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Search indexed bioRxiv preprints in genomics, neuroscience, cell biology and bioinformatics. Read source abstracts and check manuscript versions; preprints are not peer reviewed.

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bioRxiv2025

Enhancing salt tolerance in a hybrid poplar (Populus nigra x maximowiczii) through exogenous putrescine application: Growth, physiological, and biochemical responses

IntroductionPutrescine, a polyamine involved in plant growth and stress responses, has shown potential in mitigating abiotic stress effects. However, little is known about the exogenous addition of putrescine regarding salt tolerance in trees. MethodsThis study was conducted to investigate whether exogenous putrescine application via foliar spray enhances growth in a hybrid poplar (Populus nigra x maximowiczii, clone NM6) under a short duration of salt stress. Salt stress was induced by irrigating roots with 100 mM and 200 mM NaCl, followed by foliar spraying of putrescine on several days. Measurement of growth including plant height and stem diameter for each plant were recorded in the greenhouse every 15 days throughout the experiment. Gas exchange, total chlorophyll, carotenoids, soluble sugars and proteins, amino acids, polyamines, and relative water content were analyzed in foliage collected 3, 6, 7, 13, 20, 35 days after treatment. ResultsPutrescine spray on the salt-treated plants caused a significant increase in plant growth. Putrescine application caused a significant increase in fructose, glucose, and galactose in all plants, but putrescine spray had a variable impact on the sucrose content of 100 mM NaCl-treated plants. Based on metabolic responses, plants treated with 100 mM NaCl fared better when sprayed with putrescine than those treated with 200 mM NaCl. DiscussionExogenous application of putrescine mitigated growth inhibition effects of salinity. These finding highlight the potential of putrescine as a practical approach to increase salt tolerance in young poplar trees, with implications for forestry and land reclamation in saline environment.

plant biology↗

bioRxiv2025

iSleep: Continuous, binocular pupil tracking in sleep and reduced consciousness for physiological monitoring, predictions and interventions

Monitoring pupil dynamics is a key tool in understanding arousal. Pupil size can serve as a biomarker for the autonomic nervous system balance as well as for identifying brain states. While internal states can also be self-reported when awake, automated detection and non-invasive monitoring is crucial during sleep and reduced consciousness. Here, we introduce iSleep, an innovative pupil tracking and analysis framework for sleep in humans. It features comfortable, humidified eye-tracking goggles and a platform for integrated analysis and prediction capabilities. We show that iSleep allows safe and continuous access to binocular pupil size and ocular dynamics during sleep and anesthesia. iSleep reveals that pupillary fluctuations correlate tightly with brain activity, heartbeat, and breathing, and can reliably predict brain states. Pupil constrictions reflect parasympathetic drive and likely serve a protective function for deep sleep stability; while dilations indicate arousals. Unexpectedly, we observed a decoupling of binocular movements during periods of sleep, indicating alterations in reflexes which usually govern voluntary eye movements. Finally, iSleep was tested in surgery patients under general anesthesia, revealing dynamic pupil changes to noxious stimuli, suggesting the potential for nociception monitoring during surgeries. In summary, iSleep offers an easy-to-use, robust alternative to read out brain states during sleep and anesthesia, opening new avenues in monitoring, diagnostics, and treatments, previously obscured by closed eyelids.

neuroscience↗

bioRxiv2025

Human emotional odours influence horses' behaviour and physiology

Olfaction is the most widespread sensory modality animals use to communicate, yet much remains to be discovered about its role. While most studies focused on intraspecific interactions and reproduction, new evidence suggests chemosignals may influence interspecific interactions and emotional communication. This study explores this possibility, investigating olfactory signals in emotional contagion from the example of human-horse interactions. Cotton pads carrying human odours from fear and joy contexts, or unused pads (control odour) were applied to 43 horses nostrils during fear tests (suddenness and novelty tests) and human interaction tests (grooming and approach tests). Principal component analysis showed that overall, when exposed to fear-related human odours, horses exhibited significantly heightened fear responses and reduced interaction with humans compared to joy-related and control odours. More precisely, when exposed to fear-related odours, horses touched the human less in the human approach test, gazed more at the novel object, and were more startled (startle intensity and maximum heart rate) by a sudden event. These results highlight the significance of chemosignals in interspecific interactions and provides insights into questions about the impact of domestication on emotional communication.

