bioRxiv : the preprint server for biology最新文献

From smooth to buckled, nature exhibits organs of various shapes and forms. How cellular growth patterns produce smooth organ shapes such as leaves and sepals remains unclear. Here we show that unidirectional growth and comparable stiffness across both epidermal layers of Arabidopsis sepals are essential for smoothness. We identified a mutant with ectopic ASYMMETRIC LEAVES 2 (AS2) expression on the outer epidermis. Our analysis reveals that ectopic AS2 expression causes outer epidermal buckling at early stages of sepal development, due to conflicting growth directions and unequal epidermal stiffnesses. Aligning growth direction and increasing stiffness of the outer epidermis restores smoothness. Furthermore, buckling influences auxin efflux transporter protein PIN-FORMED 1 polarity to generate outgrowth in the later stages, suggesting that buckling is sufficient to initiate outgrowths. Our findings suggest that in addition to molecular cues influencing tissue mechanics, tissue mechanics can also modulate molecular signals, giving rise to well-defined shapes.

Biological function is mediated by the hierarchical organization of cell types and states within tissue ecosystems. Identifying interpretable composite marker sets that both define and distinguish hierarchical cell identities is essential for decoding biological complexity, yet remains a major challenge. Here, we present RECOMBINE, an algorithm that identifies recurrent composite marker sets to define hierarchical cell identities. Validation using both simulated and biological datasets demonstrates that RECOMBINE achieves higher accuracy in identifying discriminative markers compared to existing approaches, including differential gene expression analysis. When applied to single-cell data and validated with spatial transcriptomics data from the mouse visual cortex, RECOMBINE identified key cell type markers and generated a robust gene panel for targeted spatial profiling. It also uncovered markers of CD8+; T cell states, including GZMK+;HAVCR2-; effector memory cells associated with anti-PD-1 therapy response, and revealed a rare intestinal subpopulation with composite markers in mice. Finally, using data from the Tabula Sapiens project, RECOMBINE identified composite marker sets across a broad range of human tissues. Together, these results highlight RECOMBINE as a robust, data-driven framework for optimized marker selection, enabling the discovery and validation of hierarchical cell identities across diverse tissue contexts.

The circadian clock attunes metabolism to daily energy cycles, but how it regulates maturation of metabolic tissues is poorly understood. Here we show that DEC1, a clock transcription factor induced in adult islet β cells, coordinates their glucose responsiveness by synchronizing energetic and secretory rhythms. DEC1 binds and regulates maturity-linked genes to integrate insulin exocytosis with energy metabolism, and β-cell Dec1 ablation disrupts their transcription synchrony. Dec1-disrupted mice develop lifelong glucose intolerance and insulin deficiency, despite normal islet formation and intact Clock/Bmal1 genes. Metabolic dysfunction upon β-cell Dec1 loss stems from poor coupling of insulin secretion to glucose metabolism, reminiscent of fetal/neonatal immaturity. We link stunted maturation to a deficit in circadian bioenergetics, prompted by compromised glucose utilization, mitochondrial dynamics, and respiratory metabolism, which is rescued by increased metabolic flux. Thus, DEC1 links circadian clockwork to β-cell metabolic maturation, revealing a hierarchy for how the clock programs metabolic tissue specialization.

Mycobacterium tuberculosis (Mtb) causes 1.25 million deaths a year. However, tuberculosis (TB) pathogenesis remains poorly understood and is not fully recapitulated in standard mouse models. Here we find that gene signatures from three different Mtb-susceptible mouse models predict active TB disease in humans significantly better than a signature from resistant C57BL/6 (B6) mice. Conserved among susceptible mice, non-human primates, and humans, but largely absent from B6 mice, was Mtb-induced differentiation of macrophages into an Spp1+ differentiation state. Spp1+ macrophages expressed high levels of immunosuppressive molecules including IL-1 receptor antagonist (IL-1Ra). IL-1Ra was previously reported to cause Mtb susceptibility in one mouse model, but whether IL-1Ra is broadly important remains uncertain. Here we report that enhancement of IL-1 signaling via deletion of IL-Ra promoted bacterial control across three susceptible mouse models. We found IL-1 signaling amplified production of multiple cytokines by lymphoid and stromal cells, providing a multifactorial mechanism for how IL-1 promotes Mtb control. Our results indicate that myeloid cell expression of immunosuppressive molecules, in particular IL-1 receptor antagonist, is a conserved early mechanism limiting Mtb control in mice, non-human primates, and humans.

