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Molecules (Basel, Switzerland) Jun 2022The micrometer-scale assembly of various DNA nanostructures is one of the major challenges for further progress in DNA nanotechnology. Programmed patterns of 1D and 2D... (Review)
Review
The micrometer-scale assembly of various DNA nanostructures is one of the major challenges for further progress in DNA nanotechnology. Programmed patterns of 1D and 2D DNA origami assembly using specific DNA strands and micrometer-sized lattice assembly using cross-shaped DNA origami were performed on a lipid bilayer surface. During the diffusion of DNA origami on the membrane surface, the formation of lattices and their rearrangement in real-time were observed using high-speed atomic force microscopy (HS-AFM). The formed lattices were used to further assemble DNA origami tiles into their cavities. Various patterns of lattice-tile complexes were created by changing the interactions between the lattice and tiles. For the control of the nanostructure formation, the photo-controlled assembly and disassembly of DNA origami were performed reversibly, and dynamic assembly and disassembly were observed on a lipid bilayer surface using HS-AFM. Using a lipid bilayer for DNA origami assembly, it is possible to perform a hierarchical assembly of multiple DNA origami nanostructures, such as the integration of functional components into a frame architecture.
Topics: DNA; Lipid Bilayers; Microscopy, Atomic Force; Nanostructures; Nanotechnology; Nucleic Acid Conformation
PubMed: 35807467
DOI: 10.3390/molecules27134224 -
International Journal of Molecular... Feb 2023The integration of a DNA copy of an HIV-1 RNA genome into the host genome, carried out by the viral enzyme integrase, results in the formation of single-stranded gaps in...
The integration of a DNA copy of an HIV-1 RNA genome into the host genome, carried out by the viral enzyme integrase, results in the formation of single-stranded gaps in cellular DNA that must be repaired. Here, we have analyzed the involvement of the PI3K kinases, ATM, ATR, and DNA-PKcs, which are important players in the DNA damage response (DDR) in HIV-1 post-integrational DNA repair (PIR). The participation of the DNA-PK complex in HIV-1 PIR has been previously shown, and the formation of a complex between the viral integrase and the DNA-PK subunit, Ku70, has been found to be crucial for efficient PIR. Now, we have shown that the inhibition of both DNA-PKcs and ATM, but not ATR, significantly reduces PIR efficiency. The activation of both kinases is a sequential process, where one kinase, being activated, activates the other, and it occurs simultaneously with the integration of viral DNA. This fact suggests that the activation of both kinases triggers PIR. Most interestingly, the activation of not only DNA-PKcs, but also ATM depends on the complex formation between integrase and Ku70. The elucidation of the interactions between viruses and DDR is important both for understanding the modulation of host cell functions by these pathogens and for developing new approaches to combat viral infections.
Topics: HIV-1; Ataxia Telangiectasia Mutated Proteins; DNA-Activated Protein Kinase; DNA Repair; DNA Damage; DNA, Viral; Integrases
PubMed: 36769109
DOI: 10.3390/ijms24032797 -
Cancer Science Jan 2021Chemical carcinogenesis is focused on the formation of DNA adducts, a form of DNA damage caused by covalent binding of a chemical moiety to DNA. The detection of... (Review)
Review
Chemical carcinogenesis is focused on the formation of DNA adducts, a form of DNA damage caused by covalent binding of a chemical moiety to DNA. The detection of carcinogen-DNA adducts in human tissues, along with demonstration of mutagenicity/carcinogenicity in experimental systems, and validation of adducts as biomarkers of environmental exposure and indicators of cancer risk in molecular epidemiological studies suggests a pivotal role of DNA adducts in cancer development. However, accurate measurement of DNA adducts in varied biological samples is challenging. Advances in mass spectrometry have prompted the development of DNA adductome analysis, an emerging method that simultaneously screens for multiple DNA adducts and provides relevant structural information. In this review, we summarize the basic principle and applications of DNA adductome analysis that would contribute to the elucidation of the environmental causes of cancer. Based on parallel developments in several fields, including next-generation sequencing, we describe a new approach used to explore cancer etiology, which integrates analyses of DNA adductome data and mutational signatures derived from whole-genome/exome sequencing.