animal behavior and cognition↗

bioRxiv2025

An open-source pipeline for calcium imaging and all-optical physiology in human stem cell-derived neurons

High-throughput profiling of neuronal activity at single-cell resolution is essential for advancing our understanding of brain function, enabling large-scale functional screens, and modeling neurological disorders. However, existing approaches are limited by scalability, manual data processing, and variability, thus restricting their ability to detect disease-associated phenotypes. Here, we present a scalable, open-source platform that integrates optogenetic stimulation, calcium imaging, automated data acquisition, and a fully integrated analysis pipeline. By combining spontaneous and evoked activity profiling, the system enables robust quantification of dynamic neuronal responses across hundreds of stem cell-derived human neurons and multiple timepoints, facilitating phenotyping at both cellular and network levels. We validated the platform by recapitulating established activity phenotypes in neurodevelopmental disorders including CDKL5 Deficiency Disorder and SSADH deficiency. In addition, we generated CRISPR-Cas9 knock-in human induced pluripotent stem cell (hiPSC) lines stably expressing the genetically encoded calcium indicator GCaMP6s to model network dysfunction in Tuberous Sclerosis Complex (TSC). Using this system, we further demonstrated functional rescue of the altered neuronal activity observed in the TSC following pharmacological intervention. By linking single-cell dynamics to population-level phenotypes, this framework provides a powerful and broadly applicable tool for disease modeling, mechanistic studies, and therapeutic screening across a range of neurological disorders.

neuroscience↗

bioRxiv2025

Microfluidic Platform for Automatic Quantification of Malaria Parasite Invasion Under Physiological Flow Conditions

Understanding the impact of forces generated by blood flow on biological processes in the circulatory system, such as the invasion of human red blood cells by malaria parasites, is currently limited by the lack of experimental systems that integrate them. Recent systematic quantification of the growth of Plasmodium falciparum, the species that causes the majority of malaria mortality, under a range of shaking conditions has shown that parasite invasion of erythrocytes is affected by the shear stress to which the interacting P. falciparum merozoites and their target red blood cells are exposed. Blood flow could similarly impact shear stress and therefore invasion in vivo, but there is currently no method to test the impact of flow-induced forces on parasite invasion. We have developed a microfluidic device with four channels, each with dimensions similar to those of a post-capillary venule, but with different flow velocities. Highly synchronised P. falciparum parasites are injected into the device, and parasite egress and invasion rates are quantified using newly developed custom video analysis, which fully automates cell type identification and trajectory tracking. The device was tested with both wild-type P. falciparum lines and lines in which genes encoding proteins involved in parasite invasion had been deleted. Deletion of Erythrocyte Binding Antigen 175 (PfEBA175) has a significant impact on invasion under flow, but not in static culture. These findings establish for the first time that flow conditions can critically affect parasite invasion in a genotype-dependent manner. The method can be applied to other biological processes affected by fluid motion, such as cell adhesion, migration, and mechanotransduction.

microbiology↗

bioRxiv2025

Root vulnerability to embolism and lack of physiological recovery limit the competitive ability of an invasive palm

O_LIClimate change and invasive species both threaten forest ecosystems. While exacerbated droughts drive tree mortality, species invasion alters forest composition. Invasive plants may be more vulnerable to drought due to acquisitive traits and low embolism resistance, but their potentially superior recovery capacity raises key uncertainties in predicting future species distributions. C_LIO_LIWe compared the drought resistance and recovery of an invasive palm (Trachycarpus fortunei) with two native species (Ilex aquifolium & Tilia cordata) from the Southern Alps. Saplings were exposed to increasing intensities of drought-induced xylem embolism, then rewatered for 45 days. Among others, we tracked net assimilation (Anet), stomatal conductance (gs), and leaf water potential ({Psi}leaf) before and after the drought, and modeled the time to hydraulic failure in leaves, stems, and roots. C_LIO_LIWhile the invasive palm had exceptionally drought-tolerant leaves, its roots were the most drought-sensitive among all species. The palm maintained positive gas exchange until {Psi}leaf of -4 MPa, as some native species; however, it failed to recover from the most severe drought, with simulations showing root embolism preceding leaf failure, unlike native species. C_LIO_LIInvasive plants may match or exceed natives drought resistance, but their inability to recover from extreme droughts could limit their future spread. C_LI

ecology↗

bioRxiv2025

Uptake mechanisms and physiological effects of furanic compounds from the Maillard reaction in budding yeast

