Temporal Controls of the Asymmetric Cell Division Cycle in Caulobacter crescentus

The asymmetric cell division cycle of Caulobacter crescentus is orchestrated by an elaborate gene-protein regulatory network, centered on three major control proteins, DnaA, GcrA and CtrA. The regulatory network is cast into a quantitative computational model to investigate in a systematic fashion how these three proteins control the relevant genetic, biochemical and physiological properties of proliferating bacteria. Different controls for both swarmer and stalked cell cycles are represented in the mathematical scheme. The model is validated against observed phenotypes of wild-type cells and relevant mutants, and it predicts the phenotypes of novel mutants and of known mutants under novel experimental conditions. Because the cell cycle control proteins of Caulobacter are conserved across many species of alpha-proteobacteria, the model we are proposing here may be applicable to other genera of importance to agriculture and medicine (e.g., Rhizobium, Brucella)

In Silico Synchronization of Cellular Populations Through Expression Data Deconvolution

Cellular populations are typically heterogenous collections of cells at different points in their respective cell cycles, each with a cell cycle time that varies from individual to individual. As a result, true single-cell behavior, particularly that which is cell-cycle--dependent, is often obscured in population-level (averaged) measurements. We have developed a simple deconvolution method that can be used to remove the effects of asynchronous variability from population-level time-series data. In this paper, we summarize some recent progress in the development and application of our approach, and provide technical updates that result in increased biological fidelity. We also explore several preliminary validation results and discuss several ongoing applications that highlight the method's usefulness for estimating parameters in differential equation models of single-cell gene regulation.Comment: accepted for the 48th ACM/IEEE Design Automation Conferenc

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)

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

Modelling and calculation of dna damage and repair in mammalian cells induced by ionizing radiation of different quality

Recent experimental data have revealed a wealth of information that provides an exceptional opportunity to construct a mechanistic model of DNA repair. The cellular response to radiation exposure starts with repair of DNA damage and cell signalling that may lead to mutation, or cell death. The purpose of this work was to construct a mechanistic mathematical model of DNA repair in mammalian cells. The repair m odel is based on biochemical action of repair proteins to examin e the hypotheses regarding two or more components of double strand break ( DSB ) repair kinetics. The mechanistic mathematical model of repair proposed in this thesis is part of a bottom - up appr oach that assumes the cell is a complex system. In this approach radiation induces DNA damage, and the cellular response to radiation perturbation was modelled in terms of activating repair processes. A b iochemical kinetic method based on law of mass actio n was employed to model the repair pathways. The repair model consists of a set of nonlinear differential equations that calculates and explains protein act ivity on the damage step by step. The model takes into account complexity of the DSB, topology of da mage in the cell nucleus, and cell cycle . The solution of the model in terms of overall kinetics of DSB repair was compared with pulsed - field gel electrophoresis measurements. The repair model was integrated with the track structure model to calculate the damage spectrum and repair kinetics for every individual DSB induced by monoenergetic electrons, and ultrasoft X - rays. For this purpose we proposed a method to sample the protein repair actions for every individual DSB, and finally calculate the total repa ir time for that specific DSB. The DSB - repair kinetics for the number of DSB induced by 500 track s of monoenergetic electrons and ultrasoft X - rays were calculated and compared with experimental results for cell s irradiated with Al K , C K , and Ti K ultrasoft X - rays. The results presented here form the first example of mechanistic modelling and calculations for NHEJ, HR and MMEJ repair pathways. The results, for the first time , quantitatively confirm the hypothesis that the complex type double strand breaks play a major role in the slow kinetics of DSB repair. The results also confirm that simple DSB located in the heterocromatin delay the repair process due to a series of processes that are required for the relaxation of the heterochromatin. The rep air model established in this work provides a unique opportunity to continue this study of cellular responses to radiation furth er downstream that may have important implications for human risk estimation and radiotherapy

Vtělenost ve vztahu k novým technologiím

Tato práce je filozofickým průzkumem přivtělení technologických nástrojů, který přihlíží k empirickým výzkumům a vychází ze struktur zkušenosti objasněných raně fenomenologickou tradicí i z analýzy jednání čerpající z analytické tradice. Technologické nástroje jsou do našeho života tak hluboce včleněny, že fungují jako součást nás, přeměňují to, co si myslíme, že jsme schopní činit, a to, kdo jsme. Třebaže technologické nástroje otevírají nové prostory pro autonomní jednání, jejich vnitřní fungování může zůstat neviditelné, a vzniká riziko zakrnění našich schopností jednat. Vzhledem k tomu, že jsme zásadně zakotveni ve světě, nemůžeme se sami chápat bez odkazu na svět a svět nemůžeme pochopit bez ohledu na to, jací jsme. Jedinečnost související s lidským využíváním technologických nástrojů vyvstává z prvotní jedinečnosti, která umožňuje toto využití technologických nástrojů a odlišuje nás od našich nejbližších evolučních příbuzných. Některá zvířata rozšiřují své fyzické působení na okolní prostředí pomocí nástrojů. My lidé však používáme nástroje rovněž k rozšíření našich kognitivních schopností. A protože počítač je nejuniverzálnějším lidským nástrojem, který lze využít jak pro senzomotorické tak pro kognitivní účely, považujeme počítač za prototyp technologického nástroje. Náš popis struktur...This dissertation is an empirically responsive philosophical exploration into the incorporation of technological tools within a framework comprising the structures of experience laid out in the early phenomenological tradition and an analysis of agency drawing from the analytical tradition. Technological tools have become so deeply integrated in our lives that they function like a part of us, transforming what we feel we can do and even who we are. Although new spaces of autonomous agency have been opened up, since the inner workings of technological tools can remain invisible, we risk diminishing our own capacities. Since we are fundamentally embedded in the world, we cannot understand ourselves without reference to the world and we cannot understand the world without reference to the way we are. The uniqueness involved in our use of technological tools grows out of a more primordial uniqueness that makes technological tool use possible and sets us apart from our closest evolutionary relatives. Several animals extend their physical influence on the environment by means of tools. We humans, however, use tools to extend our cognitive abilities as well. And since the computer is the most universal human tool, which can be put to sensorimotor and cognitive purposes alike, we take the computer to be...Ústav filosofie a religionistikyInstitute of Philosophy and Religious StudiesFaculty of ArtsFilozofická fakult