A synthetic signalling network imitating the action of immune cells in response to bacterial metabolism

Abstract

State-of-the-art bottom-up synthetic biology allows us to replicate many basic biological functions in artificial cell-like devices. To mimic more complex behaviours, however, artificial cells would need to perform many of these functions in a synergistic and coordinated fashion, which remains elusive. Here we considered a sophisticated biological response, namely the capture and deactivation of pathogens by neutrophil immune cells, through the process of netosis. We designed a consortium consisting of two synthetic agents - responsive DNA-based particles and antibiotic-loaded lipid vesicles - whose coordinated action replicates the sought immune-like response when triggered by bacterial metabolism. The artificial netosis-like response emerges from a series of interlinked sensing and communication pathways between the live and synthetic agents, and translates into both physical and chemical antimicrobial actions, namely bacteria immobilisation and exposure to antibiotics. Our results demonstrate how advanced life-like responses can be prescribed with a relatively small number of synthetic molecular components, and outlines a new strategy for artificial-cell-based antimicrobial solutions.

Sculpting DNA-based synthetic cells through phase separation and phase-targeted activity

“…In a related system, Walczak et al reported the adhesion of small C-star aggregates onto the surface of cell-size Giant Unilamellar Vesicles (GUVs), 48,53 observing that C-star particles are able to permeabilise liposomes and cause their rupture. To assess whether similar destabilisation occurs for SUVs depositing onto cell-size C-star condensates, we encapsulated calcein inside SUVs lacking phospholipid fluorescent labelling.…”

“…18–20,43,44 Among condensate-forming DNA motifs, amphiphilic nanostructures, obtained by labelling DNA junctions with hydrophobic moieties, have shown significant potential owing to the stability of the self-assembled phases, their programmable nanostructure, and the possibility of engineering localised functionality and response to various environmental stimuli. 45–53…”

Co-transcriptional production of programmable RNA condensates and synthetic organelles

“…Among the many design features that can be straightforwardly controlled are nanostar valency, flexibility and arm-length, all known to predictably influence self-assembly in analogous DNA systems [21,23,28,41,67]. Control over condensatesize could be achieved by co-transcribing monovalent, surface passivating RNA constructs [68,69]. Further RNA aptamers might be embedded to recruit molecular guests, including enzymes and metabolites, while ribozymes [70] could confer catalytic properties to the synthetic MLOs.…”

Interrogation of RNA-protein interaction dynamics in bacterial growth

“…Moreover, it has been found to physically interact with previously characterised including csrB/C, arrS and chiX, all of which have a role in cell survival in acidic conditions (Aiso et al, 2014;Babitzke & Romeo, 2007;Hayes et al, 2006). As the cells were cultured in M9LB which contains glucose, we expect fermentation and acidification of the media through-out batch culture (Walczak et al, 2023), suggesting a putative upregulation of these ncRNAs. Positionally enriched k-mer analysis (PEKA) (Kuret et al, 2022) revealed a preference for AU-rich k-mers seen in the cases of ncRNAs (Fig.…”