Imperial College London

DrLorenzoDi Michele

Faculty of Natural SciencesDepartment of Chemistry

Honorary Senior Lecturer
 
 
 
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Contact

 

+44 (0)20 7594 3262l.di-michele Website

 
 
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Location

 

Molecular Sciences Research HubWhite City Campus

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Summary

 

Publications

Publication Type
Year
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64 results found

Peng Z, Iwabuchi S, Izumi K, Takiguchi S, Yamaji M, Fujita S, Suzuki H, Kambara F, Fukasawa G, Cooney A, Di Michele L, Elani Y, Matsuura T, Kawano Ret al., 2024, Lipid vesicle-based molecular robots, Lab on a Chip: miniaturisation for chemistry, physics, biology, materials science and bioengineering, Vol: 24, Pages: 996-1029, ISSN: 1473-0189

A molecular robot, which is a system comprised of one or more molecular machines and computers, can execute sophisticated tasks in many fields that span from nanomedicine to green nanotechnology. The core parts of molecular robots are fairly consistent from system to system and always include (i) a body to encapsulate molecular machines, (ii) sensors to capture signals, (iii) computers to make decisions, and (iv) actuators to perform tasks. This review aims to provide an overview of approaches and considerations to develop molecular robots. We first introduce the basic technologies required for constructing the core parts of molecular robots, describe the recent progress towards achieving higher functionality, and subsequently discuss the current challenges and outlook. We also highlight the applications of molecular robots in sensing biomarkers, signal communications with living cells, and conversion of energy. Although molecular robots are still in their infancy, they will unquestionably initiate massive change in biomedical and environmental technology in the not too distant future.

Journal article

Raguseo F, Wang Y, Li J, Petrić Howe M, Balendra R, Huyghebaert A, Vadukul DM, Tanase DA, Maher TE, Malouf L, Rubio-Sánchez R, Aprile FA, Elani Y, Patani R, Di Michele L, Di Antonio Met al., 2023, The ALS/FTD-related C9orf72 hexanucleotide repeat expansion forms RNA condensates through multimolecular G-quadruplexes, Nature Communications, Vol: 14, ISSN: 2041-1723

Amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) are neurodegenerative diseases that exist on a clinico-pathogenetic spectrum, designated ALS/FTD. The most common genetic cause of ALS/FTD is expansion of the intronic hexanucleotide repeat (GGGGCC)n in C9orf72. Here, we investigate the formation of nucleic acid secondary structures in these expansion repeats, and their role in generating condensates characteristic of ALS/FTD. We observe significant aggregation of the hexanucleotide sequence (GGGGCC)n, which we associate to the formation of multimolecular G-quadruplexes (mG4s) by using a range of biophysical techniques. Exposing the condensates to G4-unfolding conditions leads to prompt disassembly, highlighting the key role of mG4-formation in the condensation process. We further validate the biological relevance of our findings by detecting an increased prevalence of G4-structures in C9orf72 mutant human motor neurons when compared to healthy motor neurons by staining with a G4-selective fluorescent probe, revealing signal in putative condensates. Our findings strongly suggest that RNA G-rich repetitive sequences can form protein-free condensates sustained by multimolecular G-quadruplexes, highlighting their potential relevance as therapeutic targets for C9orf72 mutation-related ALS/FTD.

Journal article

Malouf L, Tanase DA, Fabrini G, Brady RA, Paez-Perez M, Leathers A, Booth MJ, Di Michele Let al., 2023, Sculpting DNA-based synthetic cells through phase separation and phase-targeted activity, Chem, Vol: 9, Pages: 3347-3364, ISSN: 2451-9308

Synthetic cells, like their biological counterparts, require internal compartments with distinct chemical and physical properties where different functionalities can be localized. Inspired by membrane-less compartmentalization in biological cells, here, we demonstrate how microphase separation can be used to engineer heterogeneous cell-like architectures with programmable morphology and compartment-targeted activity. The synthetic cells self-assemble from amphiphilic DNA nanostructures, producing core-shell condensates due to size-induced de-mixing. Lipid deposition and phase-selective etching are then used to generate a porous pseudo-membrane, a cytoplasm analog, and membrane-less organelles. The synthetic cells can sustain RNA synthesis via in vitro transcription, leading to cytoplasm and pseudo-membrane expansion caused by an accumulation of the transcript. Our approach exemplifies how architectural and functional complexity can emerge from a limited number of distinct building blocks, if molecular-scale programmability, emergent biophysical phenomena, and biochemical activity are coupled to mimic those observed in live cells.

