Search for: All records

Abstract Synthetic cells offer a versatile platform for addressing biomedical and environmental challenges, due to their modular design and capability to mimic cellular processes such as biosensing, intercellular communication, and metabolism. Constructing synthetic cells capable of stimuli‐responsive secretion is vital for applications in targeted drug delivery and biosensor development. Previous attempts at engineering secretion for synthetic cells have been confined to non‐specific cargo release via membrane pores, limiting the spatiotemporal precision and specificity necessary for selective secretion. Here, a protein‐based platform termed TEV Protease‐mediated Releasable Actin‐binding Protein (TRAP) is designed and constructed for selective, rapid, and triggerable secretion in synthetic cells. TRAP is designed to bind tightly to reconstituted actin networks and is proteolytically released from bound actin, followed by secretion via cell‐penetrating peptide membrane translocation. TRAP's efficacy in facilitating light‐activated secretion of both fluorescent and luminescent proteins is demonstrated. By equipping synthetic cells with a controlled secretion mechanism, TRAP paves the way for the development of stimuli‐responsive biomaterials, versatile synthetic cell‐based biosensing systems, and therapeutic applications through the integration of synthetic cells with living cells for targeted delivery of protein therapeutics. 

Abstract Although diverse actin network architectures found inside the cell have been individually reconstituted outside of the cell, how different types of actin architectures reorganize under applied forces is not entirely understood. Recently, bottom‐up reconstitution has enabled studies where dynamic and phenotypic characteristics of various actin networks can be recreated in an isolated cell‐like environment. Here, by creating a giant unilamellar vesicle (GUV)‐based cell model encapsulating actin networks, we investigate how actin networks rearrange in response to localized stresses applied by micropipette aspiration. We reconstitute actin bundles and branched bundles in GUVs separately and mechanically perturb them. Interestingly, we find that, when aspirated, protrusive actin bundles that are otherwise randomly oriented in the GUV lumen collapse and align along the axis of the micropipette. However, when branched bundles are aspirated, the network remains intact and outside of the pipette while the GUV membrane is aspirated into the micropipette. These results reveal distinct responses in the rearrangement of actin networks in a network architecture‐dependent manner when subjected to physical forces.