eLife最新文献

The nuclear receptor peroxisome proliferator-activated receptor gamma (PPARγ) regulates transcription in response to ligand binding at an orthosteric pocket within the ligand-binding domain (LBD). We previously showed that two covalent ligands, T0070907 and GW9662-extensively used as PPARγ inhibitors to assess off-target activity-weaken but do not completely block ligand binding via an allosteric mechanism associated with pharmacological inverse agonism (Shang and Kojetin, 2024). These covalent inhibitors shift the LBD toward a repressive conformation, where the activation function-2 (AF-2) helix 12 occupies the orthosteric pocket, competing with orthosteric ligand binding. Here, we provide additional support for this allosteric mechanism using two covalent inverse agonists, SR33065 and SR36708, which better stabilize the repressive LBD conformation and are more effective inhibitors of-but also do not completely inhibit-ligand cobinding. Furthermore, we show that ligand cobinding can occur with a previously reported PPARγ dual-site covalent inhibitor, SR16832, which appears to weaken ligand binding through a direct mechanism independent of the allosteric mechanism. These findings underscore the complex nature of the PPARγ LBD conformational ensemble and highlight the need to develop alternative methods for designing more effective covalent inhibitors.

Trained immunity (TI) refers to a state of innate immune cells that, after encountering an initial stimulus and undergoing epigenetic reprogramming and metabolic changes, allows them to respond more effectively to a subsequent challenge. TI yields a survival advantage, particularly in a pathogen-rich context. However, maladaptive TI may damage the host by exacerbating inflammatory diseases. Here we review which aspects of Western lifestyle may contribute to maladaptive TI, including a Western diet, periodontitis, chronic psychological stress, and environmental triggers such as air pollution and microplastics. Finally, we consider lifestyle intervention as a way to prevent or reduce the impact of maladaptive TI.

In lower vertebrates, retinal Müller glia (MG) exhibit a life-long capacity of cell-cycle re-entry to regenerate neurons following the retinal injury. However, the mechanism driving such injury-induced MG cell-cycle re-entry remains incompletely understood. Combining single-cell transcriptomic analysis and in vivo clonal analysis, we identified previously undescribed cxcl18b-defined MG transitional states as essential routes toward MG proliferation following green/red cone (G/R cone) ablation. Inflammation blockage abolished the triggering of these transitional states, which expressed the gene modules shared by cells of the ciliary marginal zone (CMZ), where life-long adult neurogenesis takes place. Functional studies of the redox properties of these transitional states further demonstrated the regulatory role of nitric oxide (NO) produced by Nos2b in injury-induced MG proliferation. Finally, we developed a viral-based strategy to specifically disrupt nos2b in cxcl18b-defined MG transitional states and revealed the effect of transitional state-specific NO signaling. Our findings elucidate the precision redox mechanism underlying injury-induced MG cell-cycle re-entry, providing insights into species-specific mechanisms for vertebrate retina regeneration.

ATAD2, a conserved protein which is predominantly expressed in embryonic stem (ES) cells and spermatogenic cells, emerges as a crucial regulator of chromatin plasticity. Our previous parallel studies conducted in both ES cells and S. pombe highlighted the fundamental role of ATAD2 in facilitating chromatin-bound histone chaperone turnover. Focusing on mouse spermatogenesis, we demonstrate here that ATAD2 regulates the HIRA-dependent localization of H3.3 on the genome and influences H3.3-mediated gene transcription. Moreover, by modulating histone eviction and the assembly of protamines, ATAD2 ensures proper chromatin condensation and genome packaging in mature sperm. Disruption of Atad2 function in mice leads to abnormal genome organization in mature spermatozoa. Together, these findings establish a previously overlooked level of chromatin dynamic regulation, governed by ATAD2-controlled histone chaperones binding to chromatin, which defines the balance between histone deposition and removal.

Osteoarthritis (OA) is a painful, debilitating disease with no cure or treatments that can predictably stop/reverse its progression. Treatment is particularly difficult since articular cartilage lacks intrinsic repair capacity, despite mesenchymal stem cells (MSCs) being present in the joint with robust chondrogenic potential. While heterogeneity exists among MSC subtypes within human synovium, it remains unclear which populations can regenerate cartilage or impact OA progression. We clonally isolated MSCs from normal and OA patient synovium using indexed flow cytometry, then characterized them through differentiation assays and quantitative proteomics. MSC clones were transplanted into a xenograft rat OA model and evaluated by histology and immunofluorescence. We identified heterogeneity in putative MSCs within and between patient groups and their repair capacity in the rat model. However, traditional cell surface markers could not distinguish these subtypes, highlighting the need for single-cell level understanding. Using unbiased proteomics, we identified CD47 as a novel MSC marker. CD47^Hi cells demonstrated robust treatment efficacy in the rat OA model and directly contributed to new articular cartilage formation. Characterizing MSC subtypes is essential for identifying candidates appropriate for clinical investigation and exploiting functional MSCs for cartilage regeneration strategies.