Journal article

Walczak M, Mancini L, Xu J, Raguseo F, Kotar J, Cicuta P, Di Michele Let al., 2023, A Synthetic Signaling Network Imitating the Action of Immune Cells in Response to Bacterial Metabolism, ADVANCED MATERIALS, ISSN: 0935-9648

Journal article

Rubio-Sanchez R, Mognetti BM, Cicuta P, Di Michele Let al., 2023, DNA-Origami Line-Actants Control Domain Organization and Fission in Synthetic Membranes, JOURNAL OF THE AMERICAN CHEMICAL SOCIETY, Vol: 145, Pages: 11265-11275, ISSN: 0002-7863

Journal article

Walczak M, Brady RA, Leathers A, Kotar J, Di Michele Let al., 2023, Influence of hydrophobic moieties on the crystallization of amphiphilic DNA nanostructures, JOURNAL OF CHEMICAL PHYSICS, Vol: 158, ISSN: 0021-9606

Journal article

Morzy D, Tekin C, Caroprese V, Rubio-Sanchez R, Di Michele L, Bastings MMCet al., 2023, Interplay of the mechanical and structural properties of DNA nanostructures determines their electrostatic interactions with lipid membranes, NANOSCALE, Vol: 15, Pages: 2849-2859, ISSN: 2040-3364

Journal article

Walczak M, Mancini L, Xu J, Raguseo F, Kotar J, Cicuta P, Di Michele Let al., 2023, A synthetic signalling network imitating the action of immune cells in response to bacterial metabolism

<jats:p>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,<jats:italic>artificial cells</jats:italic>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 mimics 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.</jats:p>

Journal article

Rubio-Sánchez R, Mognetti BM, Cicuta P, Di Michele Let al., 2023, DNA-origami line-actants control domain organisation and fission in synthetic membranes

<jats:title>Abstract</jats:title><jats:p>Cells can precisely program the shape and lateral organisation of their membranes using protein machinery. Aiming to replicate a comparable degree of control, here we introduce DNA-Origami Line-Actants (DOLAs) as synthetic analogues of membrane-sculpting proteins. DOLAs are designed to selectively accumulate at the line-interface between co-existing domains in phase-separated lipid membranes, modulating the tendency of the domains to coalesce. With experiments and coarse-grained simulations, we demonstrate that DOLAs can reversibly stabilise two-dimensional analogues of Pickering emulsions on synthetic giant liposomes, enabling dynamic programming of membrane lateral organisation. The control afforded over membrane structure by DOLAs extends to three-dimensional morphology, as exemplified by a proof-of-concept synthetic pathway leading to vesicle fission. With DOLAs we lay the foundations for mimicking, in synthetic systems, some of the critical membrane-hosted functionalities of biological cells, including signalling, trafficking, sensing, and division.</jats:p>

Journal article

Garcia Hernandez NV, Buccelli S, Laffranchi M, De Michieli Let al., 2023, Mixed Reality-based Exergames for Upper Limb Robotic Rehabilitation, 18th Annual ACM/IEEE International Conference on Human-Robot Interaction (HRI), Publisher: ASSOC COMPUTING MACHINERY, Pages: 447-451

Conference paper

Fletcher M, Zhu J, Rubio-Sanchez R, Sandler SE, Al Nahas K, Di Michele L, Keyser UF, Tivony Ret al., 2022, DNA-Based Optical Quantification of Ion Transport across Giant Vesicles, ACS NANO, Vol: 16, Pages: 17128-17138, ISSN: 1936-0851

Journal article

Takamori S, Cicuta P, Takeuchi S, Di Michele Let al., 2022, DNA-assisted selective electrofusion (DASE) of Escherichia coli and giant lipid vesicles, Nanoscale, Vol: 14, Pages: 14255-14267, ISSN: 2040-3364