Apurinic/apyrimidinic endonuclease I (APE1) acts as both an endonuclease and a redox factor to ensure cell survival. The two activities require different conformations of APE1. As an endonuclease, APE1 is fully folded. As a redox factor, APE1 must be partially unfolded to expose the buried residue Cys65, which reduces transcription factors including AP-1, NF-κB, and HIF-1α and thereby enables them to bind DNA. To determine a molecular basis for partial unfolding associated with APE1's redox activity, we characterized specific interactions of a known redox inhibitor APX3330 with APE1 through waterLOGSY and ¹ H- ¹⁵ N HSQC NMR approaches using ethanol and acetonitrile as co-solvents. We find that APX3330 binds to the endonuclease active site in both co-solvents and to a distant small pocket in acetonitrile. Prolonged exposure of APE1 with APX3330 in acetonitrile resulted in a time-dependent loss of ¹ H- ¹⁵ N HSQC chemical shifts (∼35%), consistent with partial unfolding. Regions that are partially unfolded include adjacent N- and C-terminal beta strands within one of the two sheets comprising the core, which converge within the small binding pocket defined by the CSPs. Removal of APX3330 via dialysis resulted in a slow reappearance of the ¹ H- ¹⁵ N HSQC chemical shifts suggesting that the effect of APX3330 is reversible. APX3330 significantly decreases the melting temperature of APE1 but has no effect on endonuclease activity using a standard assay in either co-solvent. Our results provide insights on reversible partial unfolding of APE1 relevant for its redox function as well as the mechanism of redox inhibition by APX3330.

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During aging, adipose tissue within the bone marrow expands while the trabecular red marrow contracts. The impact of these changes on blood cell formation remains unclear. To address this question, we performed single-cell and single-nuclei transcriptomic analysis on adipose-rich yellow bone marrow (BMY) and adipose-poor trabecular red marrow (BMR) from human subjects undergoing lower limb amputations. Surprisingly, we discovered two distinct hematopoietic niches, in which BMY contains a higher number of monocytes and progenitor cells expressing genes associated with inflammation. To further investigate these niches, we developed an in-vitro organoid system that maintains features of the human bone marrow. We find cells from BMY are distinct in their expression of the leptin receptor, and respond to leptin stimulation with enhanced proliferation, leading to increased monocyte production. These findings suggest that the age-associated expansion of bone marrow adipose tissue drives a pro-inflammatory state by stimulating monocyte production from a spatially distinct, leptin-responsive hematopoietic stem/progenitor cell population.

Significance: This study reveals that adipose tissue within the human bone marrow is a niche for hematopoietic stem and progenitor cells that can give rise to pro-inflammatory monocytes through leptin signaling. Expansion of bone marrow adipose tissue with age and stress may thus underlie inflammageing.

Each new human has an expected U _d = 2-10 new deleterious mutations. Using a novel approach to capture complex linkage disequilibria from high U _d using genome-wide simulations, we confirm that fitness decline due to the fixation of many slightly deleterious mutations can be compensated by rarer beneficial mutations of larger effect. The evolution of increased genome size and complexity have previously been attributed to a similarly asymmetric pattern of fixations, but we propose that the cause might be high U _d rather than the small population size posited as causal by drift barrier theory. High within-population variance in relative fitness is an inevitable consequence of high U _d ∼2-10 combined with inferred human deleterious effect sizes; two individuals will typically differ in fitness by 15-40%. The need to compensate for the deluge of deleterious mutations slows net adaptation (i.e. to the external environment) by ∼13%-55%. The rate of beneficial fixations is more sensitive to changes in the mutation rate than the rate of deleterious fixations is. As a surprising consequence of this, an increase (e.g. 10%) in overall mutation rate leads to faster adaptation; this puts to rest dysgenic fears about increasing mutation rates due to rising paternal age.

