Biology subjects

Prakash, M.

Publications and source records attributed to Prakash, M..

At least 37 records · Page 2Linked to original sources

bioRxiv2024

Optineurin-facilitated axonal mitochondria delivery promotes neuroprotection and axon regeneration

Optineurin (OPTN) mutations are linked to amyotrophic lateral sclerosis (ALS) and normal tension glaucoma (NTG), but a relevant animal model is lacking, and the molecular mechanisms underlying neurodegeneration are unknown. We found that OPTN C-terminus truncation (OPTN{Delta}C) causes late-onset neurodegeneration of retinal ganglion cells (RGCs), optic nerve (ON), and spinal cord motor neurons, preceded by a striking decrease of axonal mitochondria. Surprisingly, we discover that OPTN directly interacts with both microtubules and the mitochondrial transport complex TRAK1/KIF5B, stabilizing them for proper anterograde axonal mitochondrial transport, in a C- terminus dependent manner. Encouragingly, overexpressing OPTN/TRAK1/KIF5B reverses not only OPTN truncation-induced, but also ocular hypertension-induced neurodegeneration, and promotes striking ON regeneration. Therefore, in addition to generating new animal models for NTG and ALS, our results establish OPTN as a novel facilitator of the microtubule-dependent mitochondrial transport necessary for adequate axonal mitochondria delivery, and its loss as the likely molecular mechanism of neurodegeneration.

neuroscience↗

bioRxiv2024

Algorithmic construction of topologically complex biomineral lattices via cellular syncytia

Biomineralization is ubiquitous in both unicellular and multicellular living systems [1, 2] and has remained elusive due to a limited understanding of physicochemical and biomolecular processes [3]. Echinoderms, identified with diverse architectures of calcite-based structures in the dermis[4], present an enigma of how cellular processes control shape and form of individual structures. Specifically, in holothurians (sea cucumbers), multi-cellular clusters construct discrete single-crystal calcite ossicles ([~] 100 {micro}m length scale), with diverse morphologies both across species and even within an individual animal [5]. The local rules that might encode these unique morphologies in calcite ossicles in holothurians remain largely unknown. Here we show how transport processes in a cellular syncytium impart a top-down control on ossicle geometry via symmetry breaking, branching, and fusion in finite cellular clusters. As a unique example of cellular masonary, we show how coordination within a small cluster of cells builds calcite structures about an order of magnitude larger than any individual participating cell. We establish live imaging of ossicle growth in Apostichopus parvimensis juveniles revealing how individual crystalline seeds ([~] 1 - 2 {micro}m) grow inside a multi-cellular syncytial complex with the biomineral completely wrapped within a membrane-bound cytoplasmic sheath. Constructing a topological description of ossicle geometries from 3D micro-CT (computational tomography) data reveals the hidden growth history and conserved patterns across ossicle types. We further demonstrate vesicle transport on the surface of the ossicle, rather than cell motility, regulates material transport to the ossicle tips via a unique cytoskeletal architecture. Finally, using reduced order models of conserved transport on self-closing active branching networks, we highlight the hidden universality in the growth process of distinct ossicles. The system presented here serves as a unique playground merging top-down cellular physiology and classical branching morphogenesis [6] with bottom-up non-equilibrium mineralization [7] processes at the interface of living and non-living matter [8].

biophysics↗

bioRxiv2024

ESPressoscope: a small and powerful platform for in situ microscopy

Microscopy is essential for detecting, identifying, analyzing, and measuring small objects. Access to modern microscopy equipment is crucial for scientific research, especially in the biomedical and analytical sciences. However, the high cost of equipment, limited availability of parts, and challenges associated with transporting equipment often limit the accessibility and operational capabilities of these tools, particularly in field sites and other remote or resource-limited settings. Thus, there is a need for affordable and accessible alternatives to traditional microscopy systems. We address this challenge by investigating the feasibility of using a simple microcontroller board not only as a portable and field-ready digital microscope, but furthermore as a versatile platform which can easily be adapted to a variety of imaging applications. By adding a few external components, we demonstrate that a low-cost ESP32 camera board can be used to build an autonomous in situ platform for digital time-lapse imaging of cells. This platform, which we call the ESPressoscope, can be adapted to applications ranging from monitoring incubator cell cultures in the lab to observing ecological phenomena in the sea, and it can be adapted for other techniques such as microfluidics or spectrophotometry. The ESPressoscope achieves a low power consumption and small size, which makes it ideal for field research in environments and applications where microscopy was previously infeasible. Its Wi-Fi connectivity enables integration with external image processing and storage systems, including on cloud platforms when internet access is available. Finally, we present several web browser-based tools to help users operate and manage the ESPressoscopes software. Our findings demonstrate the potential for low-cost, portable microscopy solutions to enable new and more accessible experiments for biological and analytical applications.

