Bioactive cell-like hybrids from dendrimersomes with a human cell membrane and its components.

Cell-like hybrids from natural and synthetic amphiphiles provide a platform to engineer functions of synthetic cells and protocells. Cell membranes and vesicles prepared from human cell membranes are relatively unstable in vitro and therefore are difficult to study. The thicknesses of biological membranes and vesicles self-assembled from amphiphilic Janus dendrimers, known as dendrimersomes, are comparable. This feature facilitated the coassembly of functional cell-like hybrid vesicles from giant dendrimersomes and bacterial membrane vesicles generated from the very stable bacterial Escherichia coli cell after enzymatic degradation of its outer membrane. Human cells are fragile and require only mild centrifugation to be dismantled and subsequently reconstituted into vesicles. Here we report the coassembly of human membrane vesicles with dendrimersomes. The resulting giant hybrid vesicles containing human cell membranes, their components, and Janus dendrimers are stable for at least 1 y. To demonstrate the utility of cell-like hybrid vesicles, hybrids from dendrimersomes and bacterial membrane vesicles containing YadA, a bacterial adhesin protein, were prepared. The latter cell-like hybrids were recognized by human cells, allowing for adhesion and entry of the hybrid bacterial vesicles into human cells in vitro.

Design-functionality relationships for adhesion/growth-regulatory galectins.

Glycan-lectin recognition is assumed to elicit its broad range of (patho)physiological functions via a combination of specific contact formation with generation of complexes of distinct signal-triggering topology on biomembranes. Faced with the challenge to understand why evolution has led to three particular modes of modular architecture for adhesion/growth-regulatory galectins in vertebrates, here we introduce protein engineering to enable design switches. The impact of changes is measured in assays on cell growth and on bridging fully synthetic nanovesicles (glycodendrimersomes) with a chemically programmable surface. Using the example of homodimeric galectin-1 and monomeric galectin-3, the mutual design conversion caused qualitative differences, i.e., from bridging effector to antagonist/from antagonist to growth inhibitor and vice versa. In addition to attaining proof-of-principle evidence for the hypothesis that chimera-type galectin-3 design makes functional antagonism possible, we underscore the value of versatile surface programming with a derivative of the pan-galectin ligand lactose. Aggregation assays with N,N'-diacetyllactosamine establishing a parasite-like surface signature revealed marked selectivity among the family of galectins and bridging potency of homodimers. These findings provide fundamental insights into design-functionality relationships of galectins. Moreover, our strategy generates the tools to identify biofunctional lattice formation on biomembranes and galectin-reagents with therapeutic potential.

Direct Visualization of Vesicle Disassembly and Reassembly Using Photocleavable Dendrimers Elucidates Cargo Release Mechanisms.

Release of cargo molecules from cell-like nanocarriers can be achieved by chemical perturbations, including changes to pH and redox state and via optical modulation of membrane properties. However, little is known about the kinetics or products of vesicle breakdown due to limitations in real-time imaging at nanometer length scales. Using a library of 12 single-single type photocleavable amphiphilic Janus dendrimers, we developed a self-assembling light-responsive dendrimersome vesicle platform. A photocleavable ortho-nitrobenzyl inserted between the hydrophobic and hydrophilic dendrons of amphiphilic Janus dendrimers allowed for photocleavage and disassembly of their supramolecular assemblies. Distinct methods used to self-assemble amphiphilic Janus dendrimers produced either nanometer size small unilamellar vesicles or micron size giant multilamellar and onion-like dendrimersomes. In situ observation of giant photosensitive dendrimersomes via confocal microscopy elucidated rapid morphological transitions that accompany vesicle breakdown upon 405 nm laser illumination. Giant dendrimersomes displayed light-induced cleavage, disassembling and reassembling into much smaller vesicles at millisecond time scales. Additionally, photocleavable vesicles demonstrated rapid release of molecular and macromolecular cargos. These results guided our design of multilamellar particles to photorelease surface-attached proteins, photoinduce cargo recruitment, and photoconvert vesicle morphology. Real-time characterization of the breakdown and reassembly of lamellar structures provides insights on partial cargo retention and informs the design of versatile, optically regulated carriers for applications in nanoscience and synthetic biology.