Bio-inspired artificial synapse with chemically mediated intercompartment communication for single molecule cell-synthetic cell interaction studies

Zoupa, Aspasia 2024. Bio-inspired artificial synapse with chemically mediated intercompartment communication for single molecule cell-synthetic cell interaction studies. PhD Thesis, Cardiff University. Item availability restricted.

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Abstract

Direct transient or stable cell-cell interactions drive numerous physiological processes critical to the maintenance and function of multicellular organisms. Yet, studying cell-cell associations is challenging given the teeming activity of biological membranes. Existing biomimetic models entail structural constraints incapable of encapsulating the dynamic nature of intermembrane interactions. Inspired by direct transient intermembrane communication, where information is relayed through ligand-receptor engagement, we constructed a double membrane system with a synapse-like architecture. This bio-inspired model is comprised of two artificial lipid membranes, a droplet interface bilayer (DIB), and an encapsulated giant unilamellar vesicle (GUV), interlinked by cholesterol-tethered hybridized DNA complementary strands. The complementary single DNA strands are end-cholesterol modified for membrane integration and internally labelled for single molecule tracking. The trans-synaptic DNA-mediated coupling is characterized by single molecule Förster Resonant Energy Transfer (smFRET) measurements and diffusion coefficient (D) estimation. Chemical signal communication across interconnected bilayers is instigated by concentration gradient-dependent content (ion and ATP) flow mediated via protein channels (aHL nanopores) in both DIB and GUV membranes. Elicited vesicular biochemical responses, including initiation of actin filament growth and Ca2+-dependent change in fluorescence intensity, are measured by TIRF microscopy. This solid artificial system is expanded to a biohybrid version by tethering individual living cells to the droplet membrane via DIB- anchored integrin-targeting ligands. Artificial membrane-plasma membrane coupling is characterized by morphodynamics analysis, including Kymography and ADAPT analysis. This dual membrane system with synaptic-like intermembrane space constitutes a novel platform for studying self-organisation of mobile, paired cleft components, inter-membrane signalling, and trans-synaptic adhesion dynamics with single molecule sensitivity. Both artificial and biohybrid systems are amenable to biomolecular constituent and multicomponent ligand integration, demonstrating this model’s potentiality to interrogate fundamental membrane processes and multifaceted cellular responses. This will increase understanding of basic membrane dynamics and guide tool development for synthetic biology, biotechnology applications and next-generation cell-based therapeutic approaches.

Item Type:	Thesis (PhD)
Date Type:	Completion
Status:	Unpublished
Schools:	Schools > Pharmacy
Subjects:	Q Science > Q Science (General)
Date of First Compliant Deposit:	25 February 2025
Last Modified:	25 Feb 2025 10:01
URI:	https://orca.cardiff.ac.uk/id/eprint/176458

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