microbiology↗

bioRxiv2024

Innate Immune Activation and Mitochondrial ROS Invoke Persistent Cardiac Conduction System Dysfunction after COVID-19

BackgroundCardiac risk rises during acute SARS-CoV-2 infection and in long COVID syndrome in humans, but the mechanisms behind COVID-19-linked arrhythmias are unknown. This study explores the acute and long term effects of SARS-CoV-2 on the cardiac conduction system (CCS) in a hamster model of COVID-19. MethodsRadiotelemetry in conscious animals was used to non-invasively record electrocardiograms and subpleural pressures after intranasal SARS-CoV-2 infection. Cardiac cytokines, interferon-stimulated gene expression, and macrophage infiltration of the CCS, were assessed at 4 days and 4 weeks post-infection. A double-stranded RNA mimetic, polyinosinic:polycytidylic acid (PIC), was used in vivo and in vitro to activate viral pattern recognition receptors in the absence of SARS-CoV-2 infection. ResultsCOVID-19 induced pronounced tachypnea and severe cardiac conduction system (CCS) dysfunction, spanning from bradycardia to persistent atrioventricular block, although no viral protein expression was detected in the heart. Arrhythmias developed rapidly, partially reversed, and then redeveloped after the pulmonary infection was resolved, indicating persistent CCS injury. Increased cardiac cytokines, interferon-stimulated gene expression, and macrophage remodeling in the CCS accompanied the electrophysiological abnormalities. Interestingly, the arrhythmia phenotype was reproduced by cardiac injection of PIC in the absence of virus, indicating that innate immune activation was sufficient to drive the response. PIC also strongly induced cytokine secretion and robust interferon signaling in hearts, human iPSC-derived cardiomyocytes (hiPSC-CMs), and engineered heart tissues, accompanied by alterations in electrical and Ca2+ handling properties. Importantly, the pulmonary and cardiac effects of COVID-19 were blunted by in vivo inhibition of JAK/STAT signaling or by a mitochondrially-targeted antioxidant. ConclusionsThe findings indicate that long term dysfunction and immune cell remodeling of the CCS is induced by COVID-19, arising indirectly from oxidative stress and excessive activation of cardiac innate immune responses during infection, with implications for long COVID Syndrome.

pathology↗

bioRxiv2023

Engineering luminopsins with improved coupling efficiencies

SignificanceLuminopsins (LMOs) are bioluminescent-optogenetic tools with a luciferase fused to an opsin that allow bimodal control of neurons by providing both optogenetic and chemogenetic access. Determining which design features contribute to the efficacy of LMOs will be beneficial for further improving LMOs for use in research. AimWe investigated the relative impact of luciferase brightness, opsin sensitivity, pairing of emission and absorption wavelength, and arrangement of moieties on the function of LMOs. ApproachWe quantified efficacy of LMOs through whole cell patch clamp recordings in HEK293 cells by determining coupling efficiency, the percentage of maximum LED induced photocurrent achieved with bioluminescent activation of an opsin. We confirmed key results by multielectrode array (MEAs) recordings in primary neurons. ResultsLuciferase brightness and opsin sensitivity had the most impact on the efficacy of LMOs, and N-terminal fusions of luciferases to opsins performed better than C-terminal and multi-terminal fusions. Precise paring of luciferase emission and opsin absorption spectra appeared to be less critical. ConclusionsWhole cell patch clamp recordings allowed us to quantify the impact of different characteristics of LMOs on their function. Our results suggest that coupling brighter bioluminescent sources to more sensitive opsins will improve LMO function. As bioluminescent activation of opsins is most likely based on Forster resonance energy transfer (FRET), the most effective strategy for improving LMOs further will be molecular evolution of luciferase-fluorescent protein-opsin fusions.

