Biology subjects

Coyle, S. M.

Publications and source records attributed to Coyle, S. M..

10 recordsLinked to original sources

bioRxiv2025

A self-organizing single-cell morphology circuit optimizes Podophrya collini predatory trap structure

Cellular structure self-organizes through an interplay between internal mechanisms and external cues. In predatory ciliates, structures that capture prey obtain the resources needed for their construction, creating feedback between environmental inputs and morphological outputs. Here, we describe a self-organizing single-cell morphology circuit that adaptively optimizes the predatory trap structure of the suctorian Podophrya collini. These trap structures ensnare large cellular targets using an array of straw-like tentacles that siphon out prey cytoplasm upon contact with their tips. We find that trap architecture scales anisotropically, favoring tentacle number over length, to construct traps that maximize capture probability for the resources on hand. Drug perturbations, transcriptomics, proteomics, and expansion microscopy define distinct molecular and structural requirements that regulate trap structure maintenance and tentacle biogenesis. We integrate these findings into a mathematical model that explains the adaptive scaling of the trap and that makes predictions we confirm experimentally. More broadly, this circuits architecture provides a general-purpose control logic for organizing the number and size of sub-cellular structures applicable to other natural and engineered cellular systems.

cell biology↗

bioRxiv2025

Decoding ultrasensitive self-assembly of the calcium-regulated Tetrahymena cytoskeletal protein Tcb2 using optical actuation.

EF-hand calcium binding proteins are key macromolecular components of many unique filament systems and ultrafast contractile structures found in protists. However, our biochemical understanding of these cytoskeletal systems has been hindered by the need for assays that can controllably generate spatiotemporal calcium dynamics to probe their behavior. Here, we define the quantitative requirements for calcium-dependent self-assembly of the Tetrahymena cortical cytoskeletal protein Tcb2 using a microscopy-based spatiotemporally controlled optical calcium release assay. Light-driven uncaging of the photolabile calcium chelator DMNP-EDTA stimulates rapid localized self-assembly of Tcb2 into micron-scale protein networks. We quantify how the growth, size, and lifetime of Tcb2 networks is controlled by the duration and intensity of the applied light pulse. Incorporating the fluorescent calcium indicator Rhod-5N allows inference of the spatiotemporal distribution of calcium-bound Tcb2 monomers during the reaction and identifies a sharp, ultrasensitive transition to Tcb2 self-assembly. By applying this assay to mutants in Tcb2s four EF hand domains, we show that D184 is the key calcium binding site that licenses Tcb2 for self-assembly and define quantitative roles for other binding sites in tuning Tcb2s calcium-responsiveness. Our approach reveals a rich space of structures and regulation available to a single-protein system through coupling calcium-binding to ultrasensitive self-assembly, opening new paths forward to understanding other protist filament networks and contractile myonemes.

biophysics↗

bioRxiv2025

Noise-guided tuning of synthetic protein waves in living cells

Biological systems use protein circuits to organize cellular activities in space and time, but engineering synthetic dynamics is challenging due to stochastic effects of genetic and biochemical variation on circuit behavior. Genetically encoded oscillators (GEOs) built from bacterial MinDE-family ATPase and Activator modules generate fast orthogonal protein waves in eukaryotic cells, providing an experimental model system for genetic and biochemical coordination of synthetic protein dynamics. Here, we use budding yeast to experimentally define and model phase portraits that reveal how the breadth of frequencies and amplitudes available to a GEO are genetically controlled by ATPase and Activator expression levels and noise. GEO amplitude is encoded by ATPase absolute abundance, making it sensitive to extrinsic noise on a population level. In contrast, GEO frequency is remarkably stable because it is controlled by the Activator:ATPase ratio and thus affected primarily by intrinsic noise. These features facilitate noise-guided design of different expression strategies that act as filters on GEO waveform, enabling us to construct clonal populations that oscillate at different frequencies as well as independently tune frequency and amplitude variation within a single population. By characterizing 169 biochemically distinct GEOs, we provide a rich assortment of phase portraits as starting points for application of our waveform engineering approach. Our findings suggest noise-guided design may be a valuable strategy for achieving precision control over dynamic protein circuits.

synthetic biology↗

bioRxiv2025

Light-induced reversible assembly and actuation in ultrafast Ca2+-driven chemomechanical protein networks