Humans continuously adapt locomotor patterns. Whether energetic cost reduction is the primary objective or a by-product of locomotor adaptation is not known. If energetic cost is the primary objective, then manipulating energetic cost will affect the locomotor pattern. Our study aims to determine if information about task duration affects energetic cost and locomotor adaptation during split-belt walking. We hypothesize that information about a longer adaptation duration will result in lower metabolic costs and lower mechanical work. N=52 participants walked for 10 minutes with the belts moving at 1.5 and 0.5 m/s, followed by 6 minutes of walking with both belts at 1.0 m/s. Nineteen participants walked on the split-belt while we provided True information about time remaining every minute (Group T). Nineteen participants received False information that split-belt adaptation duration was around 30 minutes (Group F). Fourteen participants walked on a split-belt with accurate information about task duration, and one update at 5 minutes remaining (Group C). Participants in Groups C and F had a lower rate of change in metabolic cost from baseline (p=0.002) and generated less positive work (p=0.012) than individuals in Group T. Changes in positive work by the fast leg predicted metabolic cost reductions only in Group F (R ² =0.18, p=0.040). Participants in Group F showed greater split-belt aftereffects than the C and T groups (p<0.001). We conclude that walking biomechanics are adapted to support an energetic cost reduction when maintaining an energetic reserve is needed, as is the case for Group F, but not Group T.

New and noteworthy: The relationship between walking biomechanics and energetics can be modulated to maintain an energetic reserve during a novel locomotor adaptation task when individuals believe they must sustain a task for a prolonged period. When an energetic reserve is not required, individuals can use more energy than what is required for the task. Planning to sustain the adapted locomotor pattern for a prolonged time increases the aftereffects of locomotor adaptation.

We present the nELISA, a high-throughput, high-fidelity, and high-plex protein profiling platform. DNA oligonucleotides are used to pre-assemble antibody pairs on spectrally encoded microparticles and perform displacement-mediated detection. Spatial separation between non-cognate antibodies prevents the rise of reagent-driven cross-reactivity, while read-out is performed cost-efficiently and at high-throughput using flow cytometry. nELISA can measure both protein concentration and their post-translational modifications. We assembled an inflammatory panel of 191 targets that were multiplexed without cross-reactivity nor impact on performance vs 1-plex signals, with sensitivities as low as 0.1 pg/mL and measurements spanning 7 orders of magnitude. We then performed a large-scale inflammatory-secretome perturbation screen of peripheral blood mononuclear cells (PBMCs), with cytokines as both perturbagens and read-outs, measuring 7,392 samples and generating ∼1.4M protein data points in under a week; a significant advance in throughput compared to other highly multiplexed immunoassays. We uncovered 447 significant cytokine responses, including multiple putatively novel ones, that were conserved across donors and stimulation conditions. We validate nELISA for phenotypic screening, where its capacity to faithfully report hundreds of proteins make it a powerful tool across multiple stages of drug discovery.

The auditory brainstem response (ABR) is an acoustically evoked EEG potential that is an important diagnostic tool for hearing loss, especially in newborns. The ABR originates from the response sequence of auditory nerve and brainstem nuclei, and a click-evoked ABR typically shows three positive peaks ('waves') within the first six milliseconds. However, an assignment of the waves of the ABR to specific sources is difficult, and a quantification of contributions to the ABR waves is not available. Here, we exploit the large size and physical separation of the barn owl first-order cochlear nucleus magnocellularis (NM) to estimate single-cell contributions to the ABR. We simultaneously recorded NM neurons' spikes and the EEG in owls of both sexes, and found that ≳ 5,000 spontaneous single-cell spikes are necessary to isolate a significant spike-triggered average response at the EEG electrode. An average single-neuron contribution to the ABR was predicted by convolving the spike-triggered average with the cell's peri-stimulus time histogram. Amplitudes of predicted contributions of single NM cells typically reached 32.9 ± 1.1 nV (mean ± SE, range: 2.5 - 162.7 nV), or 0.07 ± 0.02% (median ± SE; range from 0.01% to 1%) of the ABR amplitude. The time of the predicted peak coincided best with the peak of the ABR wave II, independent of the click sound level. Our results suggest that individual neurons' contributions to an EEG can vary widely, and that wave II of the ABR is shaped by NM units.

Significance statement: The auditory brainstem response (ABR) is a scalp potential used for the diagnosis of hearing loss, both clinically and in research. We investigated the contribution of single action potentials from auditory brainstem neurons to the ABR and provide direct evidence that action potentials recorded in a first order auditory nucleus, and their EEG contribution, coincide with wave II of the ABR. The study also shows that the contribution of single cells varies strongly across the population.