149 research outputs found
Landino et al., Current Biology 2021
No full textMatlab codes used in the publication: J. Landino, M. Leda, ... A. Goryachev, and A. Miller, "Rho and F-actin self-organize within an artificial cell cortex" Current Biology, 2021 All Matlab .m files are in the single windows .zip file and are arranged into a directory tree according to the type of dynamics studied.Marcin Leda, & Andrew Goryachev. (2021). Landino et al., Current Biology 2021. Zenodo. https://doi.org/10.5281/zenodo.553275
Landino et al., Current Biology 2021
No full textMatlab codes used in the publication:
J. Landino, M. Leda, ... A. Goryachev, and A. Miller, "Rho and F-actin self-organize within an artificial cell cortex" Current Biology, 2021
All Matlab .m files are in the single windows .zip file and are arranged into a directory tree according to the type of dynamics studied
Curvature-driven positioning of Turing patterns in phase-separating curved membranes
No full textWe introduce a new finite difference scheme to study the dynamics of Turing patterns of a two-species activator-inhibitor system embedded on a phase-separating curved membrane, modelling for instance a lipid bilayer. We show that the underlying binary fluid can strongly affect both the dynamical and the steady state properties of the ensuing Turing patterns. Furthermore, geometry plays a key role, as a large enough local membrane curvature can both arrest the coarsening of the lipid domains and position the patterns selectively at areas of high or small local curvature. The physical phenomena we observe are due to a minimal coupling, between the diffusivity of the Turing components and the local membrane composition. While our study is theoretical in nature, it can provide a framework within which to address intracellular pattern formation in systems of interacting membrane proteins. © 2016 The Royal Society of Chemistry
Self-organisation in mixtures of microtubules and motor proteins
No full textSelf-organisation of mixtures of biological polymers and molecular motors
provides a fascinating manifestation of active matter. Microtubules re-oriented
by the molecular motors can form far-from-equilibrium cell-scale structures,
such as the mitotic spindle apparatus. It is believed that different motor types
favour formation of distinct patterns: clustering motors control the formation
of spindle poles and asters, while microtubule-sliding motors organise antiparallel
bundles presenting in the spindle central part.
The link between individual microscopic motor-induced interactions of filaments
and the macroscopic dynamics at cell-size scales is poorly understood.
Here we enhance our understanding of this problem and formulate a theoretical
approach, based on a Boltzmann-like kinetic equation, to describe pattern
formation in two-dimensional mixtures of microtubular filaments and molecular
motors.
In the first part of the thesis, we derive hydrodynamic equations that govern
the collective behaviour of microtubules in the presence of clustering motors.
We build on a kinetic method developed earlier by Aranson and Tsimring
and model the motor-induced reorientation of microtubules as collision rules.
The procedure of coarse-graining yields a set of equations for local density and
orientation of the microtubules. We study its behaviour by performing a linear
stability analysis and direct numerical simulations. We discuss the observed
patterns including asters and chaotic stripe-like structures and consider the
ensuing phase diagram.
In the second part of this study, we consider molecular motors which can
push apart antiparallel microtubules and cluster parallel ones. Using the developed
approach, we obtain a set of equations for the microtubular density,
orientation, and tensor of alignment. Through numerical simulations, we show
that this model generically creates either stable stripes with the antiparallel arrangement
of filaments inside them or an ever-evolving pattern where stripes
periodically form, rotate, self-extend and then split up. We derive a minimal
model which displays the same instability as the full model and clarifies the
underlying mechanism. We argue that our minimal model unifies various previous
observations of chaotic behaviour in the dry active matter into a general
universality class.