Programming rapid, repeatable motions in soft materials has remained a challenge in active matter and biomimetic design. Here, we present a light-controlled chemomechanical network based on Tetrahymena thermophila calcium-binding protein 2 (Tcb2), a Ca2+-sensitive contractile protein. These networks--driven by Ca2+-triggered structural rearrangements--exhibit dynamic selfassembly, spatiotemporal growth, and contraction rates comparable to actomyosin systems. By coupling light-sensitive chelators for optically triggered Ca2+ release, we achieve precise growth and repeatable mechanical contractility of Tcb2 networks, revealing emergent phenomena such as boundary-localized active regions and density gradient-driven reversals in motion. A coupled reaction-diffusion and elastic model explains these dynamics, highlighting the interplay between chemical network assembly and mechanical response. We further demonstrate active transport of particles via network-mediated forces in vitro and implement reinforcement learning to program seconds-scale spatiotemporal actuation in silico. These results establish a platform for designing responsive active materials with rapid chemomechanical dynamics and tunable optical control, with applications in synthetic cells, sub-cellular force generation, and programmable biomaterials.

bioengineering↗

bioRxiv2025

Genetically encoded protein oscillators for FM streaming of single-cell data

Wireless devices use frequency modulation (FM) to reliably transmit information. Here, we establish a biochemical analogue of this paradigm using genetically encoded protein oscillators (GEOs) as carrier signals for real-time streaming of single-cell data. The GEO platform leverages evolutionarily diverse MinDE-family ATPase and Activator modules to generate fast synthetic protein oscillations in cells, where the waveform is controlled by both circuit biochemistry and intrinsic cellular physiology. This allows for diverse data encoding strategies, including noise-resistant FM reporters that broadcast proteasomal degradation or transcriptional dynamics; and autoencoder circuits that directly couple shifts in cell physiology to GEO waveform. Deploying GEOs in human embryonic stem cells allowed us to track single-cell developmental states in living populations during differentiation, revealing distinct patterns of spatiotemporal heterogeneity along endoderm, mesoderm, and ectoderm trajectories. The GEO platform establishes a dynamically controllable biochemical carrier signal, unlocking new high-fidelity FM data-encoding paradigms for continuous single-cell analysis.

synthetic biology↗

bioRxiv2024

Injury Severity is a Key Contributor to Coagulation Dysregulation and Fibrinogen Consumption

BackgroundTraumatic injury is a leading cause of death for those under the age of 45, with 40% occurring due to hemorrhage. Severe tissue injury and hypoperfusion lead to marked changes in coagulation, thereby preventing formation of a stable blood clot and increasing hemorrhage associated mortality. ObjectivesWe aimed to quantify changes in clot formation and mechanics occurring after traumatic injury and the relationship to coagulation kinetics, and fibrinolysis. MethodsPlasma was isolated from injured patients upon arrival to the emergency department. Coagulation kinetics and mechanics of healthy donors and patient plasma were compared with rheological, turbidimetric and thrombin generation assays. ELISAs were performed to determine tissue plasminogen activator (tPA) and D-dimer concentration, as fibrinolytic markers. ResultsSixty-three patients were included in the study. The median injury severity score (ISS) was 17, median age was 37.5 years old, and mortality rate was 30%. Rheological, turbidimetric and thrombin generation assays indicated that trauma patients on average, and especially deceased patients, exhibited reduced clot stiffness, increased fibrinolysis and reduced thrombin generation compared to healthy donors. Fibrinogen concentration, clot stiffness, D-dimer and tPA all demonstrated significant direct correlation to increasing ISS. Machine learning algorithms identified and highlighted the importance of clinical factors on determining patient outcomes. ConclusionsViscoelastic and biochemical assays indicate significant contributors and predictors of mortality for improved patient treatment and therapeutic target detection. ESSENTIALSO_LITraumatic injury may lead to alterations in a patients ability to form stable blood clots C_LIO_LIA study was performed to assess how trauma severity affects coagulation kinetics C_LIO_LIKey alterations were observed in trauma patients, who exhibit weaker and slower forming clots C_LIO_LIPaired with machine learning methods, the results indicate key aspects contributing to mortality C_LI

bioengineering↗

bioRxiv2023

Comparative profiling of cellular gait on adhesive micropatterns defines statistical patterns of activity that underlie native and cancerous cell dynamics.