Maillard reaction products (MRPs) are formed during the thermal processing of foods and exhibit important sensory attributes. Furanic compounds are a subset of MRPs commonly found in food products that are toxic to eukarytoic cells, although the mechanisms of toxicity are poorly understood. We used budding yeast to explore uptake mechanisms of common furanic compounds: 5-hydroxymethylfurfural (HMF), furfural (FUR), and 2-Furyl methyl ketone (FMK). Titrations of each furanic compound were used to identify concentrations that have an inhibitory effect on growth. We identified HMF as a potential substrate of the Pdr5 multidrug resistance pump and linked HMF and FUR toxicity to surface nutrient transporter levels. Live cell imaging shows that HMF disrupts mitochondria whilst FUR affects the endolysosomal system. Results indicate these furanic compounds may have distinct uptake, efflux, and toxicity mechanisms. As many of these cellular components are conserved throughout evolution, this work could shed light on the metabolism of toxic compounds commonly found within animal food sources.

cell biology↗

bioRxiv2025

Paranemic cohesion of DNA molecules in different counter ions at room and physiological temperatures

Isothermal assembly allows the construction of DNA nanostructures at constant moderate temperatures. Further, assembly of DNA nanostructures in different counter ions expands their utility in various applications. In this work, we demonstrate the isothermal assembly of paranemic crossover (PX) DNA motifs in magnesium (Mg2+), calcium (Ca2+) and strontium (Sr2+) at 20 {degrees}C and 37 {degrees}C. Further, we study how differences in design affect the assembly of 4-stranded and 2-stranded PX molecules.

synthetic biology↗

bioRxiv2025

Pre-Sensory Spontaneous Activity Accelerates Coordinated Maturation of Synaptic Partners and Drives Transition to the Mature Physiological Phenotype

Pre-sensory (ps), peripherally-generated spontaneous activity (SA) is important for establishing basic topography of central components of sensory pathways. However, roles for psSA in sculpting connections at the single neuron level and in driving functional maturation of postsynaptic targets are difficult to isolate and therefore little explored. We capitalized on the temporal onset of cochlea-generated psSA just prior to growth of the calyx of Held (CH), the largest nerve terminal in the mammalian brain, and its well-defined circuit topography to explore the causal roles for psSA in targeted synaptogenesis and postsynaptic functional maturation. To this end, we developed a viral vector to rapidly express tetanus neurotoxin (TeNT), which blocks neurotransmission, and selectively introduced it into CH-forming neurons at the onset of psSA. TeNT expression did not prevent but delayed and altered the developmental trajectory of neural circuit maturation, which in sum prevented the postsynaptic neuron from acquiring its mature phasic firing phenotype. Thus, psSA initiates and drives circuit maturation along central sensory pathways, but alternative mechanisms can partially compensate in its absence.

neuroscience↗

bioRxiv2025

Butterfly egg plasticity in physiologically and reproductive relevant traits: adaptation to climate?

Embryos of terrestrial animals developing inside immobile eggs face variable environmental conditions. Insect egg pores, critical for gas exchange (aeropyles) and fertilisation (micropyles), are ecologically important yet understudied. While aeropyles likely adapt to climatic respiratory demands, and micropyles may respond to environmental pressures, their inter- and intra-specific variation remains poorly studied. Here, we compared egg pore morphology of three closely related Pieris butterflies sampled in the Netherlands. In addition, we compared egg traits of different P. napi populations sampled along a 4,000-km latitudinal gradient from northern to southern Europe linked these traits with available climatic data. After several generations under identical greenhouse conditions, egg pore traits remained highly variable within and between species and groups. Remarkably, P. napi populations from divergent climates differed more in pore number and size than did co-occurring Pieris species. Aeropyle number and width were strongly associated with climatic variables such as maximum temperature, independent of egg size. Micropyle width varied with seasonality. Additionally, we observed the micropylar pit of older eggs covered by a substance more often than fresh eggs. Our findings indicate that insect egg pores are highly dynamic traits, shaped by ecological, behavioural and climatic factors and an under-recognised layer of egg adaptation.

