fibroblasts show increased collagen I deposition, ECM remodeling, and secretion. Their exosomes, enriched in tumor-promoting growth factor receptors, suppress Caveolin-1 in tumor cells and induce an EMT-like phenotype that fuels HNSCC growth. In co-culture and in vivo, Gαq-silenced fibroblasts form "railroad-track" structures guiding cancer cell migration and invasion. Reduced Gαq expression in human fibroblasts recapitulates these features, identifying Gαq as a key regulator of fibroblast plasticity and tumor-stroma interactions in HNSCC progression.
Stem cell protection is key to proper plant development. The root cap tissue performs this task by enclosing delicate stem cells in the root meristem and consists of columella cells at the tip and lateral root cap (LRC)...Stem cell protection is key to proper plant development. The root cap tissue performs this task by enclosing delicate stem cells in the root meristem and consists of columella cells at the tip and lateral root cap (LRC) cells covering the meristem. Unlike other tissues, the formation of new layers in the LRC is balanced with a regular loss through sloughing and programmed cell death in the outermost layers. The molecular mechanisms regulating this balance are still largely unknown. Brassinosteroids (BR) hormones control meristem size, radial growth, and cell shape during root development by acting non-cell-autonomously from the LRC and the epidermis. Here, we show that BR act cell-autonomously in this tissue to control LRC cell number by promoting cell-cycle entry of LRC cells through the downstream signalling factor BRASSINAZOLE-RESISTANT-1 (BZR1). Via genetic, molecular, and pharmacological analysis, we show that BZR1 maintains the LRC cell number by promoting the expression of the cell cycle regulator CYCLIN D3;3 (CYCD3;3). Our results enlighten a previously unrecognised module that ensures proper lateral root cap development.
Tsuji S, Nakano S, Ito K
… +19 more, Minami S, Uemura K, Konishi Y, Wakita M, Okumura Y, Kawamoto S, Matsuki A, Nakaoka S, Ono C, Takahashi H, Anzai I, Watanabe T, Uezumi A, Matsuura Y, Kobayashi T, Okamoto T, Tsuboi A, Asagiri M, Hara E
Persistent hyposmia is a hallmark of post COVID-19 conditions, yet the mechanisms sustaining olfactory dysfunction after viral clearance remain poorly understood. Here, using mouse models of SARS-CoV-2 infection, we show...Persistent hyposmia is a hallmark of post COVID-19 conditions, yet the mechanisms sustaining olfactory dysfunction after viral clearance remain poorly understood. Here, using mouse models of SARS-CoV-2 infection, we show that virus-induced senescence-like changes in uninfected olfactory mucosal fibroblasts persist long after viral clearance and drive prolonged olfactory dysfunction. These senescence-like cells secrete SASP factors, including IFNγ, CXCL9, and CXCL11, thereby recruiting γδ T cells to the olfactory mucosa. The accumulated γδ T cells produce excessive IL-17A, which acts on IL-17 receptor A expressed on olfactory sensory neurons, leading to sustained impairment of their function. Genetic ablation of senescence pathways (p16/p21 double knockout), pharmacological elimination of senescent cells with the senolytic drug ABT263, or olfactory neuron-specific deletion of IL-17 receptor A each significantly alleviate prolonged olfactory dysfunction. These findings identify a senescence-γδ T cell-IL-17A axis as a key driver of prolonged hyposmia following SARS-CoV-2 infection in mice.
Post-transcriptional maturation of the U6 snRNA 3'-end, important for spliceosome assembly, is catalyzed by sequential actions of TUT1 and USB1. It is believed that the TUT1-catalyzed oligo(U) tail at the U6 snRNA 3'-end...Post-transcriptional maturation of the U6 snRNA 3'-end, important for spliceosome assembly, is catalyzed by sequential actions of TUT1 and USB1. It is believed that the TUT1-catalyzed oligo(U) tail at the U6 snRNA 3'-end serves merely as a substrate for USB1 to generate a final 2',3'-cyclic phosphate group to mature the U6 snRNA. However, biallelic inactivation of TUT1 or USB1 is linked to distinct human developmental disorders, suggesting that they have different physiological functions. Here, using genetically engineered mouse models, we show that Tut1 is required to maintain stem cell pools during embryogenesis, whereas unexpectedly Usb1 is dispensable for this. Loss of Tut1 weakens the interaction of the U6 snRNA with the Lsm2-8 protein complex, causes defective RNA splicing, and triggers massive DNA damage and subsequent cell death. Splicing defects and cell death can be mitigated by recombinant U6 snRNA containing an oligo(U) tail. We propose that the TUT1-catalyzed oligo(U) tail is essential for splicing and cell proliferation. Further modification of this oligo(U) tail by USB1 is ubiquitous but only functionally required in specific cell types.
Core facilities are central to life-science research, yet performance frameworks borrowed from academia or administrative settings often miss what matters most in a technology-driven, service-focused environment. This ar...Core facilities are central to life-science research, yet performance frameworks borrowed from academia or administrative settings often miss what matters most in a technology-driven, service-focused environment. This article proposes a practical approach to selecting strategic KPIs to steer priorities and guide decisions. [Image: see text]
When RNA polymerase II (RNAPII) stalls at transcription-blocking lesions, the transcription-coupled DNA repair (TCR) pathway is activated to remove the damage. After repair, efficient transcription restart requires the P...When RNA polymerase II (RNAPII) stalls at transcription-blocking lesions, the transcription-coupled DNA repair (TCR) pathway is activated to remove the damage. After repair, efficient transcription restart requires the PAF1 elongation complex (PAF1C). PAF1C promotes deposition of transcription-associated histone marks, which are enriched at active genes and proposed to support post-repair transcription recovery. Using conditional knockouts of writers of PAF1C-associated histone marks, we show that deposition of H3K79, H3K4, and H2BK120 is dispensable for transcription restart. While H3K4 levels remain mostly unchanged during DNA damage-induced transcription inhibition, H2BK120 levels decrease after damage and are co-transcriptionally restored upon transcription restart. We further find that, unlike the core subunits PAF1 and CTR9, the dissociable PAF1C subunit RTF1 does not contribute to transcription restart. Finally, we show that the TCR core factor CSB is not required for the transient PAF1C interaction during normal transcription, but specifically stabilizes PAF1C on RNAPII in response to DNA damage-induced stalling. Together, these findings indicate that transcription restoration after DNA damage is driven by PAF1C independently of transcription-associated histone mark deposition.
