Eric Sembrat's Test Bonanza

One of the fundamental problems in biology is understanding how phenotypic variations arise in individuals. Phenotypic variation is generally attributed to genetic or environmental factors. However, in several important cases, phenotypic variations are observed even among genetically identical cells in homogeneous environments. Recent research indicates that such `non-genetic individuality' can arise due to intrinsic stochasticity in the process of gene expression. Correspondingly there is a need to develop a framework for quantitative modeling of stochastic gene expression and its regulation. Of particular interest is modeling of regulation by non-coding RNAs, which is often a critical component of cellular processes such as development, differentiation and cancer.

In this talk, I will discuss approaches developed by my group that lead to new analytical results for stochastic models of gene expression. In biologically relevant limits, we develop a mapping to queueing theory to derive exact results for general models of stochastic gene expression. Focusing on specific regulatory mechanisms, we propose and analyze a comprehensive model for regulation by non-coding RNAs.  The results obtained provide new insights into the role of non-coding RNAs in fine-tuning the noise in gene expression. I will conclude with a discussion of protocols for inferring gene expression parameters from observations of mRNA and protein distributions.

The ribosome translates the genetic information encoded in messenger RNA into protein. Folded structures in the coding region of an mRNA represent
a kinetic barrier that lowers the peptide elongation rate, as the ribosome must disrupt structures it encounters in the mRNA to allow translocation to the next codon. Such structures are exploited by the cell to create diverse strategies for translation regulation. Although strand separation activity is inherent to the ribosome, requiring no exogenous helicases, its mechanism is still unknown. By using a single-molecule optical tweezers assay to follow in real time the codon-by-codon translation of mRNA hairpins, we conducted a quantitative characterization of the effect of the RNA structural stability on the peptide elongation rate, which revealed distinct mechanisms utilized by the ribosome to unwind mRNA structures. Our results establish a quantitative mechanical basis for understanding the mechanism of translational regulation of the elongation rate by structured mRNAs.

Cadherins constitute a large family of Ca2+-dependent adhesion molecules in the Inter-cellular junctions that play a pivotal role in the assembly of cells into specific three-dimensional tissues.  Although the molecular mechanisms underlying cadherin-mediated cell adhesion are still not fully understood, it seems likely that both cis dimers that are formed by binding of extracellular domains of two cadherins on the same cell surface, and trans-dimers formed between cadherins on opposing cell surfaces, are critical to trigger the junction formation.

Here we present a new multiscale computational strategy to model the process of junction formation based on the knowledge of cadherin molecular structures and its 3D binding affinities. The cell interfacial region is defined by a simplified system where each of two interacting membrane surfaces is represented as a two-dimensional lattice with each cadherin molecule treated as a randomly diffusing unit. The binding energy for a pair of interacting cadherins in this two-dimensional discrete system is obtained from 3D binding affinities through a renormalization process derived from statistical thermodynamics. The properties of individual cadherins used in the lattice model are based on molecular level simulations. Our results show that within the range of experimentally-measured binding affinities, cadherins condense into junctions driven by the coupling of cis and trans interactions. The key factor appears to be a loss of molecular flexibility during trans dimerization that increases the magnitude of lateral cis interactions.

We have also developed stochastic dynamics to study the adhesion of multiple cells. Each cell in the system is described as a mechanical entity and adhesive properties between two cells are derived from the lattice model. The cellular simulations are used to study the specific problems of tissue morphogenesis and tumor metastasis. The consequent question and upcoming challenge is to understand the functional roles of cell adhesion in intracellular signal transduction.

Clathrin-coated vesicles are the most prominent carriers of membrane traffic from cell surface to endosomes (endocytosis), a pathway by which hormones, transferrin, immunoglobulins, LDL, viruses, and their receptors enter cells. They are also important for traffic between endosomes and the trans-Golgi network. In this presentation, I will discuss (i) technological and analytical advances that I developed to directly visualize clathrin-mediated membrane traffic in three dimensions and in living cells; (ii) data obtained using these advances that defined a role for actin filament polymerization in counteracting membrane tension during clathrin-coated vesicle budding at the apical surface of polarized epithelial cells; and (iii) how these advances can be used to study a wide variety of biological processes that occur in living cells and tissues.

Living organisms are capable to sense and adapt to a wide range of environmental changes, which is essential for their survival in nature. Although numerous molecular circuits have been evolved to accomplish this sensory adaptation function in different organisms, these circuits share intrinsically the same logical construct. Using Escherichia coli cells as model system, we combined theoretical techniques with experiments to formulate biological sensory adaptation. We demonstrated that E. coli cells accurately “remember” the chemical environment by differentiating and encoding external signals into molecular modifications on specific sensory receptors. We also discovered that E. coli cells adjust their chemical sensitivity via tuning the sensory machinery assembly according to the environment. Moreover, by evaluating the energetic cost associated with sensory adaptation, we were able to derive the exponential tradeoff relation between the benefit (adaptation accuracy & adaptation speed) and the cost (energy dissipation). We believe that this set of approaches sketch a general framework for studying various biological regulatory circuits and the obtained benefit-to-cost relations could shed light on the design principles and the evolution of regulatory circuits.