molecular biology↗

bioRxiv2023

Curved crease origami and topological singularities at a cellular scale enable hyper-extensibility of Lacrymaria olor

Eukaryotic cells undergo dramatic morphological changes during cell division, phagocytosis and motility. Fundamental limits of cellular morphodynamics such as how fast or how much cellular shapes can change without harm to a living cell remain poorly understood. Here we describe hyper-extensibility in the single-celled protist Lacrymaria olor, a 40 {micro}m cell which is capable of reversible and repeatable extensions (neck-like protrusions) up to 1500 {micro}m in 30 seconds. We discover that a unique and intricate organization of cortical cytoskeleton and membrane enables these hyper-extensions that can be described as the first cellular scale curved crease origami. Furthermore, we show how these topological singularities including d- cones and twisted domain walls provide a geometrical control mechanism for the deployment of membrane and microtubule sheets as they repeatably spool thousands of time from the cell body. We lastly build physical origami models to understand how these topological singularities provide a mechanism for the cell to control the hyper-extensile deployable structure. This new geometrical motif where a cell employs curved crease origami to perform a physiological function has wide ranging implications in understanding cellular morphodynamics and direct applications in deployable micro-robotics. Significance statementHere we present the discovery of curved crease origami at the scale of a single cell. We show how topological singularities in the origami (d-cones) and twist walls in microtubule ribbons control deployment of a hyper-extensile neck in a single-celled protist. Our work establishes a direct link between geometry and cell behavior, connecting form and function of cellular morphodynamics.

biophysics↗

bioRxiv2023

Machine learning prediction of future amyloid beta positivity in amyloid-negative individuals

INTRODUCTIONThe pathophysiology of Alzheimers disease (AD) involves {beta}-amyloid (A{beta}) accumulation. Early identification of individuals with abnormal {beta}-amyloid levels is crucial, but A{beta} quantification with positron emission tomography (PET) and cerebrospinal fluid (CSF) is invasive and expensive. METHODSWe propose a machine learning framework using standard non-invasive (MRI, demographics, APOE, neuropsychology) measures to predict future A{beta}-positivity in A{beta}-negative individuals. We separately study A{beta}-positivity defined by PET and CSF. RESULTS: Cross-validated AUC for 4-year A{beta} conversion prediction was 0.78 for the CSF-based and 0.68 for the PET-based A{beta} definitions. Although not trained for the clinical status-change prediction, the CSF-based model excelled in predicting future mild cognitive impairment (MCI)/dementia conversion in cognitively normal/MCI individuals (AUCs, respectively, 0.76 and 0.89 with a separate dataset). DISCUSSIONStandard measures have potential in detecting future A{beta}-positivity and assessing conversion risk, even in cognitively normal individuals. The CSF-based definition led to better predictions than the PET-based definition.

neuroscience↗

bioRxiv2023

A Bioluminescent Activity Dependent (BLADe) Platform for Converting Neuronal Activity to Photoreceptor Activation

Genetically encoded sensors and actuators have advanced the ability to observe and manipulate cellular activity, yet few non-invasive strategies enable cells to directly couple their intracellular states to user-defined outputs. We promote a bioluminescent activity-dependent (BLADe) platform that facilitates programmable feedback through genetically encoded light generation. Using calcium (Ca{superscript 2}) flux as a model, we engineered a Ca{superscript 2}-dependent luciferase that functions as an activity-gated light source capable of photoactivating light-sensing actuators. As an initial demonstration of the versatility of this platform we present two separate use cases in neurons. In the first application, the presence of luciferin triggers Ca{superscript 2} dependent local illumination that provides activity dependent gene expression by activating a light-sensitive transcription factor. In the second application, neuronal activity-driven Ca{superscript 2} fluctuations via locally generated bioluminescence control neural dynamics through opsin activation in single cells, populations and intact tissue. BLADe can be expanded to couple any signal that bioluminescent enzymes can be engineered to detect with the wide variety of photosensing actuators. This modular strategy of coupling an activity dependent light emitter to a light sensing actuator offers, in principle, a generalizable framework for state dependent cell-autonomous control across biological systems.