ecology↗

bioRxiv2025

Comparative genome analysis of tea rhizosphere dwelling Paenarthrobacter nicotinovorans AB444: Uncovering Physiological Resilience and Ecological Adaptability

Plant growth-promoting rhizobacteria (PGPR) can be considered a promising tool for sustainable agriculture. The recent scenario of growing demand for higher crop yields, with declining soil fertility and impacts of climate change, promotes the global shift towards eco-friendly agricultural practices, emphasising the significance of PGPR as a sustainable solution. In this study, a PGPR Paenarthrobacter sp. AB444 was isolated from the rhizosphere of tea plants at the Longview and Makaibari tea estates in the Darjeeling District of West Bengal. Based on the draft genome sequence along with comprehensive chemotaxonomic analyses, strain AB444 was identified as Paenarthrobacter nicotinovorans. The pangenomic analysis with phylogenetically close strains revealed the gene clusters that are common and unique to the isolate. The plant growth-promoting attributes of AB444 were explored with different assays like phosphate solubilisation, IAA and siderophore production, ACC deaminase, nitrogen fixation, and protease activity. The colonization of the isolate to the plant root and in-planta growth were also assessed. The presence of biosynthetic gene clusters of AB444 related to plant growth was also extensively studied, highlighting the unmapped disposition of Paenarthrobacter nicotinovorans.

microbiology↗

bioRxiv2025

Endogenous corazonin signaling modulates the post-mating switch in behavior and physiology in females of the brown planthopper and Drosophila

Mating in insects typically triggers a post-mating response (PMR) in females, characterized by reduced receptivity to re-mating and increased oviposition, which ensures numerous and viable offspring and male paternity. This PMR is induced by male seminal factors, such as sex peptide in Drosophila melanogaster, as well as intrinsic female signaling components. The latter signaling remains poorly understood in most insects, including the devastating rice pest, the brown planthopper (BPH) Nilaparvata lugens. Here, we show that the neuropeptide corazonin (CRZ) and its receptor (CrzR) are critical for the PMR in female BPHs. Peptide injection, RNAi knockdown, and CRISPR/Cas9 mutagenesis confirm that intact CRZ signaling reduces re-mating frequency and increases ovulation in mated BPH females. The CrzR is highly expressed in the female reproductive tract, and CrzR knockdown phenocopies CRZ diminishment. Importantly, female CRZ/CrzR signaling is required for male seminal factors, such as the peptide maccessin, to induce the PMR; with disrupted CrzR signaling, injection of seminal fluid or maccessin fails to reduce female receptivity. Notably, CRZ is not produced in male accessory glands (MAGs) of BPHs and thus not transferred during copulation. We furthermore demonstrate that also in D. melanogaster disrupted CRZ signaling increases female re-mating and reduces oviposition, while CRZ injection suppresses virgin receptivity and increases oviposition. Finally, we detected no CRZ in the MAG of D. melanogaster, supporting its role as an endogenous signal in the female PMR also in this species. In summary, our findings reveal a conserved role of endogenous CRZ signaling in regulating the female PMR and demonstrate that female CRZ signaling and male-derived signals cooperate to induce post-mating transitions in BPHs and D. melanogaster. CRZ is a paralog of the peptide gonadotropin-releasing hormone, known to regulate reproduction in vertebrates, including humans, suggesting evolutionary conservation of an ancient function.

genetics↗

bioRxiv2025

The host range paradox of Meloidogyne incognita: a physiological and transcriptomic analysis of nine susceptible interactions across six plant orders

The plant-parasitic nematode Meloidogyne incognita is the pathogen with the broadest host range among all known biotrophic interactions. This species is also the single most damaging of a group of agriculturally important plant-parasites, which together are estimated to contribute to losses in excess of $170 billion/year to world agriculture. Understanding how M. incognita is able to infect representatives from most orders of flowering plants, covering more than 3000 species, addresses a fundamentally important question of how pathogens adapt to their host-environment, and may inform control of a pathogen which threatens global food security. Here, we analyse the plant-nematode infection phenotype, and cross-kingdom transcriptome, of nine interactions across six orders of flowering plants at 25 days post infection. At this stage, majority of nematodes found within roots were immature and mature females (49.2% - 91.8%). Our data show that the phylogenetic distribution of hosts does not explain the phenotypic distribution of parasitism. Interestingly, though, M. incognita do have distinct transcriptional responses to different groups of hosts, but in a pattern which is independent of host phylogenetic. Three distinct nematode "transcriptional programmes" - Group 1, 2, and 3 - are evident, and we find that effectors are neither uniformly deployed across hosts, nor across groups of hosts. Importantly, we show that this differential deployment of effectors can have profound consequences for host specificity. Finally, we show that there is essentially no widespread core gall transcriptome at 25 days post infection, prompting the proposal of a model best described as "all roads lead to Rome".