Synthetic biology and cellular engineering require chemical and physical alterations, which are typically achieved by fusing target cells with each other or with payload-carrying vectors. On one hand, electrofusion can efficiently induce the merging of biological cells and/or synthetic analogues via the application of intense DC pulses, but it lacks selectivity and often leads to uncontrolled fusion. On the other hand, synthetic DNA-based constructs, inspired by natural fusogenic proteins, have been shown to induce a selective fusion between membranes, albeit with low efficiency. Here we introduce DNA-assisted selective electrofusion (DASE) which relies on membrane-anchored DNA constructs to bring together the objects one seeks to merge, and applying an electric impulse to trigger their fusion. The DASE process combines the efficiency of standard electrofusion and the selectivity of fusogenic nanostructures, as we demonstrate by inducing and characterizing the fusion of spheroplasts derived from Escherichia coli bacteria with cargo-carrying giant lipid vesicles.

Journal article

Leathers A, Walczak M, Brady R, Al Samad A, Kotar J, Booth M, Cicuta P, Di Michele Let al., 2022, Reaction-diffusion patterning of DNA-based artificial cells, Journal of the American Chemical Society, Vol: 144, Pages: 17468-17476, ISSN: 0002-7863

Biological cells display complex internal architectures, with distinct micro environments that establish the chemical heterogeneity needed to sustain cellular functions. The continued efforts to create advanced cell mimics – artificial cells– demands strategies to construct similarly heterogeneous structures with localized functionalities. Here, we introduce a platform forconstructing membrane-less artificial cells from the self-assembly of synthetic DNA nanostructures, in which internal domains can be estab-lished thanks to prescribed reaction-diffusion waves. The method, rationalized through numerical modeling, enables the formation of up to five distinct, concentric environments, in which functional moieties can be localized. As a proof-of-concept, we apply this platform to build DNA-based artificial cells in which a prototypical nucleus synthesizes fluorescent RNAaptamers, which then accumulate in a surrounding storage shell, thus demonstrating spatial segregation of functionalities reminiscent of that observed in biological cells.

Journal article

Paez Perez M, Russell A, Cicuta P, Di Michele Let al., 2022, Modulating membrane fusion through the design of fusogenic DNA circuits and bilayer composition, Soft Matter, Vol: 18, Pages: 7035-7044, ISSN: 1744-683X

Membrane fusion is a ubiquitous phenomenon linked to many biological processes, and representsa crucial step in liposome-based drug delivery strategies. The ability to control, ever more precisely,membrane fusion pathways would thus be highly valuable for next generation nano-medical solutions and, more generally, the design of advanced biomimetic systems such as synthetic cells. Inthis article, we present fusogenic nanostructures constructed from synthetic DNA which, differentfrom previous solutions, unlock routes for modulating the rate of fusion and making it conditionalto the presence of soluble DNA molecules, thus demonstrating how membrane fusion can be controlled through simple DNA-based molecular circuits. We then systematically explore the relationshipbetween lipid-membrane composition, its biophysical properties, and measured fusion efficiency, linking our observations to the stability of transition states in the fusion pathway. Finally, we observethat specific lipid compositions lead to the emergence of complex bilayer architectures in the fusion products, such as nested morphologies, which are accompanied by alterations in biophysicalbehaviour. Our findings provide multiple, orthogonal strategies to program lipid-membrane fusion,which leverage the design of either the fusogenic DNA constructs or the physico/chemical propertiesof the membranes, and could thus be valuable in applications where some design parameters areconstrained by other factors such as material cost and biocompatibility, as it is often the case inbiotechnological applications.