neuroscience↗

bioRxiv2023

A FRET-based excitatory luminopsin with improved efficacy of bioluminescence-driven activation

SignificanceBioluminescent optogenetics (BL-OG) offers a unique and powerful approach to manipulate neural activity both opto- and chemogenetically using a single actuator molecule (a LuMinOpsin, LMO). AimTo further enhance the utility of BL-OG by improving the efficacy of chemogenetic (bioluminescence- driven) LMO activation. ApproachWe developed novel luciferases optimized for Forster resonance energy transfer (FRET) when fused to the fluorescent protein mNeonGreen, generating bright bioluminescent (BL) emitters spectrally tuned to Volvox Channelrhodopsin 1 (VChR1). ResultsA new LMO generated from this approach (LMO7) showed significantly stronger BL-driven opsin activation compared to previous and other new variants. We extensively benchmarked LMO7 against LMO3 (current standard), and found significantly stronger neuronal activity modulation ex vivo and in vivo, and efficient modulation of behavior. ConclusionsWe report a robust new option for achieving multiple modes of control in a single actuator, and a promising engineering strategy for continued improvement of BL-OG.

neuroscience↗

bioRxiv2023

Coupling and uncoupling of midline morphogenesis and cell flow in amniote gastrulation

Large-scale cell flow characterizes gastrulation in animal development. In amniote gastrulation, particularly in avian gastrula, a bilateral vortex-like counter-rotating cell flow, called polonaise movements, appears along the midline. Here, through experimental manipulations, we addressed relationships between the polonaise movements and morphogenesis of the primitive streak, the earliest midline structure in amniotes. Suppression of the Wnt/planar cell polarity (PCP) signaling pathway maintains the polonaise movements along a deformed primitive streak. Mitotic arrest leads to diminished extension and development of the primitive streak and maintains the early phase of the polonaise movements. Ectopically induced Vg1, an axis-inducing morphogen, generates the polonaise movements, aligned to the induced midline, but disturbs the stereotypical cell flow pattern at the authentic midline. Despite the altered cell flow, induction and extension of the primitive streak are preserved along both authentic and induced midlines. Finally, we show that ectopic axis-inducing morphogen, Vg1, is capable of initiating the polonaise movements without concomitant PS extension under mitotic arrest conditions. These results are consistent with a model wherein primitive streak morphogenesis is required for the maintenance of the polonaise movements, but the polonaise movements are not necessarily responsible for primitive streak morphogenesis. Our data describe a previously undefined relationship between the large-scale cell flow and midline morphogenesis in gastrulation.

developmental biology↗

bioRxiv2023

Energetics of the Microsporidian Polar Tube Invasion Machinery

Microsporidia are eukaryotic, obligate intracellular parasites that infect a wide range of hosts, leading to health and economic burdens worldwide. Microsporidia use an unusual invasion organelle called the polar tube (PT), which is ejected from a dormant spore at ultra-fast speeds, to infect host cells. The mechanics of PT ejection are impressive. Anncaliia algerae microsporidia spores (3-4 {micro}m in size) shoot out a 100-nm-wide PT at a speed of 300 {micro}m/sec, creating a shear rate of 3000 sec-1. The infectious cargo, which contains two nuclei, is shot through this narrow tube for a distance of [~]60-140 {micro}m (Jaroenlak et al., 2020) and into the host cell. Considering the large hydraulic resistance in an extremely thin tube and the low-Reynolds-number nature of the process, it is not known how microsporidia can achieve this ultrafast event. In this study, we use Serial Block-Face Scanning Electron Microscopy to capture 3-dimensional snapshots of A. algerae spores in different states of the PT ejection process. Grounded in these data, we propose a theoretical framework starting with a systematic exploration of possible topological connectivity amongst organelles, and assess the energy requirements of the resulting models. We perform PT ring experiments in media of varying viscosity, and use the results to rank our proposed hypotheses based on their predicted energy requirement. We also present a possible mechanism for cargo translocation, and quantitatively compare our predictions to experimental observations. Our study provides a comprehensive biophysical analysis of the energy dissipation of microsporidian infection process and demonstrates the extreme limits of cellular hydraulics. Statement of SignicanceMicrosporidia are a group of spore-forming, intracellular parasites that infect a wide range of hosts (including humans). Once triggered, microsporidian spores (3-4 {micro}m in size) shoot out a specialized organelle called the polar tube (PT) (60-140 {micro}m long, 100 nm wide) at ultrafast speed (300 {micro}m/sec), penetrating host cells and acting as a conduit for the transport of infectious cargo. Although this process has fascinated biologists for a century, the biophysical mechanisms underlying PT extrusion are not understood. We thus take a data-driven approach to generate models for the physical basis of PT ring and cargo transport through the PT. Our approach here demonstrates the extreme limits of cellular hydraulics and the potential applications of biophysical approaches to other cellular architectures.