Cell dynamics are powered by patterns of activity, but it is not straightforward to quantify these patterns or compare them across different environmental conditions or cell-types. Here we digitize the long-term shape fluctuations of metazoan cells grown on micropatterned fibronectin islands to define and extract statistical features of cell dynamics without the need for genetic modification or fluorescence imaging. These shape fluctuations generate single-cell morphological signals that can be decomposed into two major components: a continuous, slow-timescale meandering of morphology about an average steady-state shape; and short-lived "events" of rapid morphology change that sporadically occur throughout the timecourse. By developing statistical metrics for each of these components, we used thousands of hours of single-cell data to quantitatively define how each axis of cell dynamics was impacted by environmental conditions or cell-type. We found the size and spatial complexity of the micropattern island modulated the statistics of morphological events--lifetime, frequency, and orientation--but not its baseline shape fluctuations. Extending this approach to profile a panel of triple negative breast cancer cell-lines, we found that different cell-types could be distinguished from one another along specific and unique statistical axes of their behavior. Our results suggest that micropatterned substrates provide a generalizable method to build statistical profiles of cell dynamics to classify and compare emergent cell behaviors.

systems biology↗

bioRxiv2023

A synthetic biology approach defines mechanistic and regulatory roles for phase condensation in the assembly of microtubule architectures by human TPPP

Microtubules filaments are assembled into higher-order structures and machines critical for cellular processes using microtubule-associated proteins (MAPs). However, the design of synthetic MAPs that direct the formation of new structures in cells is challenging, as nanoscale biochemical activities must be organized across micron length-scales. Here we develop synthetic MAP-IDR condensates (synMAPs) that provide tunable and regulatable assembly of higher-order microtubule structures in vitro and in mammalian cells. synMAPs harness a small microtubule-binding domain from oligodendrocytes (TPPP) whose activity can be synthetically rewired by interaction with condensate-forming IDR sequences. This combination allows synMAPs to self-organize multivalent structures that bind and bridge microtubules into synthetic architectures. Regulating the connection between the microtubule-binding and condensate-forming components allows synMAPs to act as nodes in more complex cytoskeletal circuits in which the formation and dynamics of the microtubule structure can be controlled by small molecules or cell-signaling inputs. By systematically testing a panel of synMAP circuit designs, we define a two-level control scheme for dynamic assembly of microtubule architectures at the nanoscale (via microtubule-binding) and microscale (via condensate formation). synMAPs provide a compact and rationally engineerable starting point for the design of more complex microtubule architectures and cellular machines.

synthetic biology↗

bioRxiv2022

A programmable reaction-diffusion system for spatiotemporal cell signaling circuit design

Cells self-organize molecules in space and time to generate complex behaviors, but we lack synthetic strategies for engineering spatiotemporal signaling. We present a programmable reaction-diffusion platform for designing protein oscillations, patterns, and circuits in mammalian cells using two bacterial proteins, MinD and MinE (MinDE). MinDE circuits act like "single-cell radios", emitting frequency-barcoded fluorescence signals that can be spectrally isolated and analyzed using digital signal processing tools. We define how to genetically program these signals and modulate their dynamics using engineerable protein-protein interactions. By connecting MinDE to endogenous cellular pathways, we built circuits that broadcast frequency-barcoded single-cell kinase activity or that synthetically pattern actin polymerization. Our work establishes a new paradigm for probing and engineering cellular activities at length and timescales critical for biological function.

synthetic biology↗

bioRxiv2018

Coupled active systems encode emergent behavioral dynamics of the unicellular predator Lacrymaria olor

Multiple active systems in a cell work together to produce sophisticated cellular behaviors such as motility and search. However, it is often unclear how this coupling specifies the complex emergent dynamics that define such behaviors. As a model system, we analyzed the hunting strategy of Lacrymaria olor, a unicellular predatory ciliate that uses extreme morphological changes to extend, contract and whip an apparent \"cell neck\" over many body lengths to capture prey. Tracking millions of unique subcellular morphologies over time revealed that these fast dynamics encode a comprehensive local search behavior apparent only at longer timescales. This hunting behavior emerges as a tug-of-war between active sub-cellular structures that use surface cilia and cortex contractility to deform the structure of the neck. The resulting search space can be described mathematically using a small number of normal shape modes that change amplitude rapidly during hunts. The distribution of these shape modes in space and time reveals a transition point between tense and compressed neck morphologies at the mean neck length, such that new shapes are readily sampled by repeatedly extending and retracting across this critical length. Molecular perturbations to the cell-signaling controller show that coupling between ciliary and contractile programs is needed to maintain this length/shape relationship; neither system alone provides the dynamic repertoire of shapes necessary for comprehensive search. Our results highlight the utility of coupling antagonistic active systems as a strategy for encoding or engineering complex behaviors in molecular machines.\n\nOne Sentence Summary: Analysis of millions of unique cellular morphologies of the highly dynamic single-celled predator Lacrymaria olor reveals that it programs a comprehensive search space and emergent hunting behavior through coupling surface based active cilia and cortex based contractile molecular systems together.

systems biology↗