Phase resetting reveals network dynamics underlying a bacterial cell cycle

Genomic and proteomic methods yield networks of biological regulatory interactions but do not provide direct insight into how those interactions are organized into functional modules, or how information flows from one module to another. In this work we introduce an approach that provides this complementary information and apply it to the bacterium Caulobacter crescentus, a paradigm for cell-cycle control. Operationally, we use an inducible promoter to express the essential transcriptional regulatory gene ctrA in a periodic, pulsed fashion. This chemical perturbation causes the population of cells to divide synchronously, and we use the resulting advance or delay of the division times of single cells to construct a phase resetting curve. We find that delay is strongly favored over advance. This finding is surprising since it does not follow from the temporal expression profile of CtrA and, in turn, simulations of existing network models. We propose a phenomenological model that suggests that the cell-cycle network comprises two distinct functional modules that oscillate autonomously and couple in a highly asymmetric fashion. These features collectively provide a new mechanism for tight temporal control of the cell cycle in C. crescentus. We discuss how the procedure can serve as the basis for a general approach for probing network dynamics, which we term chemical perturbation spectroscopy (CPS)

Programmable base editing of zebrafish genome using a modified CRISPR-Cas9 system.

Precise genetic modifications in model animals are essential for biomedical research. Here, we report a programmable "base editing" system to induce precise base conversion with high efficiency in zebrafish. Using cytidine deaminase fused to Cas9 nickase, up to 28% of site-specific single-base mutations are achieved in multiple gene loci. In addition, an engineered Cas9-VQR variant with 5'-NGA PAM specificities is used to induce base conversion in zebrafish. This shows that Cas9 variants can be used to expand the utility of this technology. Collectively, the targeted base editing system represents a strategy for precise and effective genome editing in zebrafish.The use of base editing enables precise genetic modifications in model animals. Here the authors show high efficient single-base editing in zebrafish using modified Cas9 and its VQR variant with an altered PAM specificity

Functional Roles of Pulsing in Genetic Circuits

A fundamental problem in biology is to understand how genetic circuits implement core cellular functions. Time-lapse microscopy techniques are beginning to provide a direct view of circuit dynamics in individual living cells. Unexpectedly, we are discovering that key transcription and regulatory factors pulse on and off repeatedly, and often stochastically, even when cells are maintained in constant conditions. This type of spontaneous dynamic behavior is pervasive, appearing in diverse cell types from microbes to mammalian cells. Here, we review recent work showing how pulsing is generated and controlled by underlying regulatory circuits and how it provides critical capabilities to cells in stress response, signaling, and development. A major theme is the ability of pulsing to enable time-based regulation analogous to strategies used in engineered systems. Thus, pulsatile dynamics is emerging as a central, and still largely unexplored, layer of temporal organization in the cell

Combinatorial gene regulation by modulation of relative pulse timing

Studies of individual living cells have revealed that many transcription factors activate in dynamic, and often stochastic, pulses within the same cell. However, it has remained unclear whether cells might exploit the dynamic interaction of these pulses to control gene expression. Here, using quantitative single-cell time-lapse imaging of Saccharomyces cerevisiae, we show that the pulsatile transcription factors Msn2 and Mig1 combinatorially regulate their target genes through modulation of their relative pulse timing. The activator Msn2 and repressor Mig1 showed pulsed activation in either a temporally overlapping or non-overlapping manner during their transient response to different inputs, with only the non-overlapping dynamics efficiently activating target gene expression. Similarly, under constant environmental conditions, where Msn2 and Mig1 exhibit sporadic pulsing, glucose concentration modulated the temporal overlap between pulses of the two factors. Together, these results reveal a time-based mode of combinatorial gene regulation. Regulation through relative signal timing is common in engineering and neurobiology, and these results suggest that it could also function broadly within the signalling and regulatory systems of the cell

Thickness-dependent Dielectric Constant of Few-layer In2Se3 Nano-flakes

The dielectric constant or relative permittivity of a dielectric material, which describes how the net electric field in the medium is reduced with respect to the external field, is a parameter of critical importance for charging and screening in electronic devices. Such a fundamental material property is intimately related to not only the polarizability of individual atoms, but also the specific atomic arrangement in the crystal lattice. In this letter, we present both experimental and theoretical investigations on the dielectric constant of few-layer In2Se3 nano-flakes grown on mica substrates by van der Waals epitaxy. A nondestructive microwave impedance microscope is employed to simultaneously quantify the number of layers and local electrical properties. The measured dielectric constant increases monotonically as a function of the thickness and saturates to the bulk value at around 6 ~ 8 quintuple layers. The same trend of layer-dependent dielectric constant is also revealed by first-principle calculations. Our results of the dielectric response, being ubiquitously applicable to layered 2D semiconductors, are expected to be significant for this vibrant research field.Comment: 15 pages, 4 figures, 1 table in Nano letters, 2015 ASA