biophysics↗

bioRxiv2022

Thermotaxis in an apolar, non-neuronal animal

Neuronal circuits are hallmarks of complex decision making processes in the animal world. How animals without neurons process information and respond to environmental cues promises a new window into studying precursors of neuronal control and origin of the nervous system as we know it today. Robust decision making in animals, such as in chemotaxis or thermotaxis, often requires internal symmetry breaking (such as A-P axis) provided naturally by a given body plan of an animal. Here we report the discovery of robust thermotaxis behavior in Trichoplax adhaerens, an early-divergent, enigmatic animal with no anterior-posterior symmetry breaking (apolar), and no known neurons or muscles. We present the first quantitative and robust behavioral response assay in placozoan, which present an apolar flat geometry. By exposing T. adhaerens to a thermal gradient under a long-term imaging setup, we observe robust thermotaxis that occurs over timescale of hours, independent of any circadian rhythms. We quantify that T. adhaerens can detect thermal gradients of at least 0.1 {degrees}C/cm. Positive thermotaxis is observed for a range of baseline temperatures from 17-22.5 {degrees}C with motility trajectories to be well-described by a Levy distribution. Interestingly, the organism does not maintain a fixed orientation while performing thermotaxis. Utilizing natural diversity in size of adult organisms (100um to a few mm), we also demonstrate a critical animal size above which thermotaxis behavior is hindered. Several TRP family homologs have been previously reported to be conserved in metazoans, including in T. adhaerens. We discover naringenin, a known TRPM3 antagonist, inhibits thermotaxis in T. adhaerens. The discovery of robust thermotaxis in T. adhaerens provides a tractable handle to interrogate information processing in a brainless animal. Understanding how divergent marine animals process thermal cues is also critical due to rapid temperature rise in our oceans.

animal behavior and cognition↗

bioRxiv2022

A topologically complex cytoplasm enables inflation-based escape from a gravity trap

The daily vertical migrations of plankton play a crucial role in shaping marine ecosystems and influencing global biogeochemical cycles. They also form the foundation of the largest daily biomass movement on Earth. Surprisingly, amongst this diverse group of organisms, some single cell protists transit these depths exceeding 50 meters without employing flagella or cilia, and the underlying mechanisms remain poorly understood. It has been previously proposed that this capability relies on the cells ability to regulate its internal density relative to seawater. Here, using Pyrocystis noctiluca as a model system, we demonstrate the primary mechanism for this density control is a rapid cellular inflation event, during which a single plankton cell expands its volume six-fold in less than 10 minutes. This self-regulated cellular inflation selectively imports fluid less dense than surrounding seawater, and can effectively sling-shot a cell and reverse sedimentation within minutes. This ability is made possible by a reticulated cytoplasmic architecture in Pyrocystis noctiluca that enables this rapid increase in overall cell volume without dilution of its cytoplasmic content. We further present a generalized mathematical framework that unifies cell cycle driven density regulation, stratified ecology, and associated cell behavior in the open ocean. Our study unveils an ingenious strategy employed by non-motile plankton to evade the gravitational sedimentation trap, highlighting how precise control of cell size was essential for survival in the ocean.

cell biology↗

bioRxiv2022

Afadin and zyxin contribute to coupling between cell junctions and contractile actomyosin networks during apical constriction

