Physical Biology Circle Meeting

Biological systems not only have the remarkable capacity to build and maintain complex spatio-temporal structures in noisy environments, they can also rapidly break up and rebuild such structures. Here, using primitive societies of Polistes wasps as a model system we show that both robust specialisation and rapid plasticity are emergent properties of a multi-scale dynamics. We employ a unique strategy combining theory with an experimental approach that, after perturbing the social structure by removing the queen, correlates time-resolved multi-omics to video recordings at the level of individual insects during the re-establishment of the social steady state. Using theory, we show that the interplay between molecular and colony-scale processes allows Polistes to be stable against intrinsic perturbations of molecular states while reacting plastically to extrinsic cues affecting the society as a whole. Long-term stability of the social structure is further facilitated by dynamic DNA methylation. Our study provides a general principle of how both specialization and plasticity can be achieved in biological systems.

DNA methylation is an epigenetic modification of cytosines in a CpG context. In important biological contexts, such as during the loss of pluripotency during early development and the differentiation of blood stem cells antagonistic players of the DNA methylation machinery are co-expressed, resulting in active turnover of DNA methylation. The functional relevance of such turnover is not understood. Here, we show that the interplay between long-range interactions resulting from methylation dependent chromatin states and irreversibility due to active turnover facilitates kinetic proof reading of DNA methylation patterns. Specifically, we use analytical calculations we predict an order of magnitude effect of these processes on the fidelity of the recapitulation of an underlying binding energy landscape for plausible biological parameters. We qualitatively confirm our theoretical findings using single-cell sequencing of mutant embryonic stem cells.

Entropy production plays a fundamental role in the study of non-equilibrium systems by offering a quantitative handle on the time-reversal symmetry breaking induced by energy dissipation at microscopic spatio-temporal scales. The question of whether features of this local dissipation propagate across scales to induce genuinely non-equilibrium (rather than equilibrium-like) macroscopic behaviour is however beyond the scope of the existing entropy production toolkit. We address this question by introducing a systematic procedure to characterise the scaling of entropy production under iterative coarse graining, which we apply to a minimal driven-diffusion model on lattices of arbitrary dimensions. Our approach unveils a natural criterion to distinguish equilibrium-like and genuinely non-equilibrium macroscopic phenomena based on the sign of the scaling exponent of the entropy production per mesostate.

Living cells maintain and change shape, adhere, move and divide. At the basis of these processes there is the cytoskeleton, a dynamic meshwork of filamentous protein assemblies. The proteins involved in the polymerisation of those filaments are actin and tubulin. The filaments interact with molecular motors. The motors interacting with actin and tubulin filaments are respectively called myosin and kinesin. These motors consume adenosine triphosphate (ATP) in order to generate mechanical stress. Of particular interest is a thin actin layer beneath the plasma membrane, which goes under the name of actin cortex. Indeed, it plays an important role in the cellular morphogenesis and determines the mechanical properties of the cells. Also, instability of the actin cortex might generate dynamic protrusions and the contractile ring that cleaves animal cells into two daugther cells during division. We study the dynamics of the actin filaments in this layer and the potential impacts of actin cortex instabilities. For this purpose, we use a hydrodynamic description of active gels. Depending on the contractile activity, its anisotropy and the friction of the gel along the membrane we observe five different phases. In addition, we obtained an orientation change of actin filaments between the membrane and the inner boundary of the actin cortex, physically corresponding to a both nematic and polar phase transition.

