Biology subjects

Wright, C. S.

Publications and source records attributed to Wright, C. S..

10 recordsLinked to original sources

bioRxiv2026

Contemporaneity of the past in stochastic intergenerational homeostasis

What recurring patterns of behaviors are hidden in the stochastic intergenerational dynamics of individual bacterial cells in different environments? Embracing the inherently stochastic nature of the homeostasis process, first we reconceptualize homeostasis as representing a "standing pattern of variation in trajectory space" of a naturally occurring adaptive complex system (rather than the simple "set-point" of a self-regulating apparatus), then delineate two mechanistically distinct potential routes to achieving homeostasis (either elastic, reflexive, memory-free adaptation or plastic, reflective, memoryful adaptation) and show that both schemes are simultaneously utilized during multigenerational stochastic growth and division of an individual cell. From experimental data we identify an intergenerational scaling law which directly yields the exact stochastic map governing stochastic intergenerational cell size homeostasis of individual bacterial cells. Its broad applicability across bacterial species, growth conditions, and microenvironments suggests that the organizational motif representing the nature of coupling of growth to division is effectively the same in all of these scenarios, despite apparent differences in actualization through molecular circuitry. The precise parameters characterizing the intergenerational scaling law vary from condition to condition and provide early hints of two tradeoffs: precision-speed and precision-energy.

biophysics↗

bioRxiv2026

Observing concurrent subcellular dynamics in large living tissues

An outstanding question in eukaryotic biology is the mechanistic connection between events occurring at (sub)cellular levels (time scales of milliseconds to minutes) to those at the tissue levels (tens of minutes to months). Deciphering such mechanisms requires imaging approaches capable of simultaneously achieving high spatial and temporal resolutions for large samples over long periods of time. Here, we demonstrate Airy beam-based light sheet microscopy of organelles in tens to hundreds of cells in a few hundred micrometre-wide tissue environments. We achieve a typical resolution of 320 nm over 266 x 266 x 100 m3 volumes at a temporal rate of 0.05 Hz, typically with generally used fluorophores such as Green Fluorescent Protein, over extended periods of time that allow tracking of organelle and protein dynamics. We validated our approach across different length and time scales by imaging mitochondria and endosome dynamics in very large fields of view in zebrafish tissue, molecular assemblies of myosin as gastrulation proceeds in Drosophila embryos, 3D mitochondrial streaming in mouse oocytes, pressure-driven motility and protrusions in amoebae, mitochondrial dynamics in cancer spheroids, 5 -colour fast imaging in iBlastoids, and endosomal dynamics in single cells. Through these model systems, we demonstrate the versatility of Airy beam light sheet microscopy to image large tissues at unprecedented high resolution; to capture dynamics in photosensitive, delicate samples; and to screen 3D samples. We anticipate that our Airy beam-based approach will represent a pivotal advance in cellular biology--especially developmental biology--as it provides, for the first time, true subcellular resolution over large imaging volumes with high temporal resolution.

developmental biology↗

bioRxiv2023

Scaling of stochastic growth and division dynamics: A comparative study of individual rod-shaped cells in the Mother Machine and SChemostat platforms

Microfluidic platforms enable long-term quantification of stochastic behaviors of individual bacterial cells under precisely controlled growth conditions. Yet, quantitative comparisons of physiological parameters and cell behaviors of different microorganisms in different experimental and device modalities is not readily possible owing to experiment-specific details affecting cell physiology in confounding ways. To rigorously assess the effects of mechanical confinement, we designed, engineered, and performed side-by-side experiments under otherwise identical conditions in the Mother Machine (with confinement) and the SChemostat (without confinement), using the latter as the ideal comparator. We established a protocol to cultivate a suitably engineered rod-shaped mutant of Caulobacter crescentus in the Mother Machine, and benchmarked the differences in stochastic growth and division dynamics in the Mother Machine with respect to the SChemostat. While the single-cell growth rate distributions are remarkably similar, the mechanically confined cells in the Mother Machine experience a substantial increase in interdivision times. However, we find that the division ratio distribution precisely compensates for this increase in the interdivision times, which in turn reflects identical emergent simplicities governing stochastic intergenerational homeostasis of cell sizes across device and experimental configurations, provided the cell sizes are appropriately mean-rescaled in each condition. Our results provide insights into the nature of the robustness of the bacterial growth and division machinery.

biophysics↗

bioRxiv2023

Architectural underpinnings of stochastic intergenerational homeostasis

Living systems are naturally complex and adaptive, and offer unique insights into the strategies for achieving and sustaining stochastic homeostasis in different conditions. Here, we focus on homeostasis in the context of stochastic growth and division of individual bacterial cells. We take advantage of high-precision longterm dynamical data that have recently been used to extract emergent simplicities and to articulate empirical intra- and in-tergenerational scaling laws governing these stochastic dynamics. We identify the core motif in the mechanistic coupling between division and growth, which naturally yields these precise rules, thus also bridging the intra- and intergenerational phenomenologies. By developing and utilizing novel techniques for solving a broad class of first passage processes, we derive the exact analytic necessary and sufficient condition for sustaining stochastic intergenerational cell size homeostasis within this framework. Furthermore, we provide predictions for the precision kinematics of cell size homeostasis, and the shape of the interdivision time distribution, which are compellingly borne out by the high-precision data. Taken together, these results provide insights into the functional architecture of control systems that yield robust yet flexible stochastic homeostasis.