Journal article

Kaufhold WT, Pfeifer W, Castro CE, Di Michele Let al., 2022, Probing the mechanical properties of DNA nanostructures with metadynamics, ACS Nano, Vol: 16, Pages: 8784-8797, ISSN: 1936-0851

Molecular dynamics simulations are often used to provide feedback in the design workflow of DNA nanostructures. However, even with coarse-grained models, the convergence of distributions from unbiased simulation is slow, limiting applications to equilibrium structural properties. Given the increasing interest in dynamic, reconfigurable, and deformable devices, methods that enable efficient quantification of large ranges of motion, conformational transitions, and mechanical deformation are critically needed. Metadynamics is an automated biasing technique that enables the rapid acquisition of molecular conformational distributions by flattening free energy landscapes. Here we leveraged this approach to sample the free energy landscapes of DNA nanostructures whose unbiased dynamics are nonergodic, including bistable Holliday junctions and part of a bistable DNA origami structure. Taking a DNA origami-compliant joint as a case study, we further demonstrate that metadynamics can predict the mechanical response of a full DNA origami device to an applied force, showing good agreement with experiments. Our results exemplify the efficient computation of free energy landscapes and force response in DNA nanodevices, which could be applied for rapid feedback in iterative design workflows and generally facilitate the integration of simulation and experiments. Metadynamics will be particularly useful to guide the design of dynamic devices for nanorobotics, biosensing, or nanomanufacturing applications.

Journal article

Fabrini G, Minard A, Brady R, Di Antonio M, Di Michele Let al., 2022, Cation-responsive and photocleavable hydrogels from non-canonical amphiphilic DNA nanostructures, Nano Letters: a journal dedicated to nanoscience and nanotechnology, Vol: 22, Pages: 602-611, ISSN: 1530-6984

Thanks to its biocompatibility, versatility and programmable interactions, DNA has been proposed as a building block for functional, stimuli-responsive frameworks with applications in biosensing, tissue engineering and drug delivery. Of particular importance for in vivo applications is the possibility of making such nano-materials responsive to physiological stimuli. Here we demonstrate how combining non-canonical DNA G-quadruplex (G4) structures with amphiphilic DNA constructs yields nanos-tructures, which we termed “Quad-Stars”, capable of assembling into responsive hydrogel particles via a straightforward, enzyme-free, one-pot reaction. The embedded G4 structures allow one to trigger and control the assembly/disassembly in a reversible fashion by adding or removing K+ ions. Furthermore, the hydrogel aggregates can be photo disassembled upon near-UV irradiation in the presence of a porphyrin photosensitiser. The combinedreversibility of assembly, responsiveness and cargo-loading capabilities of the hydrophobic moieties make Quad-Stars a promising candidate for biosensors and responsive drug delivery carriers.

Journal article

Rubio-Sanchez R, Fabrini G, Cicuta P, Di Michele Let al., 2021, Amphiphilic DNA nanostructures for bottom-up synthetic biology, Chemical Communications, Vol: 57, Pages: 12725-12740, ISSN: 1359-7345

DNA nanotechnology enables the construction of sophisticated biomimetic nanomachines that are increasingly central to the growing efforts of creating complex cell-like entities from the bottom-up. DNA nanostructures have been proposed as both structural and functional elements of these artificial cells, and in many instances are decorated with hydrophobic moieties to enable interfacing with synthetic lipid bilayers or regulating bulk self-organisation. In this feature article we review recent efforts to design biomimetic membrane-anchored DNA nanostructures capable of imparting complex functionalities to cell-like objects, such as regulated adhesion, tissue formation, communication and transport. We then discuss the ability of hydrophobic modifications to enable the self-assembly of DNA-based nanostructured frameworks with prescribed morphology and functionality, and explore the relevance of these novel materials for artificial cell science and beyond. Finally, we comment on the yet mostly unexpressed potential of amphiphilic DNA-nanotechnology as a complete toolbox for bottom-up synthetic biology – a figurative and literal scaffold upon which the next generation of synthetic cells could be built.

Journal article

Rubio-Sanchez R, O'Flaherty DK, Wang A, Coscia F, Petris G, Di Michele L, Cicuta P, Bonfio Cet al., 2021, Thermally driven membrane phase transitions enable content reshuffling in primitive cells, Journal of the American Chemical Society, Vol: 143, Pages: 16589-16598, ISSN: 0002-7863

