title: PNIPAM-containing droplets: The route towards an artificial cytoskeleton assembly based on synthetic and natural components
creator: Sauter, Désirée
subject: 540
subject: 540 Chemistry and allied sciences
subject: 570
subject: 570 Life sciences
description: Although cells represent the smallest building blocks of life, they are already exhibiting a high level of complexity. It is therefore not surprising that researchers tend to utilize simplified systems for investigating, but also reconstituting, cellular processes in artificially constructed cells. The conception of those artificial cells can be done using bottom-up biological approaches. Currently, the reconstitution of cellular process is  achieved using natural or nature-derived components like the actomyosin-complex for the reconstitution of cellular motility. Although a reduction of complexity can be accomplished in this way, natural systems still require specialized conditions and sophisticated buffers.  Ultimately, the reduction of complexity can only be realized by the reconstitution of cellular processes using completely synthetic materials.  In this thesis, a completely artificial cytoskeleton based on thermo-responsive poly(N-isopropylacrylamide) (PNIPAM) to trigger motility of synthetic cells was established. To this end, a PNIPAM-based composite material containing gold nanorods was generated and its physiochemical behavior was characterized. The composite material was introduced and assembled inside water-in-oil emulsion droplets that have been stabilized by either a custom-made PNIPAM-containing surfactant (PNS) or a commercially available fluorosurfactant (CS). Besides the reversibility of the PNIPAM phase transition in bulk and droplet-based synthetic cells, the on-demand inducible deformation of droplets due to this phase transition was shown. Additionally, droplet  migration was triggered using asymmetric PNIPAM-containing droplets in combination with fluorophilic-coated surfaces. The production of those cell polarization-mimicking  droplets was achieved via targeted fusion of stable and unstable droplets during polymerization. Following the assessment of PNIPAM-mediated droplet motility, for the  first time, the artificial cytoskeleton material was combined with the natural cytoskeleton protein actin by a sequential pico-injection approach thus elucidating the biocompatibility of the presented system.  Herein, a versatile PNIPAM-based artificial cytoskeleton for synthetic cell applications was designed and implemented.  The control over the PNIPAM volume transition processes allowed to achieve the motility of the droplet-based synthetic cells by their dynamic deformation. The biocompatibility of the system by combining natural and synthetic cytoskeleton components could be shown.  The tolerance of the PNIPAM system also towards specialized buffer systems, might expand the bandwidth of the bottom-up synthetic biology tool kit, enhancing the assembly of hybrid cellular modules. Therefore, the developed approach represents an important milestone in the development of bottom-up synthetic biology applications.
date: 2021
type: Dissertation
type: info:eu-repo/semantics/doctoralThesis
type: NonPeerReviewed
format: application/pdf
identifier: https://archiv.ub.uni-heidelberg.de/volltextserverhttps://archiv.ub.uni-heidelberg.de/volltextserver/30507/7/DissertationDesireeSauter.pdf
identifier: DOI:10.11588/heidok.00030507
identifier: urn:nbn:de:bsz:16-heidok-305070
identifier:   Sauter, Désirée  (2021) PNIPAM-containing droplets: The route towards an artificial cytoskeleton assembly based on synthetic and natural components.  [Dissertation]     
relation: https://archiv.ub.uni-heidelberg.de/volltextserver/30507/
rights: info:eu-repo/semantics/openAccess
rights: http://archiv.ub.uni-heidelberg.de/volltextserver/help/license_urhg.html
language: eng