TBA

The structure of the spindle apparatus is conserved among metazoans, with two spindle poles flanking a central region of parallel or anti-parallel aligned microtubules. Force-producing machinery is involved in maintaining the organization and proper functioning of this array. These motors drive polarity sorting and focus the poles. However, our understanding of the contribution of different motor activities to successful spindle assembly is still in progress; experiments with purified proteins can help clarify the activities of spindle component subsets. We purify two mitotic kinesins which reconstitute extensile and contractile phases: the plus-end-oriented KIF11, and the minus- end-oriented HSET. Previously our lab showed that purified KIF11 and HSET can drive extension and contraction, respectively, mimicking separately the organizational modes of the extensile midzone and contractile poles of the spindle (Roostalu and Rickman, et al, 2018). Here, varying the ratio of the oppositely-directed motors in a combined experiment showed a sharp transition between contraction and extension of the resulting network. Coexistence of the phases was not observed, indicating that further control is required, with respect to control of microtubule dynamics, physical constraints, or localization of activities. The resulting phase space nonetheless reveals unexpected interplay or cooperation of the two motor activities in some regimes.

Living cells host many membrane-less compartments which originate via liquid-liquid phase separation in cells. From theoretical perspective much is known about classical emulsions, which coarsen through fusion or Ostwald ripening. Recent experimental work revealed a new class of active emulsions with controllable lifetime and droplet kinetics. This class of active emulsions involves a fuel that drives a chemical reaction from thermodynamically stable precursor molecules to metastable building blocks. In recent years we have shown that this class of emulsions exhibit accelerated Ostwald ripening. Further, a population of such emulsions can exert a parasitic-host behavior, that changes the lifetime of the emulsion and its robustness against the environment. Our most recent study focuses on theoretical analysis how the distribution of droplet sizes differs in different classes of active emulsions, for which the growth law is nontrivial. Current finding is a bimodal distribution of droplet sizes, where we see a simultaneous enhancement of smaller and larger droplets in the system. The bimodality arises as the larger droplets grow faster than the smaller ones. The peak at smaller radii of the bimodal distribution vanishes due to the dissolution of smaller droplets because of the fast increase of the critical radius or when the changes in the critical radius cannot outcompete the growth of the smaller droplets that ripen by the Ostwald law. This new characteristic of the droplet size distribution in active emulsions is a new mechanism to control the populations of emulsions. Our results pave a way to construct an origin of life model of interacting populations of active emulsions.

In vertebrates, the sense of balance and hearing rely on “cellular microphones”: the hair cells. On the top of their cell body, hair cells have a bundle of few tens of micrometric actin filled cilia suitably named hair bundle. The hair bundle is a mechanical sensor. A sound, transmitted as a force, will make the cilia pivot and open mechanosensitive ion channels in the cilia membrane. Potassium and calcium ions in the medium flow in and change the cell membrane potential; sound is detected. For my PhD project, I am studying the mechanics of hair bundles. Using the frog as a model organism, it is possible to study hair bundles in ex vivo preparations wherein physiological ionic environments can be recreated. Thanks to microfabricated glass fibers, it is also possible to apply a force of few piconewtons to a single hair bundle leading to a deflection of several tens of nanometers. In these conditions, hair bundle stiffness depends on the transduction current, therefore the mechanics is nonlinear. Close to the extreme values of transduction current, when ion channels are all closed or all opened, stiffness is constant. Conversely, when half of the ion channels are opened, the hair bundle is softer. Stiffness can even be negative, the hair bundle is not stable anymore for this fraction of opened channels. Previous studies have shown that this nonlinearity coupled to a mechanism of adaptation makes the bundle behave as an active oscillator: it oscillates spontaneously and amplifies weak sinusoidal stimuli at the frequency of spontaneous oscillations. Nevertheless, in other vertebrate species and in less controlled ionic environment, hair bundle mechanics is linear even if the ion channels can open and close. This suggests that other parameters control the interplay between ion channel opening and hair bundle mechanics. My project aims to study how nonlinearity evolves upon change of experimental conditions. Because it is finely tuned in the inner ear of mammals, we chose to vary the voltage difference between the fluid around the hair bundle and the fluid on the other side of the cell body. I have preliminary results suggesting that the nonlinearity amplitude significatively changes with the voltage difference. These changes are not predicted by the standard model describing hair bundle mechanics. Therefore, If they were to be confirmed, my results would question the classic picture of the transduction machinery. Furthermore they highlight how important it is to control experimental conditions in order to observe an eventual nonlinearity in hair bundle mechanics.