Capsaspora owczarzaki is a protist that may both reveal aspects of animal evolution and curtail the spread of schistosomiasis. Capsaspora exhibits a regulated aggregative behavior reminiscent of cellular aggregation in s...Capsaspora owczarzaki is a protist that may both reveal aspects of animal evolution and curtail the spread of schistosomiasis. Capsaspora exhibits a regulated aggregative behavior reminiscent of cellular aggregation in some animals-a process that might have contributed to the origin of animals. This aggregative behavior may also be vital for Capsaspora to colonize the intermediate host of parasitic schistosomes and potentially prevent the spread of schistosomiasis. Both applications demand elucidation of the mechanisms underlying Capsaspora aggregation. Toward this goal, we evaluated the chemical properties of lipid cues that activate aggregation. We found that a range of zwitterionic lipids induced this behavior, revealing that aggregation is activated by diverse lipid-rich conditions. Furthermore, we demonstrated that aggregation in Capsaspora requires clathrin-mediated endocytosis, highlighting the potential significance of endocytosis-linked cellular signaling in recent animal ancestors. Finally, we found that aggregation is initiated independently of protein translation, suggesting post-translational activation of filopodial retraction. Together, our findings illuminate the molecular and cellular basis of Capsaspora's aggregative behavior, with implications for the evolution of animal multicellularity and the transmission of parasites.
Genome-wide association studies (GWASs) have identified hundreds of obesity-associated SNPs, but establishing their causality remains challenging. Here, we demonstrate that rs11676272, located in the ADCY3 gene, is a fun...Genome-wide association studies (GWASs) have identified hundreds of obesity-associated SNPs, but establishing their causality remains challenging. Here, we demonstrate that rs11676272, located in the ADCY3 gene, is a functional causal variant for obesity susceptibility. Bioinformatic analyses and dual-luciferase reporter assays indicate that the rs11676272 region may act as a human-gained enhancer regulating ADCY3 expression. In HEK293T cells, CRISPR-Cas9-mediated single-nucleotide editing of rs11676272 (T > C) reduces ADCY3 expression. Moreover, the rs11676272-T allele is preferentially bound by the transcription factor E2F3 to upregulate ADCY3 expression, whereas the rs11676272-C risk allele loses this binding. In vivo, the rs11676272 T > C variant in human ADCY3 (hADCY3) knock-in mice accelerates weight gain under high-fat diet conditions and shortens primary cilia in the ventromedial hypothalamus (VMH). CRISPRa-mediated activation of the hADCY3 promoter region rescues ciliary length in both the VMH and hypothalamic arcuate nucleus of Mut-hADCY3 mice. Our data reveal a causal role for rs11676272 in obesity, offering insight into potential therapeutic strategies.
In 1926, Thomas Hunt Morgan published , which provided a mechanism for Mendel’s laws of inheritance, and paved the ground for modern genetics. It enabled numerous discoveries and new technologies, adding to our knowledge...In 1926, Thomas Hunt Morgan published , which provided a mechanism for Mendel’s laws of inheritance, and paved the ground for modern genetics. It enabled numerous discoveries and new technologies, adding to our knowledge and putting more nuance to Morgan’s original theory. [Image: see text]
Ribonucleoprotein (RNP) granules are dynamic, membraneless organelles that sequester translationally repressed mRNAs and RNA-binding proteins, playing a pivotal role in the regulation of localized protein synthesis. Whil...Ribonucleoprotein (RNP) granules are dynamic, membraneless organelles that sequester translationally repressed mRNAs and RNA-binding proteins, playing a pivotal role in the regulation of localized protein synthesis. While disassembly of RNP granules is essential for reactivating translation, the mechanisms by which neuronal activity regulates this process remain poorly understood. In this study, we show that stimulation of N-methyl-D-aspartate (NMDA) receptor (NMDAR) triggers calcium influx, leading to activation of calpain 1 and subsequent proteolytic cleavage of Ras-GTPase-activating protein binding protein 1 (G3BP1), a core component of stress granules. This cleavage results in the disassembly of G3BP1 granules in the neurites and promotes mTOR-dependent local translation, thereby linking synaptic activity to spatially restricted protein synthesis. Finally, we demonstrate that the NMDAR-calpain 1-G3BP1-mTOR signaling axis contributes to axonal regeneration, establishing proteolytic remodeling of RNP granules as a key mechanism of activity-dependent neural repair.
Comment on “Reevaluating the Senolytic Activity of a GLS1 Inhibitor and an Anti-PD-1 Antibody: Toward Greater Reproducibility and Methodological Rigor” by Kawamoto et al. [Image: see text]Comment on “Reevaluating the Senolytic Activity of a GLS1 Inhibitor and an Anti-PD-1 Antibody: Toward Greater Reproducibility and Methodological Rigor” by Kawamoto et al. [Image: see text]