Self-assembling single-chain amphiphiles available in the prebiotic environment likely played a fundamental role in the advent of primitive cell cycles. However, the instability of prebiotic fatty acid-based membranes to temperature and pH seems to suggest that primitive cells could only host prebiotically relevant processes in a narrow range of nonfluctuating environmental conditions. Here we propose that membrane phase transitions, driven by environmental fluctuations, enabled the generation of daughter protocells with reshuffled content. A reversible membrane-to-oil phase transition accounts for the dissolution of fatty acid-based vesicles at high temperatures and the concomitant release of protocellular content. At low temperatures, fatty acid bilayers reassemble and encapsulate reshuffled material in a new cohort of protocells. Notably, we find that our disassembly/reassembly cycle drives the emergence of functional RNA-containing primitive cells from parent nonfunctional compartments. Thus, by exploiting the intrinsic instability of prebiotic fatty acid vesicles, our results point at an environmentally driven tunable prebiotic process, which supports the release and reshuffling of oligonucleotides and membrane components, potentially leading to a new generation of protocells with superior traits. In the absence of protocellular transport machinery, the environmentally driven disassembly/assembly cycle proposed herein would have plausibly supported protocellular content reshuffling transmitted to primitive cell progeny, hinting at a potential mechanism important to initiate Darwinian evolution of early life forms.

Journal article

Walczak M, Brady RA, Mancini L, Contini C, Rubio-Sanchez R, Kaufhold W, Cicuta P, Di Michele Let al., 2021, Responsive core-shell DNA particles trigger lipid-membrane disruption and bacteria entrapment, Nature Communications, Vol: 12, Pages: 1-11, ISSN: 2041-1723

Biology has evolved a variety of agents capable of permeabilising and disrupting lipid membranes, from amyloid aggregates, to antimicrobial peptides, to venom compounds. While often associatedwith disease or toxicity, these agents are also central to many biosensing and therapeutic tech nologies. Here, we introduce a class of synthetic, DNA-based particles capable of disrupting lipid membranes. The particles have finely programmable size, and self-assemble from all-DNA and cholesterol-DNA nanostructures, the latter forming a membrane-adhesive core and the former a protective hydrophilic corona. We show that the corona can be selectively displaced with a molecu19 lar cue, exposing the ‘sticky’ core. Unprotected particles adhere to synthetic lipid vesicles, which in turn enhances membrane permeability and leads to vesicle collapse. Furthermore, particle-particle coalescence leads to the formation of gel-like DNA aggregates that envelop surviving vesicles. This response is reminiscent of pathogen immobilisation through immune cells secretion of DNA networks, as we demonstrate by trapping E. coli bacteria.

Journal article

Morzy D, Rubio-Sanchez R, Joshi H, Aksimentiev A, Di Michele L, Keyser UFet al., 2021, Cations regulate membrane attachment and functionality of DNA nanostructures, Journal of the American Chemical Society, Vol: 143, Pages: 7358-7367, ISSN: 0002-7863

The interplay between nucleic acids and lipids underpins several key processes in molecular biology, synthetic biotechnology, vaccine technology, and nanomedicine. These interactions are often electrostatic in nature, and much of their rich phenomenology remains unexplored in view of the chemical diversity of lipids, the heterogeneity of their phases, and the broad range of relevant solvent conditions. Here we unravel the electrostatic interactions between zwitterionic lipid membranes and DNA nanostructures in the presence of physiologically relevant cations, with the purpose of identifying new routes to program DNA–lipid complexation and membrane-active nanodevices. We demonstrate that this interplay is influenced by both the phase of the lipid membranes and the valency of the ions and observe divalent cation bridging between nucleic acids and gel-phase bilayers. Furthermore, even in the presence of hydrophobic modifications on the DNA, we find that cations are still required to enable DNA adhesion to liquid-phase membranes. We show that the latter mechanism can be exploited to control the degree of attachment of cholesterol-modified DNA nanostructures by modifying their overall hydrophobicity and charge. Besides their biological relevance, the interaction mechanisms we explored hold great practical potential in the design of biomimetic nanodevices, as we show by constructing an ion-regulated DNA-based synthetic enzyme.