The Drosophila wing undergoes a series of complex shape changes during its development. One of these events is known as eversion in which the wing disc pouch, an epithelial monolayer tissue undergoes shape change from a hemispherical configuration to a flattened structure with the two dome flanks facing each other. Segmentation of Imaging data of the apical cell area distributions show an interesting area gradient which changes during development. Using a computational spring lattice model, we wish to check whether this change in cell area distributions can explain the 3D shape change during eversion.

Living cells use compartments(droplets) to spatially organise molecules that can undergo fuel-driven chemical reactions. Not much is known about the mechanisms underlying such spatial control of chemical reactions and how much the properties of chemical reactions are altered by the compartments relative to homogeneous systems. Here, we derive a theoretical framework to study fuel driven chemical reactions in the presence of compartments.We study two state transitions like phosphorylation via hydrolysis of ATP and enzymatic reactions. For two state transitions, we find that the ratio of phosphorylated product can be regulated by droplets by two orders of magnitude relative to the homogeneous state. In the case of enzymatic reactions, we show that the initial rate of product formation can be increased by more than ten fold. We further calculate analytically the optimal conditions of designing the system. Our studies exemplify the enormous potential of phase separated compartments as biochemical reactors in living cells and enhancing the effect of enzymes. Understanding the control of biochemical reactions via compartments is key to elucidate the functionality of stress granules for the cell and is also crucial for biochemical communication among synthetic cells and RNA catalysis in coacervate protocells.

The plasma membrane and the underlying actomyosin cortex constitute the surface of animal cells, and their mechanical properties are key for a plethora of cell processes. Although they are often studied as independent structures, the plasma membrane and the cortex are physically tethered to each other via linker proteins, globally referred to as membrane-to-cortex attachment (MCA). MCA contributes to membrane tension and regulates biological processes such as cell migration and stem cell differentiation. Nevertheless, it remains the most elusive element of the animal cell surface. Aiming at investigating MCA in the context of cell surface mechanics, we have developed artificial signaling-inert linker proteins to specifically perturb it. Strikingly, we have found an unforeseen role for MCA in the regulation of cortex mechanics. Specifically, it modulates cortical tension and cortical stiffness in a density-dependent manner. We further employed in-cell cryo-electron tomography to decipher how MCA affects the architecture of the cortex in cells. Finally, we show the relevance of our findings for early mouse embryo cell specification.

TBA

Many compartments in cells are protein-rich biomolecular condensates demixed from the cyto- or nucleoplasm. Although the defining feature of a condensate is its molecular composition, traditional approaches to measure this quantitatively are inefficient or require confounding labels. By combining quantitative phase microscopy (QPM) with the physics of sessile droplets, we developed a label-free method to measure the shape and composition of micron-sized model condensates. This method has a precision better than 2%, requires 1000-fold less material than bulk approaches, and exposes systematic errors as large as 40-fold in common estimates based on fluorescence intensity ratios. In addition to salt- and temperature-dependent binodals, the method reveals complex aging dynamics in individual condensates as well as a large sequence-encoded diversity of compositions in condensates formed with distinct full-length proteins. We anticipate that this new approach will enable understanding the physical properties of biomolecular condensates and their function.