Finally, we discuss obtained models, compare them to identify common
features, and offer the directions of future advances
Computational study of electrostatic contribution to membrane dynamics
No full textElectrostatics plays a crucial role in the membrane biology. Negatively charged
lipids (such as PS, PA and PIP2) are subject to redistribution under the action
of electrostatic forces during various signalling events. Membrane recruitment
of multiple signalling proteins (such as MARCKS or Src kinase) is often maintained
by positively charged polybasic domains (PD). Even though adsorption of
these proteins to the cellular membrane has been extensively investigated, very
little is known about how electrostatic interactions contribute to their membrane
lateral dynamics. This thesis presents an investigation of the contribution of
electrostatic interactions to the membrane lateral dynamics by means of novel
computational tools. First, I developed a dynamic Monte-Carlo automaton that
faithfully simulates lateral diffusion of the adsorbed positively charged PD of a
peripheral membrane protein, as well as the dynamics of mono- (PS, PA) and
polyvalent (PIP2) anionic lipids within the bilayer. This model allowed to investigate
the major characteristics of protein-membrane diffusion on the uniform
membrane. In agreement with earlier results, the simulations revealed the following
microscopic phenomena: 1) Electrostatic lipid demixing in the vicinity of
the PD; 2) PD interacts with PIP2 stronger than with monovalent lipids. On the
spatially heterogeneous membrane the automaton predicted a directional drift
of the PD, which was validated by a simple mean-field analytical model. The
predicted phenomenon could potentially play a major role in membrane domain formation. To test this hypothesis and to investigate the membrane dynamics on
larger scales I developed a continuous model, which was based on the results of
the automaton simulations. The results of the continuous model and the Monte-Carlo simulations were shown to be in quantitative agreement. The continuous
model allows one to simulate the electrostatic membrane dynamics on micrometer
scales and can be used to describe various biologically important processes, such
as endocytic cup initiation
Symmetry Breaking in Cells and Tissues
No full text“Symmetry Breaking in Cells and Tissues” presents a collection of seventeen reviews, opinions and original research papers contributed by theoreticians, physicists and mathematicians, as well as experimental biologists, united by a common interest in biological pattern formation and morphogenesis. The contributors discuss diverse manifestations of symmetry breaking in biology and showcase recent developments in experimental and theoretical approaches to biological morphogenesis and pattern formation on multiple scales
Ca²⁺/Calmodulin signalling during colony initiation in Neurospora crassa
No full textThe primary research aims of this thesis were to analyse the mechanism of
Ca²⁺/calmodulin (CaM) signalling during conidial germination and conidial
anastomosis tube (CAT)-mediated fusion in Neurospora crassa. Ca²⁺ is an ubiquitous
signalling molecule that regulates many important processes in filamentous fungi
including spore germination, hyphal growth, mechanosensing, stress responses,
circadian rhythms, and the virulence of pathogens. Transient increases in cytosolic
free calcium ([Ca²⁺]c) act as intracellular signals. As the primary intracellular Ca²⁺
receptor, calmodulin (CaM) converts these Ca²⁺ signals into responses by regulating
the activities of numerous target proteins. Ca²⁺-free medium, antagonists of L-type
Ca²⁺ channels, CaM and calcineurin were found to inhibit CAT fusion. In addition, my
results showed that CAT chemotropism is dependent on extracellular Ca²⁺.
65 genes were identified as likely components of the Ca²⁺ signalling machinery
of N. crassa based on a comparative genomic analysis of S. cerevisiae, A. fumigatus
and C. albicans. Deletion mutants of 29 of these genes were characterized in relation
to their possible roles during colony initiation and development. Four of these
mutants (Δcna-1, Δcnb-1, Δcamk-1, Δplc-2, and Δrgs-1), which were homokaryons,
exhibited strong morphological phenotypes associated with CAT fusion.
To identify the protein machinery involved in Ca²⁺/CaM signalling during colony
initiation, proteins that directly or indirectly interacted with CaM were isolated from
germlings by immunoprecipitation and analyzed by mass spectroscopy. A total of 286
putative Ca²⁺/CaM-interacting proteins were identified in this way and 30 of these
proteins contained CaM-binding motifs. This proteomics analysis provided evidence
for Ca²⁺/CaM signalling playing a role in regulating the activity of a wide range of
proteins including MAP kinases in the cell integrity pathway, Ras/Rho signalling
pathway, and microtubule and actin cytoskeletal proteins.
GFP labelled CaM localized as dynamic spots associated with the plasma
membrane and cytoplasm in both germ tubes and CATs. Significant CaM
accumulation was observed in the tips of CATs growing towards each other, around
fusion pores at sites of CAT fusion, and at developing septa in germ tubes. CaM
localization was influenced by the actin and microtubule cytoskeleton during the
colony initiation. Inhibition of F-actin polymerization with latrunculin-A suppressed
the pronounced accumulation of CaM at growing germ tube and CAT tips. The
movement of CaM associated with spindle pole bodies was prevented by treatment
with the microtubule polymerization inhibitor benomyl. The absence of myo-5
resulted in reduced CAT fusion and the lack recruitment of CaM at growing tips
indicating a role for the motor protein, myosin-5, in these processes.
Finally, by expressing the genetically encoded Ca²⁺ sensor GCaMP6s under the
control of tef-1 promoter in N. crassa, I have been able to image [Ca²⁺]c dynamics in
this fungus for the first time. Using this I have been able to detect localized [Ca²⁺]c
spikes and waves in conidia, germ tubes and CATs. However, I obtained no clear
evidence for localized [Ca²⁺]c changes being associated with CAT chemotropism or
fusion
Spatio-temporal organisation and dynamics of centromere
No full textEukaryotes package their genomes into chromosomes, which must be replicated and segregated equally during each cell division. The centromere is the specialised locus for faithful chromosome segregation. The spatio-temporal organisation of proteins is a common strategy that biological systems adopt to increase their complexity given a certain amount of genetic information, which provides the degrees of freedom required to meet the diverse challenges they face. The same applies to the centromere, whose function of ensuring accurate chromosome segregation is partially, if not largely, realised by regulating the spatial organisation of critical proteins. Mechanistic understanding of many of these fascinating processes is still lacking. In this work, I studied the spatio-temporal organisation and dynamics of two key centromeric proteins, CENP-A and Shugoshin, attempting to uncover their respective underlying mechanisms.