The function of the neocortex is fundamentally determined by its repeating microcircuit motif, but also by its rich, interregional connectivity. We present a data-driven computational model of the anatomy of non-barrel primary somatosensory cortex of juvenile rat, integrating whole-brain scale data while providing cellular and subcellular specificity. The model consists of 4.2 million morphologically detailed neurons, placed in a digital brain atlas. They are connected by 14.2 billion synapses, comprising local, mid-range and extrinsic connectivity. We delineated the limits of determining connectivity from neuron morphology and placement, finding that it reproduces targeting by Sst+ neurons, but requires additional specificity to reproduce targeting by PV+ and VIP+ interneurons. Globally, connectivity was characterized by local clusters tied together through hub neurons in layer 5, demonstrating how local and interegional connectivity are complicit, inseparable networks. The model is suitable for simulation-based studies, and the model is made openly available to the community.

Cortical dynamics underlie many cognitive processes and emerge from complex multiscale interactions, which are challenging to study in vivo. Large-scale, biophysically detailed models offer a tool that can complement laboratory approaches. We present a model comprising eight somatosensory cortex subregions, 4.2 million morphological and electrically detailed neurons, and 13.2 billion local and mid-range synapses. In silico tools enabled reproduction and extension of complex laboratory experiments under a single parameterization, providing strong validation. The model reproduced millisecond-precise stimulus-responses, stimulus-encoding under targeted optogenetic activation, and selective propagation of stimulus-evoked activity to downstream areas. The model's direct correspondence with biology generated predictions about how multiscale organization shapes activity; for example, how cortical activity is shaped by high-dimensional connectivity motifs in local and mid-range connectivity, and spatial targeting rules by inhibitory subpopulations. The latter was facilitated using a rewired connectome that included specific targeting rules observed for different inhibitory neuron types in electron microscopy. The model also predicted the role of inhibitory interneuron types and different layers in stimulus encoding. Simulation tools and a large subvolume of the model are made available to enable further community-driven improvement, validation, and investigation.

Since the trabecular meshwork (TM) is central to intraocular pressure (IOP) regulation and glaucoma, a deeper understanding of its genomic landscape is needed. We present a multimodal, single-cell resolution analysis of mouse limbal cells (includes TM). In total, we sequenced 9,394 wild-type TM cell transcriptomes. We discovered three TM cell subtypes with characteristic signature genes validated by immunofluorescence on tissue sections and whole-mounts. The subtypes are robust, being detected in datasets for two diverse mouse strains and in independent data from two institutions. Results show compartmentalized enrichment of critical pathways in specific TM cell subtypes. Distinctive signatures include increased expression of genes responsible for (1) extracellular matrix structure and metabolism (TM1 subtype), (2) secreted ligand signaling to support Schlemm's canal cells (TM2), and (3) contractile and mitochondrial/metabolic activity (TM3). ATAC-sequencing data identified active transcription factors in TM cells, including LMX1B. Mutations in LMX1B cause high IOP and glaucoma. LMX1B is emerging as a key transcription factor for normal mitochondrial function, and its expression is much higher in TM3 cells than other limbal cells. To understand the role of LMX1B in TM function and glaucoma, we single-cell sequenced limbal cells from Lmx1b^V265D/+ mutant mice (2491 TM cells). In Lmx1b^V265D/+ mice, TM3 cells were uniquely affected by pronounced mitochondrial pathway changes. Mitochondria in TM cells of Lmx1b^V265D/+ mice are swollen with a reduced cristae area, further supporting a role for mitochondrial dysfunction in the initiation of IOP elevation in these mice. Importantly, treatment with vitamin B3 (nicotinamide), which enhances mitochondrial function and metabolic resilience in other contexts, significantly protected Lmx1b mutant mice from IOP elevation.

Proteomics experiments on Drosophila reveal sex-specific effects in aging, and an important role for a protein called DIP-β.

Mitotic surveillance pathways monitor the duration of mitosis (M phase) in the cell cycle. Prolonged M phase, caused by spindle attachment defects or microtubule-targeting drugs, triggers formation of the ternary 'mitotic stopwatch pathway' complex (MSP) consisting of 53BP1, USP28, and p53. This complex stabilizes p53, leading to cell cycle arrest or apoptosis in daughter cells. In cancers that are resistant to paclitaxel, a microtubule-targeting agent, cells bypass mitotic surveillance activation, allowing unchecked proliferation, although the underlying mechanisms remain poorly understood. Here, we identify GMCL1 as a key negative regulator of MSP signaling. We show that 53BP1 physically interacts with GMCL1, but not its paralog GMCL2, and we map their interaction domains. CRL3^GMCL1 functions as a ubiquitin ligase that targets 53BP1 for degradation during the M phase, thereby reducing p53 accumulation in daughter cells. Depletion of GMCL1 inhibits cell cycle progression upon release from prolonged mitotic arrest, a defect that is rescued by co-silencing 53BP1 or USP28. Moreover, GMCL1 depletion sensitizes cancer cells to paclitaxel in a p53-dependent manner. Together, our findings support a model in which dysregulated CRL3^GMCL1-mediated degradation of 53BP1 prevents proper MSP function, leading to p53 degradation and continued proliferation. Targeting GMCL1 may, therefore, represent one possible avenue for addressing paclitaxel resistance in cancer cells with functional p53.