One of the most common cell shape changes driving morphogenesis in diverse animals is the constriction of the apical cell surface. Apical constriction depends on contraction of an actomyosin network in the apical cell cortex, but such actomyosin networks have been shown to undergo continual, conveyor belt-like contractions before the shrinking of an apical surface begins. This finding suggests that apical constriction is not necessarily triggered by the contraction of actomyosin networks, but rather can be triggered by unidentified, temporally-regulated mechanical links between actomyosin and junctions. Here, we used C. elegans gastrulation as a model to seek genes that contribute to such dynamic linkage. We found that -catenin and {beta}-catenin initially failed to move centripetally with contracting cortical actomyosin networks, suggesting that linkage is regulated between intact cadherin-catenin complexes and actomyosin. We used proteomic and transcriptomic approaches to identify new players, including the candidate linkers AFD-1/afadin and ZYX-1/zyxin, as contributing to C. elegans gastrulation. We found that ZYX-1/zyxin is among a family of LIM domain proteins that have transcripts that become enriched in multiple cells just before they undergo apical constriction. We developed a semi-automated image analysis tool to describe rates of myosin and membrane movement, which we then used to discover that ZYX-1/zyxin contributes to cell-cell junctions centripetal movement in concert with contracting actomyosin networks. These results identify several new genes that contribute to C. elegans gastrulation, and they identify zyxin as a key protein important for actomyosin networks to effectively pull cell-cell junctions inward during apical constriction. The transcriptional upregulation of ZYX-1/zyxin in specific cells in C. elegans points to one way that developmental patterning spatiotemporally regulates cell biological mechanisms in vivo. Because zyxin and related proteins contribute to membrane-cytoskeleton linkage in other systems, we anticipate that its roles in regulating apical constriction in this manner may be conserved.

genetics↗

bioRxiv2022

Emergent Programmable Behavior and Chaos in Dynamically Driven Active Filaments

AO_SCPLOWBSTRACTC_SCPLOWHow the behavior of single cells emerges from their constituent sub-cellular biochemical and physical parts is an outstanding challenge at the intersection of biology and physics. A remarkable example of single-cell behavior is seen in the ciliate Lacrymaria olor, which hunts by striking its prey via rapid movements and protrusions of a slender neck, many times the size of the original cell body. This dynamics of the cell neck is powered by active injection of energy into this slender filamentous structure via a coat of cilia across its length and specialized cilia at the tip. How a cell can program this dynamical active filament to produce desirable behaviors like search and homing to a target remains unknown. By constructing a coupled active-elastic and hydrodynamic model of a slender filament with activity at the tip, here we uncover how cell behavior (filament shape dynamics) can be controlled via activity dynamics. Our model captures two key features of this system - dynamic activity patterns (extension and compression cycles) and active stresses that are uniquely aligned with the filament geometry - leading to a so-called "follower force" constraint. We show that active filaments under deterministic, time-varying follower forces display rich behaviors including periodic and aperiodic shape dynamics over long times. We further show that aperiodic dynamics occur due to a transition to chaos in regions of a biologically accessible parameter space. By further dissecting the non-linear dynamics of this active filament system, we discover a simple iterative map of filament shape that predicts long-term behavior. Lastly, using these iterative maps as a design tool, we demonstrate examples of how a cell could "program" filament behaviors by using frequency and amplitude modulated activity patterns. Overall, our results serve as a framework to mechanistically understand behavior in single cells such as L. olor and present a novel chaotic dynamical system in active elastohydrodynamics. Our work also offers a direct framework for designing programmable active matter systems using filament geometries. Significance statementSingle-celled protozoa display remarkable animal-like behaviors without the aid of neurons. Mechanistically understanding how this dynamic behavior emerges from underlying physical and biochemical components is an outstanding challenge. In this work, using an active filament model, we uncover the fundamental non-linear dynamics and non-variational mechanics that underlie the complex behaviors of single cells like Lacrymaria olor. In doing so we discover a novel route to chaos in active elastohydrodynamic systems and the first-ever description of how chaos can drive single-cell behaviors. Lastly, we present a framework for how filament behaviors can be "programmed" using dynamic, modulated activity patterns. Overall our work provides mechanistic insights into single-cell behaviors and offers a new framework for the design of filamentous active matter systems to achieve diverse functions.

biophysics↗

bioRxiv2021

Entangled architecture of rough endoplasmic reticulum (RER) and vacuoles enables topological damping in cytoplasm of an ultra-fast giant cell

