Research

An Oxo-mechanical regulation of cell state

Across myriad habitats and diverse lifeforms, oxygen and mechanics are the most ubiquitously varying and profoundly influential environmental regulators. While physiological niches feature both varying oxygen levels and heterogeneous mechanics, laboratory experiments typically interrogate these as independent variables. Here, we show that combinatorial regimes defined by varying oxygen partial pressures and environmental mechanics—an oxo-mechanical cue—induce functionally-distinct cellular states in 3D ECM-like contexts. Single-cell morphometrics combined with multi-omics reveal that cellular response to oxygen deprivation depends on external mechanical milieus, whereas, cellular engagement with different mechanical microenvironments depends on oxygen availability. Independently perturbing both hypoxic signaling and cytoskeletal activity further reveals a reciprocal oxo-mechanical regulatory coupling, which operates by differentially altering the global chromatin accessibility for transcriptional regulation in response to specific combinations of oxygen partial pressures and external mechanical milieus. Together, our findings establish that a coupling between oxygen and mechanics drives the emergence of microenvironmentally-defined cell states. Read more about this work here.

A new force of selection: bacterial growth under 3D confinement

Bacteria reside in diverse natural environments with complex material properties, such as soil, infected tissues, and mucus. Since we observed them 300-odd years ago, a vast body of work has investigated how microbial communities in such niches are regulated by different selection pressures. However, most such experimental evidence derives from bacteria cultured in either homogeneously mixed liquid broths or 2D flat plates. Neither of these recreate the complex mechanical regimes of their natural settings, which are mostly disordered, granular, and porous 3D matrices with viscoelastic properties. To tackle this problem, we engineer transparent mucus-like mimics using a mechanically tunable synthetic hydrogel-based 3D growth medium that matches the viscoelastic shear moduli and pore-scale confinement of mucus. Using these, we discover that 3D growth under physical confinement selectively favors rod-shaped bacteria that form elongated colonies and enjoy increased access to nutrients, as opposed to spherical bacteria that form compact, rounded colonies. Importantly, we find that the structure of bacterial colonies in 3D space is dictated by the shape of a single cell - while rod-shaped bacteria form elongated and spread out colonies, spherical bacteria form compact and rounded colonies. Remarkably, this shape-dependent pattern allows rod-shaped bacteria to grow more successfully in highly confined 3D environments, helping them outcompete the spherical bacteria. 

This work provides the first-such experimental evidence that physical confinement plays a selective role in determining bacterial growth fitness – completely altering the way we traditionally think about how microbial populations survive and adapt across diverse ecological settings such as soil, aquifers, mucus, and infected tissues. Importantly, our research provides a new framework for understanding how the mechanical properties of an environment can actively regulate its resident biological matter. Read the paper here, as well as its coverage in the national media.  Read more about this work here.

Timing the size: physically controlling the cell cycle

The eukaryotic cell cycle is one of biology’s best-studied processes. And the budding yeast represents its very epitome. Naturally, understanding how this process can be dysregulated has massive implications for cell biology. Decades of research have shown how mutations, nutrient stresses, and compressive mechanical stresses alter the cell cycle progression in yeast. However, each of these typically impose either genetic, metabolic, or chemical insults that fundamentally alter the organism’s biological state while affecting the proliferative dynamics. But what if there was a way around - altering the division kinetics without perturbing the cell’s genome, metabolome, or biochemical signaling? 

In a brand new piece from our lab, we show a stunning example of purely physical regulation over cell cycle. Using tunable viscoelastic 3D growth media, we accomplish over 3-4 fold prolongations in cell cycle durations, without invoking any classical mechanosensory responses or any major alterations to the transcriptomic states. Rather, the degree of confinement changes the rate at which cells achieve volumetric growth during budding – which directly gates the division times! With significant implications for redefining how we understand cellular division and aging, our work signals an exciting new direction for exploring environmental regulation of proliferative growth. Read more here!

Squishy spheroids and crafty cells:
chemically controlling morphological stability and plasticity

Ovarian cancer – particularly high-grade serous ovarian carcinoma – is one of the most aggressive and challenging pathologies. Clinically, this often presents enigmatic forms, where metastatic cells organize into solid, morula-like masses or hollow, blastula-like ECM-covered masses. Whether and how these morphologically-distinct ensembles alter disease progression remains an open question. Moreover, the mechanistic determinants guiding the cellular reprogramming necessary to achieve such spatially well-defined aggregate organizations remain unclear.

Recently, we have characterized a key biochemical regulatory axis underlying the structural stability and morphological plasticity of these collective forms. We find that simple chemical cues – calcium and pH – exert a profound influence on the transitions between these forms, as well as re-program cells within these collectives. We also demonstrate a remarkable “structural memory” for the individual cells within these aggregates, which enables accelerated recovery of collective organizations even after catastrophic disruptions. Together, this work holds immense value towards identifying regulatory modules controlling both single cell states and multicellular organisation, as well as understanding how specialized cell populations emerge within multicellular ensembles. Read more about this work here.

To thrash or crawl:
environmental mechanics govern transitions in worm motion

Undulatory motion is a common locomotory mechanism, manifesting from worms to snakes. The mechanics of this have long fascinated physicists and biologists, given the immense complexities of efficiently achieving energetically-expensive bodily contortions while simultaneously exerting neuro-muscular control that also takes into account environmental feedback. A classic model for studying undulatory motion is the C. elegans – by itself a workhorse for development biology and neurobiology. On flat agar pads and in liquids, this worm typically exhibits either crawling-like or thrashing-like swimming behaviors, which are achieved via muscular activity leading to undulations of its body. While the neuronal basis for these behaviors has been very well characterized, how more complex mechanical regimes – especially granular and viscoelastic 3D environments such as soil, where C. elegans naturally reside – influence its motion remain insufficiently understood. 