plant biology↗

bioRxiv2025

Artificial selection for resistance to copper and off-target physiological and behavioral effects in Drosophila melanogaster

Pollution resulting from mining, industry, and agriculture has played an important role in shaping the evolution of diverse organisms. The pathway to evolved resistance can involve polygenic shifts that affect multiple traits. Heavy metal contamination is of particular concern due to the health effects for humans and cross-tolerance effects that influence pesticide resistance. We used a replicated artificial selection approach to examine the response to copper selection in Drosophila melanogaster collected from a retired mine and active fruit orchard. We tracked shifts in resistance to the target trait, copper, as well as off-target effects on cadmium and lead resistance, starvation resistance, lifespan, and feeding aversion to copper contaminated food. Selection for copper resistance increased the focal trait and maintained resistance to non-essential heavy metals while control flies lost metal resistance overtime, implying a cost associated with maintaining resistance to heavy metals. Starvation resistance was correlated with copper resistance, but the correlation was not sufficient to explain the gain in copper resistance. Lifespan was correlated with increased copper resistance in flies collected from one of the two collection sites, suggesting that life history traits may be influenced by repeated heavy metal exposure. Future genomic analysis will help clarify the genetic control of the selection response. Together, our results underscore the complexity of adaptive shift in polygenic traits and provide a basis for further exploration of costs and correlative change following copper selection.

evolutionary biology↗

bioRxiv2025

Cingulate-centered flexible control: physiologic correlates and enhancement by internal capsule stimulation

Flexible cognitive control requires rapid adjustment when current demands diverge from the brains predictions. Deficits in control adjustment are prominent in psychiatric disorders, yet the neural mechanisms supporting such adjustment and their modifiability remain unclear. We analyzed two intracranial electroencephalography datasets, one with brief internal capsule stimulation (ICS), to identify a human circuit mechanism for flexible adjustment after control-demand change. We further used two clinical internal capsule deep brain stimulation (IC DBS) cohorts to validate our proposed mechanism. Across intracranial datasets, phase-amplitude coupling (PAC) anchored to the theta phase of right rostral anterior cingulate cortex (rACC-R), particularly theta-gamma coupling with dorsolateral prefrontal cortex and dorsal ACC, was associated with faster performance specifically when control demand changed. This effect generalized across datasets and tasks and was robust across analytic pipelines. An adaptive drift-diffusion model accounted for this adjustment cost in terms of unsigned control prediction error. In Dataset 1, brief ICS selectively enhanced the same rACC-R theta-centered PAC configuration on control-state-change trials, and computational modeling indicated that stimulation improved control flexibility under high-mismatch conditions. In a primarily treatment-resistant depression IC DBS cohort, stimulation-induced enhancement of the flexibility-related behavioral parameter, rather than a more general-control parameter, was associated with clinical response (N = 14; AUC = 0.90). In a separate treatment-resistant obsessive-compulsive disorder IC DBS cohort, clinically selected stimulation settings preferentially increased the same flexibility-related parameter across all patients (N = 5). These findings identify rACC-centered theta-phase coordination as a candidate human mechanism for flexible adjustment, show that capsule stimulation can augment this mechanism when flexibility is required, and suggest a causal link between flexibility-related behavioral and neural signals and psychiatric symptoms.

neuroscience↗

bioRxiv2025

Development of long-term oyster tissue cultures reveals cellular plasticity, regenerative potential, and sustained physiological activity