The heart is an electro-mechanical system in which, under normal conditions, electrical waves propagate in a coordinated manner to initiate an efficient contraction. In pathologic states, propagation can destabilize and exhibit period-doubling bifurcations that can result in both quasiperiodic and spatiotemporally chaotic oscillations. In turn, these oscillations can lead to single or multiple rapidly rotating spiral or scroll waves that generate complex spatiotemporal patterns of activation that inhibit contraction and can be lethal if untreated. Despite much study, little is known about the actual mechanisms that initiate, perpetuate, and terminate reentrant waves in cardiac tissue.

In this talk, I will discuss experimental and theoretical approaches to understanding the dynamics of cardiac arrhythmias. Then I will show how state-of-the-art voltage-sensitive fluorescent dyes can be used to image the electrical waves present in cardiac tissue, leading to new insights about their underlying dynamics. I will establish a relationship between the response of cardiac tissue to an electric field and the spatial distribution of heterogeneities in the scale-free coronary vascular structure. I will discuss how in response to a pulsed electric field E, these heterogeneities serve as nucleation sites for the generation of intramural electrical waves with a source density ρ(E) and a characteristic time τ for tissue excitation that obeys a power law. These intramural wave sources permit targeting of electrical turbulence near the cores of the vortices of electrical activity that drive complex fibrillatory dynamics. Therefore, rapid synchronization of cardiac tissue and termination of fibrillation can be achieved with a series of low-energy pulses. I will finish with results showing the efficacy and clinical application of this novel mechanism in vitro and in vivo.

Embryogenesis and regeneration are among the most striking and beautiful phenomena in nature. For a physicist, this brings together many major themes—pattern formation, information processing, the mechanics of complex fluid-like materials—that are essential for our understanding of life more broadly. In my talk I will give two examples on the important role of tissue mechanics for these phenomena.

First, I will discuss how a macroscopic tissue property, specifically tissue surface tension, is connected to the properties of the constituent cells, such as cortical tension and adhesion. I will directly compare theoretical predictions with experimental data and discuss the relationship between tissue surface tension and tissue dynamics using primarily zebrafish embryonic tissues as the experimental system.

In the second part of my talk, I'll switch gears over to regeneration and asexual reproduction in planarians. Asexual reproduction and the ability to regenerate are intrinsically connected, but despite this important link, little is known about the physical process of reproduction due to experimental difficulties. We have overcome some of these difficulties and I will present preliminary data on the physical mechanisms of dividing planarians. Finally, I will discuss our current understanding of the asexual population dynamics based on a large-scale experiment in which we have been tracking >10,000 reproductive events over the course of >2.5 years and up to 51 generations using a custom-built Scan-Add-Print database system.

Self-assembly of amphiphilic peptides designed during the last ten years by different research  groups leads to a large variety of 3D-structures that already found applications in e.g. stabilization of large protein complexes, cell culturing systems etc. Our group has recently suggested a new type of short amphiphilic peptides that exhibits clear charge separation controllable by the pH of the environment. An intricate interplay between electrostatic and hydrophobic interactions and the packing parameter of the peptide molecule leads to a rich pattern of self-assembling behavior ranging from nucleated and pH-dependent self-assembly into tubular and spherical micelles up to pH-independent isodesmic polymerization into thin ribbons.

Another interesting development came from one of the short antimicrobial peptides (AMP), indolicidin. We found that indolicidin, as well as some of its derivatives can assemble on the DNA surface forming smooth and continuous coverage. In nature this phenomenon might be responsible for efficient knocking down the DNA replication and transcription processes in the invading cells while from nanotechnological prospective, it can help designing functional DNA electronics.

1. L. Gurevich, T.W. Poulsen, O. Z. Andersen, N. L. Kildeby and P. Fojan, “pH-dependent self-assembly of the short surfactant-like peptide KA6”, J.Nanoscience and Nanotechnology 10, 7946-7950 (2010)

This talk will describe new results on the properties of colloidal crystals, both on their solidification and on their melting.  It will describe how hard-sphere like colloids crystallize, and will explore the huge discrepancy between the nucleation rates predicted by theory and measured in simulation and those measured experimentally.  The discrepancy can be as large as 150 orders of magnitude!  A simple modification to the theory, suggested by experiment, is able to account for this behavior and to rectify the discrepancy.  It will also describe how perfect colloidal crystals, formed in a Wigner lattice, melt.  Since there are no grain boundaries for the crystals, melting occurs in a different fashion, one that seems to have some second order character to it.

There are over 28,000 species of fishes, and a key feature of this remarkable evolutionary diversity is a great variety of propulsive systems used by fishes for maneuvering in the aquatic environment.  Fishes have numerous control surfaces (fins) which act to transfer momentum to the surrounding fluid.  In this presentation I will discuss the results of recent experimental kinematic and hydrodynamic studies of fish fin function, and their implications for the construction of robotic models of fishes.  Recent high-resolution video analyses of fish fin movements during locomotion show that fins undergo much greater deformations than previously suspected and fish fins possess an clever active surface control mechanism.  Fish fin motion results in the formation of vortex rings of various conformations, and quantification of vortex rings shed into the wake by freely-swimming fishes has proven to be useful for understanding the mechanisms of propulsion. Experimental analyses of propulsion in freely-swimming fishes have led to the development of a variety of self-propelling robotic models: pectoral fin and caudal fin (tail) robotic devices, and a flapping foil model fish of locomotion.  Data from these devices will be presented and discussed in terms of the utility of using robotic models for understanding fish locomotor dynamics.