Cellular systems are known to exhibit some of the fastest movements in biology - but little is known as to how single cells can dissipate this energy rapidly and adapt to such large accelerations without disrupting internal architecture. To address this, we investigate Spirostomum ambiguum - a giant cell (1-4 mm in length) well-known to exhibit ultrafast contractions (50% of body length) within 5 msec with a peak acceleration of 15g. Utilizing transmitted electron microscopy (TEM) and confocal imaging, we discover a novel association of rough endoplasmic reticulum (RER) and vacuoles throughout the cell - forming a contiguous fenestrated membrane architecture that topologically entangles these two organelles. A nearly uniform inter-organelle spacing of 60 nm is observed between RER and vacuoles, closely packing the entire cell. Inspired by the entangled organelle structure, we study the mechanical properties of entangled deformable particles using a vertex-based model, with all simulation parameters matching 10 dimensionless numbers to ensure dynamic similarity. We demonstrate how entangled deformable particles respond to external loads by an increased viscosity against squeezing and help preserve spatial relationships. Because this enhanced damping arises from the entanglement of two networks incurring a strain-induced jamming transition at subcritical volume fractions, which is demonstrated through the spatial correlation of velocity direction, we term this phenomenon "topological damping". Our findings suggest a new mechanical role of RER-vacuolar meshwork as a metamaterial capable of damping an ultra-fast contraction event. Significance StatementLittle is known about how single-cell organisms with extreme motility can decelerate or dissipate energy, as they lack connective tissues. Our study discovered a novel entangled rough endoplasmic reticulum (RER)-vacuolar meshwork architecture in Spirostomum ambiguum, an ultrafast giant cell that can contract itself with 15g accelerations. We demonstrate through an entangled deformable particle model that the entangled architecture increases the squeeze-flow viscosity of particle systems and helps dampen the motion, a phenomenon we called "topological damping". For biologists, our study suggests the mechanical role of RER through topological constraints on nearby organelles. For physicists, we point out a new way to create a system with strain-induced jamming. For engineers, we present a novel architecture that can provide braking functions.

cell biology↗

bioRxiv2021

Selective Control of Synaptically-Connected Circuit Elements by All-Optical Synapses

Understanding percepts, engrams and actions requires methods for selectively modulating synaptic communication between specific subsets of interconnected cells. Here, we develop an approach to control synaptically connected elements using bioluminescent light: Luciferase-generated light, originating from a presynaptic axon terminal, modulates an opsin in its postsynaptic target. Vesicular-localized luciferase is released into the synaptic cleft in response to presynaptic activity, creating a real-time Optical Synapse. Light production is under experimenter-control by introduction of the small molecule luciferin. Signal transmission across this optical synapse is temporally defined by the presence of both the luciferin and presynaptic activity. We validate synaptic Interluminescence by multi-electrode recording in cultured neurons and in mice in vivo. Interluminescence represents a powerful approach to achieve synapse-specific and activity-dependent circuit control during behavior in vivo.

neuroscience↗

bioRxiv2021

Tissue scale properties of the extracellular matrix regulates nuclear shape, organisation and fate in the embryonic midline sutures

Balancing self-renewal and differentiation is a key feature of every stem cell niche and one that is tuned by mechanical interactions of cells with their neighbors and surrounding extracellular matrix. The fibrous stem cell niches that develop as sutures between skull bones must balance the complex extracellular environment that emerges to define them with self-renewal and bone production. Here, we address the role for physical stimuli in suture development by probing the relationship between nuclear shape, organization and gene expression in response to a developing collagen network in embryonic midline sutures. This work complements genetic approaches used to study sutures and provides the first quantitative analyses of physical structure in these sutures. By combining multiple imaging modalities with novel shape description, in addition to network analysis methods, we find the early emergence of a complex extracellular collagen network to have an important role in regulating morphogenesis and cell fate. We show that disrupted collagen crosslinking can alter ECM organization of midline sutures as well as stimulate expression of bone differentiation markers. Further, our findings suggest that in vivo, skeletal tissues can uncouple the response of the nuclear lamina from collagen mediated tissue stiffening seen in vitro. Our findings highlight a crucial relationship between the cellular microenvironment, tissue stiffness and geometry with gene expression in normal development and maintenance of progenitor fate in embryonic sutures.

developmental biology↗