Molluscs are ecologically and economically significant, yet progress in understanding their cellular biology has been constrained by the lack of reliable molluscan cell culture systems. Insights into key molluscan biology such as immunity, biomineralization, and host-pathogen interactions, has previously been limited by short term survival of primary culture systems. Here, we establish reproducible long-term in vitro and ex vivo cultures of cells and tissues from the oyster, Ostrea edulis, including whole heart, visceral mass, mantle, and gill explants. Explanted tissues remained viable for over two months, with whole hearts maintaining rhythmic, stimulus-responsive contractions for up to nine months. Cultures maintained structural and functional features while uncovering cellular plasticity, including haemocyte granule turnover, auricular epithelial remodelling with novel ciliation, and the emergence of de novo multicellular microtissues. Previously under-characterized tissue-specific cell types were maintained in culture, including adipocytes from the visceral mass and myocytes from the mantle, thereby expanding the cellular repertoire accessible in vitro. Mineralized structures were observed across tissue cultures, highlighting the potential of this system for studying biomineralization. By sustaining diverse cell types in vitro, these cultures provide a novel tractable platform for studying molluscan diseases, while also laying the foundation for deeper investigation into comparative invertebrate biology and the development of immortalised molluscan cell lines.

cell biology↗

bioRxiv2025

Pharmacological and Physiological Characteristics of Synaptic Transmissions from the Medial Prefrontal Cortex onto Noradrenergic Neurons and Their Presynaptic Neurons in the Mouse Locus Coeruleus

The locus coeruleus (LC) is the primary source of norepinephrine in the brain and is known to modulate brain-wide arousal state. Recent evidence suggests that it also regulates immediate attentional responses by resetting related cortical networks to optimize behavioral outcomes. Cortical regions of high cognitive function, such as the medial prefrontal cortex (mPFC), are theorized to directly influence LC output for the purpose of behavioral regulation. However, the available evidence is insufficient to provide a comprehensive understanding of the underlying mechanisms and properties. To provide further comprehensive data on this issue, we combined ex vivo whole-cell recording with an optogenetic approach to study the synaptic transmission of mPFC inputs to LC neurons, including noradrenergic (NA) neurons and GABAergic neurons presynaptic to them (preLC neurons). Our findings indicate that the mPFC exhibits monosynaptic connections with both NA and GABAergic preLC neurons. These synaptic connections demonstrate cell-type-specific disparities in glutamate release properties. In comparison to those on GABAergic preLC neurons, the mPFC fibers synapsing on LC-NA exhibit a lower release probability (higher paired-pulse ratio) and demonstrate a presynaptic enhancement of glutamate release efficacy during behavior. The features of simultaneous connections onto LC-NA and GABAergic preLC neurons, which exhibit cell-type-specific differences in plastic function of the transmitter release, enable the mPFC to effectively multiplex information to the LC for the adaptive regulation of behavior.

neuroscience↗

bioRxiv2025

SHORT DISORDERED PEPTIDES ARE SUFFICIENT TO CONVERT PROTEINS INTO MECHANOSENSORS THAT RESPOND TO PHYSIOLOGICAL CELLULAR FORCES

Mechanical forces regulate many biological processes and a handful of mechanosensor domains have been identified in proteins that respond to cellular forces. The Notch signaling pathway, a key regulator of cell communication processes ranging from animal development to cancer, is activated by mechanical forces generated by cell-cell interactions. The mechanosensing element of Notch is thought to be a 300 amino acid extracellular domain called negative regulatory region (NRR). Ligand binding exerts mechanical forces that are believed to partially unfold NRR, rendering it susceptible to cleavage by extracellular metalloproteases that leads to activation of signaling. However, some engineered notch-like receptors lacking the NRR can be activated upon ligand binding, although the mechanisms underlying this activation are not known. Here we observe that Notch molecules without the NRR, but with a 12 amino acid sequence present in the extracellular juxta-transmembrane domain (eJTMD) are also cleaved and activated upon interaction with their ligands in cultured cells and in transgenic animals. Furthermore, the [~]12 aa eJTMD from notch genes from other species (chicken, xenopus, zebrafish and Drosophila), from other unrelated transmembrane proteins (erbB4, N-CAM, and E-Cadherin), or multiple artificial sequences of [~]12 amino acids without any apparent structure or specific sequence can also convert proteins into ligand-dependent mechanosensors. These results indicate that short, disordered amino acid sequences that are commonly found in many proteins are capable of imparting mechanosensing capabilities into proteins, suggesting that mechanical force may regulate many more cellular processes than previously suspected.

cell biology↗