Journal article

Rubio R, Eizagirre Barker S, Walczak M, Cicuta P, Di Michele Let al., 2021, A modular, dynamic, DNA-based platform for regulating cargo distribution and transport between lipid domains, Nano Letters, Vol: 21, Pages: 2800-2808, ISSN: 1530-6984

Cell membranes regulate the distribution of biological machinery between phase-separated lipid domains to facilitate key processes including signaling and transport, which are among the life-like functionalities that bottom-up synthetic biology aims to replicate in artificial-cellular systems. Here, we introduce a modular approach to program partitioning of amphiphilic DNA nanostructures in coexisting lipid domains. Exploiting the tendency of different hydrophobic “anchors” to enrich different phases, we modulate the lateral distribution of our devices by rationally combining hydrophobes and by changing nanostructure size and topology. We demonstrate the functionality of our strategy with a bioinspired DNA architecture, which dynamically undergoes ligand-induced reconfiguration to mediate cargo transport between domains via lateral redistribution. Our findings pave the way to next-generation biomimetic platforms for sensing, transduction, and communication in synthetic cellular systems.

Journal article

Rubio-Sánchez R, Barker SE, Walczak M, Cicuta P, Di Michele Let al., 2021, A modular, dynamic, DNA-based platform for regulating cargo distribution and transport between lipid domains

Cell membranes regulate the distribution of biological machinery between phase-separated lipid domains to facilitate key processes including signalling and transport, which are among the life-like functionalities that bottom-up synthetic biology aims to replicate in artificial-cellular systems. Here, we introduce a modular approach to program partitioning of amphiphilic DNA nanostructures in co-existing lipid domains. Exploiting the tendency of different hydrophobic “anchors” to enrich different phases, we modulate the lateral distribution of our devices by rationally combining hydrophobes, and by changing nanostructure size and its topology. We demonstrate the functionality of our strategy with a bio-inspired DNA architecture, which dynamically undergoes ligand-induced reconfiguration to mediate cargo transport between domains via lateral re-distribution. Our findings pave the way to next-generation biomimetic platforms for sensing, transduction, and communication in synthetic cellular systems.

Journal article

Clowsley AH, Kaufhold WT, Lutz T, Meletiou A, Di Michele L, Soeller Cet al., 2021, Repeat DNA-PAINT suppresses background and non-specific signals in optical nanoscopy, Nature Communications, Vol: 12, Pages: 1-10, ISSN: 2041-1723

DNA-PAINT is a versatile optical super-resolution technique relying on the transient binding of fluorescent DNA ‘imagers’ to target epitopes. Its performance in biological samples is often constrained by strong background signals and non-specific binding events, both exacerbated by high imager concentrations. Here we describe Repeat DNA-PAINT, a method that enables a substantial reduction in imager concentration, thus suppressing spurious signals. Additionally, Repeat DNA-PAINT reduces photoinduced target-site loss and can accelerate sampling, all without affecting spatial resolution.

Journal article

Lanfranco R, Jana PK, Bruylants G, Cicuta P, Mognetti BM, Di Michele Let al., 2020, Adaptable DNA interactions regulate surface triggered self assembly, Nanoscale, Vol: 12, Pages: 18616-18620, ISSN: 2040-3364

DNA-mediated multivalent interactions between colloidal particles have been extensively applied for their ability to program bulk phase behaviour and dynamic processes. Exploiting the competition between different types of DNA–DNA bonds, here we experimentally demonstrate the selective triggering of colloidal self-assembly in the presence of a functionalised surface, which induces changes in particle–particle interactions. Besides its relevance to the manufacturing of layered materials with controlled thickness, the intrinsic signal-amplification features of the proposed interaction scheme make it valuable for biosensing applications.

Journal article

Clowsley AH, Kaufhold WT, Lutz T, Meletiou A, Di Michele L, Christian Set al., 2020, Detecting nanoscale distribution of protein pairs by proximity dependent super-resolution microscopy, Journal of the American Chemical Society, Vol: 142, Pages: 12069-12078, ISSN: 0002-7863