By engineering a mechanically tunable 3D medium which spans over three orders of magnitude in its degree of confinement, we achieve the first-such systematic interrogation of worm motion within granular viscoelastic systems. Intriguingly, we start observing a distinct non-monotonic dependence of the worm swimming speeds on the mechanical properties of the environment – as the degree of confinement increases, worms initially speed up before slowing down. Combining slender body theory and quantitative experiments to construct a non-dimensionalized phase space, we find that worms optimize for the efficiency of motion as the degree of physical confinement increases. Crucially, we capture a smooth transition from thrashing-like behavior under low confinement to crawling-like behavior under higher confinement – which is unprecedented across such a broad mechanical regime. This work holds significant interest towards exploring how complex mechanical environments alter behavioral outcomes, as well as raises interesting future questions concerning the neuromuscular basis of tradeoffs between optimal speeds and optimal efficiency of motion. Read more about this work here.

Jammed agarose microgel as 3D cell growth media

Jammed microgels are a promising class of biomaterials well-suited for 3D cell culture, tissue bioengineering, and 3D bioprinting. However, existing protocols for fabricating such microgels either involve complex synthesis steps, long preparation times, or polyelectrolyte hydrogel formulations that sequester ionic elements from the cell growth media. Hence, there is an unmet need for a universally biocompatible, high-throughput, and widely accessible manufacturing process. We address these demands by introducing a rapid, high-throughput, and remarkably straightforward method to synthesize jammed microgels composed of flash-solidified agarose granules directly prepared in a culture medium of choice. Read more about this work here.

Morphological instability and roughening of growing 3D bacterial colonies

The morphogenesis of two-dimensional bacterial colonies has been well studied. However, little is known about the colony morphologies of bacteria growing in three dimensions, despite the prevalence of three-dimensional environments (e.g., soil, inside hosts) as natural bacterial habitats. Using experiments on bacteria in granular hydrogel matrices, we find that dense multicellular colonies growing in three dimensions undergo a common morphological instability and roughen, adopting a characteristic broccoli-like morphology when they exceed a critical size. Analysis of a continuum “active fluid” model of the expanding colony reveals that this behavior originates from an interplay of competition for nutrients with growth-driven colony expansion, both of which vary spatially. These results shed light on the fundamental biophysical principles underlying growth in three dimensions. Read more about this work here.

Chemotactic smoothing of collective migration

Collective migration—the directed, coordinated motion of many self-propelled agents—is a fascinating emergent behavior exhibited by active matter with functional implications for biological systems. However, how migration can persist when a population is confronted with perturbations is poorly understood. Here, we address this gap in knowledge through studies of bacteria that migrate via directed motion, or chemotaxis, in response to a self-generated nutrient gradient. We find that bacterial populations autonomously smooth out large-scale perturbations in their overall morphology, enabling the cells to continue to migrate together. This smoothing process arises from spatial variations in the ability of cells to sense and respond to the local nutrient gradient—revealing a population-scale consequence of the manner in which individual cells transduce external signals. Altogether, our work provides insights to predict, and potentially control, the collective migration and morphology of cellular populations and diverse other forms of active matter. Read more about this work here. 

Bacterial Chemotaxis in Porous Media

Chemotactic migration of bacteria—their ability to direct multicellular motion along chemical gradients—is central to processes in agriculture, the environment, and medicine. However, studies are typically performed in bulk liquid, despite the fact that most bacteria inhabit heterogeneous porous media such as soils, sediments, and biological gels. By using direct visualization and 3D bioprinting, we find that cellular chemotaxis drives collective migration while confinement in a porous medium fundamentally alters chemotactic migration in two ways. First, cells bias their motion through a different primary mechanism in confinement than in bulk liquid. Second, confinement markedly alters the dynamics and morphology of the migrating population—features that can be described by a continuum model, but only when standard motility parameters are substantially altered from their bulk liquid values. Our work thus provides a framework to predict and control the migration of bacteria, and active matter in general, in heterogeneous environments. Read more about this work here. 

Bacterial Dynamics in 3D Porous Media

While bacterial motility is well-studied for motion on flat surfaces or in unconfined liquid media, most bacteria are found in heterogeneous porous media, such as biological gels and tissues, soils, sediments, and subsurface formations. Understanding how confinement alters bacterial motility is therefore critical to model the progression of infections, apply beneficial bacteria for drug delivery, and bioremediation. Unconfined bacteria move via runs and tumbles, leading to random walk-like motion; in a porous medium, previous research has assumed bacteria still move via runs and tumbles, but with a reduced diffusivity due to collisions with obstacles. However, this assumption has never been directly tested due to the inability to visualize processes in opaque 3D media. Here, we directly visualize the motion of single E. coli cells inside a model 3D porous medium, having controlled pore structure. By analyzing the individual cell trajectories, we find that the bacteria do not move via a run and tumble process, but instead via intermittent hops and traps reminiscent of thermally-activated transport in disordered media. We will present how bacterial motility depends sensitively on pore-scale confinement. Our findings overturn standard assumptions made in the field and provide guidance for the development of more accurate macroscopic models of bacterial motion. Our recent work can be found in Nature Communications and Soft Matter.