To navigate through tissues, migrating cells must balance persistent migration with changes in direction to circumvent obstacles. The protrusion at the leading edge of a moving cell is driven by actin polymerization and displays dynamic membrane patterns that constantly probe the environment. Here, we asked whether cells read out their membrane topography to decode their environment and decide if they should move ahead or turn away. To test this, we first parametrized the curvature of the plasma membrane of migrating immune cells to match microscopy data. Then, we created a theoretical model to explore what types of feedback between topography and protrusion could lead to the observed patterns. Our model predicts that negative coupling of positive (inward) curvature of the plasma membrane to actin polymerization would explain our data. To identify the putative positive curvature sensor, we screened for proteins with suitable membrane binding domains that are expressed during cell migration. We found that the BAR domain protein Snx33 localized to the leading edge, and destabilized it by inhibiting the major actin nucleation promoting factor WAVE2. This mechanism was indeed required for navigation, as Snx33 knockout cells fail to change direction when hitting inert or cellular obstructions but continued to migrate persistently. Our results show how cells can read out their surface topography to interpret their environment, allowing them to rapidly switch between persistent and exploratory migration in order to circumvent obstacles.

The actin cortex is an active adaptive material, embedded with complex regulatory networks that can sense, generate and transmit mechanical forces. The cortex can exhibit a wide range of dynamic behaviours, from generating pulsatory contractions and traveling waves to forming highly organised structures such as ordered fibers, contractile rings and networks that must adapt to the local cellular environment. Despite the progress in characterising the biochemical and mechanical components of the actin cortex, our quantitative understanding of the emergent dynamics of this mechanochemical system is limited. Here we develop a mathematical model for the RhoA signalling network, the upstream regulator for actomyosin assembly and contractility, coupled to an active polymer gel, to investigate how the interplay between chemical signalling and mechanical forces govern the propagation of contractile stresses and patterns in the cortex. We demonstrate that mechanical feedback in the excitable RhoA system, through dilution and concentration of chemicals, acts to destabilise homogeneous states and robustly generate pulsatile contractions. While moderate active stresses generate spatial propagation of contraction pulses, higher active stresses assemble localised contractile structures. Moreover, mechanochemical feedback induces memory in the active gel, enabling long-range propagation of transient local signals.

Hearing relies on the operation of cellular microphones—the hair cells of the inner ear —to transduce mechanical vibrations evoked by sound into electrical signals. At the level of a single hair cell, mechanoelectrical transduction results from the deflection of the hair bundle—a tuft of a few tens to a few hundred stereocilia that protrude from the apical surface of each hair cell into the surrounding fluid. Hair-bundle deflections modulate tension in proteinaceous “tip links” that interconnect the stereocilia near their tips and pull on mechanosensitive ion channels, evoking an electrical response. There is currently no detailed understanding about how the individual contributions of the tip links sum up to shape the collective response of a hair cell, in particular in the mammalian cochlea where the hair bundles are known to be weakly cohesive. A first aim of this PhD project is to characterize individual stereocilia movements as well as the field of tension in the tip links. In addition, past evidence suggests that the mechanical properties of hair bundles, which condition the ear’s sensitivity and frequency selectivity to sound, strongly depend on their complex local ionic environment. In the case of the mammalian hearing organ—the cochlea, reproducing such environment ex-vivo remains an experimental challenge, resulting in a long-standing limitation for the study of hair-cell mechanosensitivity under physiological conditions. In this PhD project, I aim at developing a microfluidic ear-on-chip approach to provide unprecedented control over key physiological parameters of the hair cells, while enabling mechanical stimulation of single hair bundles to characterize their mechanosensitivity. A combination of innovative microfluidic tools with numerical simulations will shed light on the physiological and physical parameters that control the mechanosensitivity of the hair bundle.