Interactions between biomolecules such as proteins underlie most cellular processes. It is crucial to visualize these molecular-interaction complexes directly within the cell, to show precisely where these interactions occur and thus improve our understanding of cellular regulation. Currently available proximity-sensitive assays for in situ imaging of such interactions produce diffraction-limited signals and therefore preclude information on the nanometer-scale distribution of interaction complexes. By contrast, optical super-resolution imaging provides information about molecular distributions with nanometer resolution, which has greatly advanced our understanding of cell biology. However, current co-localization analysis of super-resolution fluorescence imaging is prone to false positive signals as the detection of protein proximity is directly dependent on the local optical resolution. Here we present proximity-dependent PAINT (PD-PAINT), a method for subdiffraction imaging of protein pairs, in which proximity detection is decoupled from optical resolution. Proximity is detected via the highly distance-dependent interaction of two DNA constructs anchored to the target species. Labeled protein pairs are then imaged with high-contrast and nanoscale resolution using the super-resolution approach of DNA-PAINT. The mechanisms underlying the new technique are analyzed by means of coarse-grained molecular simulations and experimentally demonstrated by imaging DNA-origami tiles and epitopes of cardiac proteins in isolated cardiomyocytes. We show that PD-PAINT can be straightforwardly integrated in a multiplexed super-resolution imaging protocol and benefits from advantages of DNA-based super-resolution localization microscopy, such as high specificity, high resolution, and the ability to image quantitatively.

Journal article

Mognetti BM, Cicuta P, Di Michele L, 2019, Programmable interactions with biomimetic DNA linkers at fluid membranes and interfaces., Reports on Progress in Physics, Vol: 82, Pages: 1-34, ISSN: 0034-4885

At the heart of the structured architecture and complex dynamics of biological&amp;#13; systems are specific and timely interactions operated by biomolecules. In many&amp;#13; instances, biomolecular agents are spatially confined to flexible lipid membranes where,&amp;#13; among other functions, they control cell adhesion, motility and tissue formation.&amp;#13; Besides being central to several biological processes, multivalent interactions mediated&amp;#13; by reactive linkers confined to deformable substrates underpin the design of synthetic-&amp;#13; biological platforms and advanced biomimetic materials. Here we review recent&amp;#13; advances on the experimental study and theoretical modelling of a heterogeneous&amp;#13; class of biomimetic systems in which synthetic linkers mediate multivalent interactions&amp;#13; between fluid and deformable colloidal units, including lipid vesicles and emulsion&amp;#13; droplets. Linkers are often prepared from synthetic DNA nanostructures, enabling&amp;#13; full programmability of the thermodynamic and kinetic properties of their mutual&amp;#13; interactions. The coupling of the statistical effects of multivalent interactions with&amp;#13; substrate fluidity and deformability gives rise to a rich emerging phenomenology that,&amp;#13; in the context of self-assembled soft materials, has been shown to produce exotic phase&amp;#13; behaviour, stimuli-responsiveness, and kinetic programmability of the self-assembly&amp;#13; process. Applications to (synthetic) biology will also be reviewed.

Journal article

del Barrio J, Liu J, Brady RA, Tan CSY, Chiodini S, Ricci M, Fernandez-Leiro R, Tsai C-J, Vasileiadi P, Di Michele L, Lairez D, Toprakcioglu C, Scherman OAet al., 2019, Emerging Two-Dimensional Crystallization of Cucurbit[8]uril Complexes: From Supramolecular Polymers to Nanofibers, JOURNAL OF THE AMERICAN CHEMICAL SOCIETY, Vol: 141, Pages: 14021-14025, ISSN: 0002-7863

Journal article

Kaufhold WT, Brady RA, Tuffnell JM, Cicuta P, Di Michele Let al., 2019, Membrane Scaffolds Enhance the Responsiveness and Stability of DNA-Based Sensing Circuits, BIOCONJUGATE CHEMISTRY, Vol: 30, Pages: 1850-1859, ISSN: 1043-1802

Journal article

Talbot EL, Kotar J, Di Michele L, Cicuta Pet al., 2019, Directed tubule growth from giant unilamellar vesicles in a thermal gradient, SOFT MATTER, Vol: 15, Pages: 1676-1683, ISSN: 1744-683X

Journal article

Lanfranco R, Jana PK, Tunesi L, Cicuta P, Mognetti BM, Di Michele L, Bruylants Get al., 2019, Kinetics of Nanoparticle-Membrane Adhesion Mediated by Multivalent Interactions, LANGMUIR, Vol: 35, Pages: 2002-2012, ISSN: 0743-7463

Journal article

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