Scaling laws governing stochastic growth and division of single bacterial cells

Uncovering the quantitative laws that govern the growth and division of single cells remains a major challenge. Using a unique combination of technologies that yields unprecedented statistical precision, we find that the sizes of individual Caulobacter crescentus cells increase exponentially in time. We also establish that they divide upon reaching a critical multiple ($\approx$1.8) of their initial sizes, rather than an absolute size. We show that when the temperature is varied, the growth and division timescales scale proportionally with each other over the physiological temperature range. Strikingly, the cell-size and division-time distributions can both be rescaled by their mean values such that the condition-specific distributions collapse to universal curves. We account for these observations with a minimal stochastic model that is based on an autocatalytic cycle. It predicts the scalings, as well as specific functional forms for the universal curves. Our experimental and theoretical analysis reveals a simple physical principle governing these complex biological processes: a single temperature-dependent scale of cellular time governs the stochastic dynamics of growth and division in balanced growth conditions.Comment: Text+Supplementar

Synthesis, Characterization, Structural and Phase Analysis of Novel Antiferroelectric Solid Solution

Complex structural transformations or phase transitions and interest for high-power energy storage density makes silver niobate (AgNbO3) an interesting material. In AgNbO3, various phase transitions are mainly associated with orthorhombic phases (in cubic structure, parameter a=b=c, where in orthorhombic, a≠b≠c). These transitions dominate the electrical behavior of AgNbO3 to a significant level. Here, by substituting lead and zirconium in place of silver and niobium, the solid solution system (Ag(1-x)Pb(x))Nb(1-x)Zr(x)O3 (where 0 ≤ x ≤ 0.12) was formed in form of ceramic, using conventional solid state synthesis. The Dielectric behavior (ability of a material to store energy as a function of temperature) and structural behavior suggested that with the increase in composition of lead zirconate PbZrO3, all the transition temperature in AgNbO3 moved towards the lower temperature. The frequency dependent dispersion in one of the orthorhombic phases, M1 phase, became more prominent till x ≤ 0.04 after that it was beyond the detection limit. Investigations of X-ray diffraction revealed the presence of different orthorhombic symmetries at room temperature with the varying substitution of PbZrO3. Analysis of X-ray diffraction peaks confirmed the phase transition from Pmc21 to the mixture of Pbcm and Pmc21 space group to Pbcm with the increase in composition of PbZrO3. To further study the interesting phases of AgNbO3, single crystals were successfully grown. An optical microscopy technique called Polarized Light Microscopy result confirmed the existence of room temperature orthorhombic symmetries. To summarize, the aim of experiment is to understand the effect of structural changes over electrical properties. Faculty Supervisor:Dr. Zuo-Guang Ye, Department of Chemistry, Simon Fraser Universit

Highway Discretionary Lane-change Decision and Control Using Model Predictive Control

To enable autonomous vehicles to perform discretionary lane change amidst the random traffic flow on highways, this paper introduces a decision-making and control method for vehicle lane change based on Model Predictive Control (MPC). This approach divides the driving control of vehicles on highways into two parts: lane-change decision and lane-change control, both of which are solved using the MPC method. In the lanechange decision module, the minimum driving costs for each lane are computed and compared by solving the MPC problem to make lane-change decisions. In the lane-change control module, a dynamic bicycle model is incorporated, and a multi-objective cost function is designed to obtain the optimal control inputs for the lane-change process. Additionally, A long-short term memory (LSTM) model is used to predict the trajectories of surrounding vehicles for both the MPC decision and control modules. The proposed lane-change decision and control method is simulated and validated in a driving simulator under random highway traffic conditions

Gesture counteracts gender stereotypes conveyed through subtle linguistic cues

Despite increased attempts to express equality in speech, biases often leak out through subtle linguistic cues. For example, the subject–complement statement (SCS, “Girls are as good as boys at math”) is used to advocate for equality but often reinforces gender stereotypes (boys are the standard against which girls are judged). We ask whether stereotypes conveyed by SCS can be counteracted by gesture. Two preregistered studies with 8- to 11-y-old children (N = 320 total) investigate whether an equal gesture—two palms placed at the same height—mitigates the gender stereotype induced by SCS. Children who saw the equal gesture along with SCS were more likely to express egalitarian beliefs than children who saw no gesture or an unequal gesture. Children can extract meaning from gesture when making stereotypical inferences, suggesting that the equal gesture may prove to be an innovative, and simple, intervention to counteract stereotypes introduced by subtle language

Single-gene tuning of Caulobacter cell cycle period and noise, swarming motility, and surface adhesion

We established that the sensor histidine kinase DivJ has an important role in the regulation of C. crescentus cell cycle period and noise. This was accomplished by designing and conducting single-cell experiments to probe the dependence of cell cycle noise on divJ expression and constructing a simplified cell cycle model that captures the dependence of cell cycle noise on DivJ with molecular details.In addition to its role in regulating the cell cycle, DivJ also affects polar cell development in C. crescentus, regulating swarming motility and surface adhesion. We propose that pleiotropic control of polar cell development by the DivJ–DivK–PleC signaling pathway underlies divJ-dependent tuning of cell swarming and adhesion behaviors.We have integrated the study of single-cell fluorescence dynamics with a kinetic model simulation to provide direct quantitative evidence that the DivJ histidine kinase is localized to the cell pole through a dynamic diffusion-and-capture mechanism during the C. crescentus cell cycle