Animal morphogenesis is a highly dynamical process spanning multiple spatial and temporal scales. It is well known that external environmental factors affect this developmental process leading to a variety of morphological phenotypes. The behavior an animal displays is often considered an emergent effect, stemming from developmental events. How organismal scale behavior during development effects morphogenesis is only beginning to be studied. Investigating the connection and coordination between organismal behavior and morphogenesis has so far been challenging due to lack of right multimodal optical technologies and biological systems of right level of complexity to study these processes with sufficient spatio-temporal resolution. Owing to its size, body plan, optical access as well as established tools for culturing and experimental manipulation, Nematostella Vectensis - an organism from the cnidarian phylum, is a promising model system to study morphobehavioural development. Here we present a tailored implementation of a known optical imaging, namely Optical Coherence Microscopy (OCM) that enables quick, label free, volumetric imaging at organismal scales to study mesoscale animal morpho-behavior. Using the volumetric imaging we could quantitatively show a significant increase in the body cavity size during the larva to adult transition in Nematostella driven by the behavioural hydraulics of the developing animal. We further establish the ontogenic and static morphological scaling relationships as the embryo develops into a polyp - a foundation for further mechanistic studies of allometric scaling. OCM technology complements our custom selective plane illumination microscope (SPIM) to perform fluorescence imaging. The multimodal technique opens up possibilities of live 3d structural and functional imaging to study ethology of development.

The auditory organ of chick, called the basilar papilla, comprises of two terminally differentiated cells: sensory hair cells and non-sensory support cells. In prenatal development, these cells arrange into a highly regular pattern where hair cells stand isolated from each other separated by support cells. This patterning of the hair cells becomes even more pronounced as the apical surface area of the hair cells increases 10-fold. We suggest that non-muscle myosin, found to be located at certain junctions, drives this regular packing by active contraction. Here, we use a mathematical vertex model, that considers mechanical forces at the scale of single cells and cellular junctions, to identify the contributions of forces, that are actively expressed by cells, to the process of cellular rearrangement. Active forces of cell size regulation and myosin contraction stand in contrast to global shear forces, which are known to be involved in similar processes of tissue reorganisation but are experimentally only observed to a limited extend in the basilar papilla.

Viruses can establish both acute and persistent chronic infections, and some viruses have developed the ability to switch between the two. However, the molecular mechanisms that trigger a transition from a benign chronic infection into pathogenesis remain unknown. Here we investigated the role of the cellular stress response in provoking a chronic-to-acute transition in viral replication in a model of mumps infection. Using a combination of cell biology, whole-cell proteomics and cryo-electron tomography we show that stress induces phosphorylation of the disordered viral Phosphoprotein, which we suggest facilitates partitioning of the viral polymerase into preformed viral condensates, constituting the core components of the viral replication machinery. This occurs concomitantly with a conformational change in the viral nucleocapsids that exposes the viral genome and can further facilitate its replication. We propose that these changes in the viral condensate upon exogenous stress, accompanied by downregulation of the host antiviral response, provide an environment that supports upregulation of viral replication and virion release. Thus, we elucidate molecular and structural mechanisms of a stress-mediated switch that disrupts the equilibrium between the virome and the host in chronic infection.

In living cells, protein-rich condensates can wet the cell membrane and surfaces of membrane-bound organelles. Interestingly, many phase-separating proteins also bind to membranes leading to a molecular layer of bound molecules. Here we investigate how binding to membranes affects surface phase transitions such as wetting and prewetting. We derive a thermodynamic theory for a three-dimensional bulk in the presence of a two-dimensional, flat membrane. Above the saturation concentration, we find that membrane binding facilitates complete wetting and lowers the wetting angle. Moreover, below the saturation concentration, binding facilitates the formation of a thick layer at the membrane and thereby shifts the prewetting phase transition far below the saturation concentration. The distinction between bound and unbound molecules near the surface leads to a large variety of prewetted states. Our work suggests that surface phase transition combined with molecular binding represents a versatile mechanism to control the formation of protein-rich domains at intra-cellular surfaces.

We investigate alignment of $d$-dimensional vectors on a sphere subject to coupling and correlated noise forces. Alignment of vectors on the circle in two dimensions due to common noise is known as noise-induced synchronization. In general, common noise and attractive coupling promote alignment or correlation between different vectors while individual noise and repulsive coupling have the opposite effect. The limit cases of vanishing coupling, vanishing individual noise or vanishing common noise can be treated analytically. The exact distribution of the scalar product between two vectors subject to correlated noise can be viewed as an example of the Moran effect in a non-trivial dynamical system.