CENP-A is the histone H3 variant whose presence epigenetically defines centromere identity. How CENP-A nucleosomes are established and maintained on chromatin is one of the key questions in centromere biology. As with other epigenetics marks, CENP-A nucleosomes are diluted because of DNA replication and have to be replenished in each cell cycle. A self-templating local deposition mechanism has emerged from experimental evidence but it fails to explain several characteristics of CENP-A spatial organisation. In this study, I used a theoretical approach to identify potentially underlying mechanism(s). I modified the classic theoretical model for epigenetics to describe the dynamics of CENP-A nucleosomes’ dilution and replenishment. It was found that local auto-amplification alone is insufficient to recapitulate the experimental observations, suggesting the existence of other mechanisms. Further analysis indicated that cooperativity and arbitrary number control are unlikely to be one of them. The combination of spontaneous conversion and threshold-based stabilisation achieved the best mimicry of the target characteristics. These results could provide directions for future experiments aiming to understand the establishment and maintenance of CENP-A nucleosomes.
Shugoshin is a peri-centromeric adaptor protein crucial for accurate chromosome segregation. The localisation of Shugoshin is responsive to tension, a mechanical force unique to bi-orientated sister chromatids, in various species. Despite the functional characterisation of this tension-dependent re-localisation, the underlying mechanisms are still elusive. I studied the question using experimental approaches in the model organism budding yeast Saccharomyces cerevisiae. There is only one Shugoshin variant named Sgo1 in budding yeast. The spindle assembly checkpoint (SAC) kinase Bub1 and its substrate H2A-S121 have been shown to be important for the peri-centromeric localisation of Sgo1. In this study, I found that, upon the establishment of tension, the kinetochore localisation of Bub1 is largely reduced and that H2A-S121 phosphorylation re-distributes from the centromere to chromosome arms, suggesting a sequential re-localisation of Bub1, H2A-S121 phosphorylation and Sgo1. Consistently, the removal of Bub1 promptly abolished H2A-S121 phosphorylation and Sgo1 peri-centromere localisation. This model indicates the existence of at least one phosphatase antagonising the activity of Bub1. Although PP1 is required for the re-localisation of Sgo1, the requirement was shown to be because of its role in regulating Bub1 localisation. The Sgo1 interactor PP2A-Rts1 was also found to be unimportant in this process, leaving the phosphatase(s) unidentified. Since Sgo1 has been proposed to promote chromosome condensation, our sequential re-distribution model predicts a more condensed chromosome status upon tension. Re-analysis of published Hi-C data confirmed the prediction, further supporting the model. Taken together, this work suggested a qualitative model for the tension-dependent re-localisation of Sgo1, provided insights into the mechanism of tension sensing and implied a potential link between SAC and chromosome condensation
A dynamic physical model of cell migration, differentiation and apoptosis in Caenorhabditis elegans
No full textThe germ line of the nematode C. elegansprovides a paradigm to study essential developmental concepts like stem cell differentiation and apoptosis. Here, we have created a computational model encompassing these developmental landmarks and the resulting movement of germ cells along the gonadal tube. We have used a technique based on molecular dynamics (MD) to model the physical movement of cells solely based on the force that arises from dividing cells. This novel way of using MD to drive the model enables calibration of simulation and experimental time. Based on this calibration, the analysis of our model shows that it is in accordance with experimental observations. In addition, the model provides insights into kinetics of molecular pathways within individual cells as well as into physical aspects like the cell density along the germ line and in local neighbourhoods of individual germ cells. In the future, the presented model can be used to test hypotheses about diverse aspects of development like stem cell division or programmed cell death. An iterative process of evolving this model and experimental testing in the model system C. eleganswill provide new insights into key developmental aspects
Global Parameter Identification of Stochastic Reaction Networks from Single Trajectories
No full textWe consider the problem of inferring the unknown parameters of a stochastic biochemical network model from a single measured time-course of the concentration of some of the involved species. Such measurements are available, e.g., from live-cell fluorescence microscopy in image-based systems biology. In addition, fluctuation time-courses from, e.g., fluorescence correlation spectroscopy provide additional information about the system dynamics that can be used to more robustly infer parameters than when considering only mean concentrations. Estimating model parameters from a single experimental trajectory enables single-cell measurements and quantification of cell–cell variability. We propose a novel combination of an adaptive Monte Carlo sampler, called Gaussian Adaptation, and efficient exact stochastic simulation algorithms that allows parameter identification from single stochastic trajectories. We benchmark the proposed method on a linear and a non-linear reaction network at steady state and during transient phases. In addition, we demonstrate that the present method also provides an ellipsoidal volume estimate of the viable part of parameter space and is able to estimate the physical volume of the compartment in which the observed reactions take place
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