biophysics↗

bioRxiv2023

Prrx1-driven LINC complex disruption in vivo does not significantly alter bone properties in 8-week male mice nor after 6-weeks voluntary wheel running

The Linker of Nucleoskeleton and Cytoskeleton (LINC) complex serves to connect the nuclear envelope and the cytoskeleton, influencing cellular processes such as nuclear arrangement, architecture, and mechanotransduction. The role LINC plays in mechanotransduction pathways in bone progenitor cells has been well studied; however, the mechanisms by which LINC complexes govern in vivo bone formation remain less clear. To bridge this knowledge gap, we established a murine model disrupting LINC using transgenic Prx-Cre mice and floxed Tg(CAG-LacZ/EGFP-KASH2) mice. Prx-Cre mice express the Cre recombinase enzyme controlled by the paired-related homeobox gene-1 promoter (Prrx1), a pivotal regulator of skeletal development. Prx-Cre animals have been widely used in the bone field to target bone progenitor cells. Tg(CAG-LacZ/EGFP-KASH2) mice carry a lox-stop-lox flanked LacZ gene allowing for the overexpression of an EGFP-KASH2 fusion protein via cre recombinase mediated deletion of the LacZ cassette. This disrupts endogenous Nesprin-Sun binding in a dominant negative manner disconnecting nesprin from the nuclear envelope. By combining these lines, we generated a Prrx1(+) cell-specific LINC disruption model to study its impact on the developing skeleton and subsequently exercise-induced bone accrual. The findings presented here indicate Prx-driven LINC disruption (PDLD) cells exhibit no change in osteogenic and adipogenic potential compared to controls in vitro nor are there bone quality changes when compared to in sedentary animals at 8 weeks. While PDLD animals displayed increased voluntary running activity andPrrx1(+) cell-specific LINC disruption abolished the exercise-induced increases in osteoid volume and surface after a 6-week exercise intervention, no other changes in bone microarchitecture or mechanical properties were found.

bioengineering↗

bioRxiv2023

Non-Markovian memory and emergent simplicities in the stochastic and plastic adaptation of individual cells to dynamic environments

Do individual bacterial cells retain memories of the history of environmental conditions experienced in previous generations? Here we directly address this question through a synthesis of physics theory and high-precision experiments on statistically identical, non-interacting individual bacterial cells, which grow and divide with intrinsic stochasticity in precisely controlled conditions. From these data, we extract "emergent simplicities" in the seemingly complex interplay between history dependence, persistence, and transience in the stochastic memories of the dynamic environments experienced by individuals over multiple generations. First, we find that the instantaneous single-cell growth rate is the key physiologically relevant quantity where intergenerational memory is stored. In contrast, the cell size dynamics are memory free, or Markovian, over intergenerational timescales. Next, we find that the effect of experiencing dynamic environments can be captured quantitatively by recal-ibrating the cellular unit of time by the measured mean instantaneous growth rate; the dynamically rescaled cell age distributions undergo a scaling collapse. Moreover, in a given condition, an individual bacterial cell retains history-dependent, or non-Markovian, memory of its growth rate over tens of generations. We derive from first principles a physically-motivated metric to quantify the degree of non-Markovianity. Furthermore, when conditions change, the instantaneous single-cell growth distribution becomes bimodal, as the bacteriums memory of past environment encountered is reset stochastically and plastically, prior to achieving a new homeostasis.

biophysics↗

bioRxiv2023

Emergent Simplicities in Stochastic Intergenerational Homeostasis

How do complex systems maintain key emergent "state variables" at desired target values to within specified tolerances? This question was first posed in the context of homeostasis in living systems over a century ago, and yet the precise quantitative rules governing this phenomenon have remained fiercely debated. We herein present a direct solution through a synthesis of high-precision experiments and first principles-based physics theory. After introducing a general approach that incorporates the inherently stochastic and dynamic nature of organismal homeostasis, we provide direct experimental evidence that stochastic intergenerational homeostasis is indeed maintained. Next, we identify a series of emergent simplicities hidden in these data. Remarkably, the dynamics of intergenerational homeostasis of organismal sizes are Markovian, or history-independent. The precision data reveal an intergenerational scaling law that fully determines, with no fine-tuning parameters, the exact stochastic map governing homeostasis, as borne out by compelling data- theory matches. These emergent simplicities in turn yield the necessary and sufficient condition for stochastic homeostasis, with surprising implications for the architecture of the underlying control system. Validation across different growth conditions, cell morphologies, experimental modalities, and organisms comprehensively establishes the universality of the results presented here.

