Yan Qiao

Protocells provide a promising platform for constructing life‐like systems from the bottom up and understanding the physicochemical principles underlying cellular organization. Early protocell models primarily focused on minimal compartmentalization and simple biochemical functions, however, increasing evidence suggests that the emergence of complex cellular behaviours requires higher‐order structural organization across multiple length scales. Recent advances in synthetic cells, soft matter, and systems chemistry have therefore shifted the field toward engineering structurally complex protocells with enhanced functional and collective capabilities. In this Review, we summarize recent progress in the construction of protocells with increasing structural complexity, including membrane‐bounded condensate systems, multicompartmentalized architectures, and hierarchically organized assemblies. We further discuss how these structural features enable emergent functionalities such as spatially programmed reactions, signal transduction, adaptive communication, and collective organization. Particular emphasis is placed on the role of physicochemical interactions and matrix‐mediated assembly in directing protocell organization into higher‐order communities and tissue‐like materials. Finally, we highlight current challenges and future opportunities in developing dynamically adaptive, non‐equilibrium protocell systems that bridge the gap between simplified biomimetic compartments and autonomous synthetic living matter. This Review underscores structural complexity as a central design principle for driving protocells toward emergent functionality and life‐like behaviours.

Biomolecular condensates undergo dynamic maturation, transitioning from liquid-like droplets to gel-like or solid-like assemblies, exhibiting structural heterogeneity in response to biochemical cues. Synthetic coacervate droplets formed via liquid-liquid phase separation (LLPS) have emerged as simplified models of these condensates and of protocells. Yet, the understanding of complex phase behaviors of liquid-like droplets in both biological and chemical contexts remains less explored, limiting our understanding of biological function and the design of droplet-based soft materials and protocells. Here, we introduce a chemically programmable strategy to dynamically modulate droplet phase transitions through controlled polymer-network cross-linking. Reactive cross-linkers selectively engage polymers within liquid-like droplets, progressively increasing internal microviscosity and inducing a liquid-to-solid transition. Beyond uniform phase regulation, network cross-linking drives spatially heterogeneous phase separation via polymer demixing and Ostwald ripening, with reaction-diffusion dynamics critically shaping both thermodynamically stable and metastable droplets. Using orthogonal cleavable cross-linkers, we further demonstrate chemical control over droplet liquefaction and the generation of multiphasic structures with pathway-dependent configurations. Integration of photoresponsive cross-linkers with digital-micromirror device (DMD) technology enables precise spatial photopatterning of droplet networks. This work establishes a versatile framework for elucidating the structural principles of microphase separation within coacervates and provides a blueprint for designing dynamic soft materials and synthetic protocells.

Liquid crystalline droplets with molecularly crowded interiors capable of selective biomolecular sequestration and interfacial wetting have been recently developed for the construction of artificial cells (protocells) and chain-like protocell networks^1-3. The controlled division of these synthetic protocells^4-16, however, remains a challenge. Symmetric fission of vesicles and droplets has been shown^17-26 using thermal gradients, dissipative self-assembly, wetting energies and chemical reactions^27-31, but asymmetric division is rare. For example, osmotic pressure has been used to induce the asymmetric division of lipid vesicles containing a polyethylene glycol-dextran aqueous two-phase system³² as well as giant unilamellar vesicles prepared with lipid-phase-separated bilayers³³. Here we show that structured liquid droplets exhibit asymmetric division in the absence of reconstituted protein machinery. In the presence of alkaline phosphatase or multivalent metal cations, individual multilamellar droplets split to produce two morphologically distinct progeny (droplet and vesicle). We show that heteromorphic division occurs by circumferential growth of a single surface caveola along a latent core-shell domain boundary because of induced changes in lipid headgroup-nucleotide counterion interactions and demonstrate that functional biomolecules are transferred between the different protocell generations. Taken together, our results provide a step towards the bottom-up assembly of proliferating artificial cells.

Synthetic cells aim to emulate living systems by reconstituting essential cellular processes within lipid-bound architectures. However, their functional complexity remains constrained by a key challenge: the synthesis and correct integration of hydrophobic membrane proteins via cell-free approaches. Here, inspired by natural cells, we developed a spatially regulated translation strategy in which membrane-anchored mRNAs recruit ribosomes to drive the cotranslational insertion of membrane proteins into lipid bilayers. This design enables efficient in situ synthesis and integration of multiple transmembrane proteins within giant unilamellar vesicles, supporting selective small-molecule transport across membranes. Importantly, the method allows for precise stoichiometric control of membrane protein composition. Together, this work establishes a minimal yet versatile framework for the direct synthesis and integration of membrane proteins, advancing the construction of functional synthetic cells.

We summarize mechanisms of coacervates crossing phospholipid membranes including vesicular engulfment via electrostatic interaction, photoswitchable phospholipids, caveolin and actin polymerization, and direct penetration via lipid rafts and defects.

Pursuing small catalyst sizes has remained a constant endeavor to maximize reaction yields by exposing more active sites and/or enhancing charge extraction. Organic particle photocatalysts typically remain tens to hundreds of nanometers in size, constrained by the quasi-infinite conjugation of commonly used polymers. Here, we reveal the size-minimized organic heterojunction nanoparticles by using all small molecule photovoltaic materials, and significantly enhance their photocatalytic activities for solar-driven hydrogen evolution. Owing to the intrinsically weak intermolecular forces and the absence of molecular entanglement of small molecules, (sub)nanometer-scale diameters of polymer-free nanoparticles are yielded, representing a size reduction of 1-2 orders of magnitude than traditional polymer-containing nanoparticles. Optimized polymer-free nanoparticles attaching on covalent frameworks achieve photocatalytic mass-united hydrogen evolution rates of up to 527.2 ~ 3180.7 mmol h⁻¹ g⁻¹ at varied concentrations under simulated sunlight, and the external quantum efficiency is up to 32.8 % at near-infrared light, marking a competitive performance for organic photocatalysts.

Coacervates formed through liquid–liquid phase separation represent a fundamental model for protocell and serve as a membraneless organelle in living cells, modulating various biological processes. During aging or under stress, protein misfolding and oligomerization trigger aberrant liquid‐to‐solid phase transition (LSPT), a process driven by multivalent interactions. These phase transitions disrupt cellular equilibrium, leading to neurodegenerative diseases such as Alzheimer's disease, Parkinson's disease, and amyotrophic lateral sclerosis. The regulatory strategies is summarize to enhance molecular interactions in coacervate LSPT into three categories, including physical stimulation, molecular modulation, and sequence regulation. This review aims to establish a conceptual framework for modulating coacervate LSPT and further explores potential clinical treatments for neurodegenerative diseases.

In nature, life is inherently dissipative. Cells continuously consume energy (such as ATP) to sustain homeostasis, drive metabolism, and respond dynamically to environmental cues. Inspired by this principle, we develop a synthetic protocell system that exhibits dissipative behavior and initiates metabolic-like processes. Our design features synthetic vesicles formed from a cationic surfactant, which undergo a fuel-driven transformation into coacervate protocells via liquid-liquid phase separation. Dissipation is achieved through alkaline phosphatase (ALP)-catalyzed ATP hydrolysis, driving the reverse transition from coacervates back to vesicles. The distinct physicochemical properties and internal organization of vesicle and coacervate protocells enable us to design functional regulators capable of producing secondary signals, such as fluorescence and enzymatic products. This work offers a strategy for engineering enzymatic reaction-regulated dissipative behaviors of protocell systems that emulate key aspects of cellular metabolism, representing a step toward synthetic life-like systems with dynamic behavior and functional complexity.