biophysics↗

bioRxiv2022

Gabapentin Disrupts Binding of Perlecan to the α2δ1 Voltage Sensitive Calcium Channel Subunit and Impairs Skeletal Mechanosensation

Our understanding of how osteocytes, the principal mechanosensors within bone, sense and perceive force remains unclear. Previous work identified "tethering elements" (TEs) spanning the pericellular space of osteocytes and transmitting mechanical information into biochemical signals. While we identified the heparan sulfate proteoglycan perlecan (PLN) as a component of these TEs, PLN must attach to the cell surface to induce biochemical responses. As voltage-sensitive calcium channels (VSCCs) are critical for bone mechanotransduction, we hypothesized that PLN binds the extracellular 2{delta}1 subunit of VSCCs to couple the bone matrix to the osteocyte membrane. Here, we showed co-localization of PLN and 2{delta}1 along osteocyte dendritic processes. Additionally, we quantified the molecular interactions between 2{delta}1 and PLN domains and demonstrated for the first time that 2{delta}1 strongly associates with PLN via its domain III. Furthermore, 2{delta}1 is the binding site for the commonly used pain drug, gabapentin (GBP), which is associated with adverse skeletal effects when used chronically. We found that GBP disrupts PLN::2{delta}1 binding in vitro, and GBP treatment in vivo results in impaired bone mechanosensation. Our work identified a novel mechanosensory complex within osteocytes composed of PLN and 2{delta}1, necessary for bone force transmission and sensitive to the drug GBP. This work provides insights into the mechanisms underlying mechanotransduction and will inform future studies to understand the mechanisms responsible for the negative effects of GBP on bone.

molecular biology↗

bioRxiv2022

Heterotypic Endosomal Interactions Drive Emergent Early Endosomal Maturations

Endosomal maturation is critical for robust and timely cargo transport to specific cellular compartments. The most prominent model of early endosomal maturation involves phosphoinositide-driven gain or loss of specific proteins on individual endosomes, emphasising an autonomous and stochastic description. However, limitations in fast, volumetric imaging long hindered direct whole-cell measurements of absolute numbers of maturation events. Here, we use lattice light-sheet imaging and bespoke automated analysis to track individual very early (APPL1-positive) and early (EEA1-positive) endosomes over the entire population, demonstrating that direct interendosomal contact drives maturation. Using fluorescence lifetime, we show that this interaction is underpinned by asymmetric EEA1 binding to very early and early endosomes through its N- and C-termini, respectively. In combination with agent-based simulation that confirms a trigger-and-convert model, our findings indicate that APPL1-to EEA1-positive maturation is driven not by autonomous events but by heterotypic EEA1-mediated interactions, providing a mechanism for temporal and population-level control of maturation.

biophysics↗

bioRxiv2020

Generation of two Multipotent Mesenchymal Progenitor Cell Lines Capable of Osteogenic, Mature Osteocyte, Adipogenic, and Chondrogenic Differentiation

Differentiation of multi-potent mesenchymal progenitor cells give rise to several tissue types including bone, cartilage, and adipose. In addition to the complication arising from the numerous spatial, temporal, and hormonal factors that regulate lineage allocation, targeting of these cells in vivo is challenging, making mesenchymal progenitor cell lines valuable tools to study both tissue development and the differentiated cell types. Mesenchymal stem cells (MSCs) can be isolated from humans and animals; however, obtaining homogenous, responsive cells in a reproducible fashion can be problematic. As such, we have developed two novel mesenchymal progenitor cell (MPC) lines, MPC1 and MPC2, which were generated from the bone marrow of male C57BL/6 mice. These cells were immortalized using the temperature sensitive large T-antigen, allowing for thermal control of proliferation and differentiation. Both MPC1 and MPC2 cell lines are capable of osteogenic, adipogenic, and chondrogenic differentiation. Under osteogenic conditions both cell lines formed discrete mineralized nodules, staining for alizarin red and alkaline phosphatase, while expressing high levels of osteogenic genes including Sost, Fgf23, and Dmp1. Sost and Dmp1 mRNA levels were drastically reduced with parathyroid hormone, thus recapitulating in vivo responses. MPC cells secreted both the intact (iFGF23) and C-terminal (cFGF23) forms of endocrine hormone FGF23, which was upregulated in the presence of 1,25 dihydroxy vitamin D (1,25D). In addition to osteogenic differentiation, both cell lines also rapidly entered the adipogenic lineage, expressing several adipose markers after only 4 days in adipogenic media. MPC cells were also capable of chondrogenic differentiation, displaying increased expression of common cartilage genes including aggrecan, sox9, and cartilage oligomeric matrix protein. With the ability to differentiate into multiple mesenchymal lineages and mimic in vivo responses of key regulatory genes/proteins, MPC cells are a valuable model to study factors that regulate mesenchymal lineage allocation as well as the mechanisms that dictate transcription, protein modification, and secretion of these factors.

cell biology↗