Topics: Animals; DNA; DNA Adducts; DNA Damage; Environmental Exposure; Humans; Mutation; Neoplasms
PubMed: 32978845
DOI: 10.1111/cas.14666 -
Nature Protocols Aug 2022Molecular simulation has become an integral part of the DNA/RNA nanotechnology research pipeline. In particular, understanding the dynamics of structures and... (Review)
Review
Molecular simulation has become an integral part of the DNA/RNA nanotechnology research pipeline. In particular, understanding the dynamics of structures and single-molecule events has improved the precision of nanoscaffolds and diagnostic tools. Here we present oxView, a design tool for visualization, design, editing and analysis of simulations of DNA, RNA and nucleic acid-protein nanostructures. oxView provides an accessible software platform for designing novel structures, tweaking existing designs, preparing them for simulation in the oxDNA/RNA molecular simulation engine and creating visualizations of simulation results. In several examples, we present procedures for using the tool, including its advanced features that couple the design capabilities with a coarse-grained simulation engine and scripting interface that can programmatically edit structures and facilitate design of complex structures from multiple substructures. These procedures provide a practical basis from which researchers, including experimentalists with limited computational experience, can integrate simulation and 3D visualization into their existing research programs.
Topics: DNA; Nanostructures; Nucleic Acid Conformation; Nucleic Acids; RNA; Software
PubMed: 35668321
DOI: 10.1038/s41596-022-00688-5 -
International Journal of Molecular... Oct 2023Hepatitis B virus (HBV) remains a dominant cause of hepatocellular carcinoma (HCC). Recently, it was shown that HBV and woodchuck hepatitis virus (WHV) integrate into... (Review)
Review
Hepatitis B virus (HBV) remains a dominant cause of hepatocellular carcinoma (HCC). Recently, it was shown that HBV and woodchuck hepatitis virus (WHV) integrate into the hepatocyte genome minutes after invasion. Retrotransposons and transposable sequences were frequent sites of the initial insertions, suggesting a mechanism for spontaneous HBV DNA dispersal throughout the hepatocyte genome. Several somatic genes were also identified as early insertional targets in infected hepatocytes and woodchuck livers. Head-to-tail joints (HTJs) dominated amongst fusions, indicating their creation by non-homologous end-joining (NHEJ). Their formation coincided with the robust oxidative damage of hepatocyte DNA. This was associated with the activation of poly(ADP-ribose) polymerase 1 (PARP1)-mediated dsDNA repair, as reflected by the augmented transcription of PARP1 and XRCC1; the PARP1 binding partner OGG1, a responder to oxidative DNA damage; and increased activity of NAD, a marker of PARP1 activation, and HO1, an indicator of cell oxidative stress. The engagement of the PARP1-mediated NHEJ repair pathway explains the HTJ format of the initial merges. The findings show that HBV and WHV are immediate inducers of oxidative DNA damage and hijack dsDNA repair to integrate into the hepatocyte genome, and through this mechanism, they may initiate pro-oncogenic processes. Tracking initial integrations may uncover early markers of HCC and help to explain HBV-associated oncogenesis.
Topics: Humans; Hepatitis B virus; Carcinoma, Hepatocellular; Liver Neoplasms; Hepatocytes; Cell Transformation, Neoplastic; Carcinogenesis; Genomics; DNA, Viral; Hepatitis B; X-ray Repair Cross Complementing Protein 1
PubMed: 37834296
DOI: 10.3390/ijms241914849 -
Electrophoresis May 2023A novel microfluidic DNA extraction protocol based on integrated diaphragm microvalves/pumps and silica-deposited open-channel columns was developed specifically for...
A novel microfluidic DNA extraction protocol based on integrated diaphragm microvalves/pumps and silica-deposited open-channel columns was developed specifically for automated and parallel DNA solid-phase extraction (SPE). The method uses microfluidic chips with a sandwiched structure containing three layers, which are the upper fluidic layer with surface-deposited silica on glass open channels as the extraction phase, the lower actuation layer with valve actuation channels on a glass wafer, and the middle poly(dimethylsiloxane) (PDMS) membrane for reversible bonding of the two glass substrates. These two glass substrates can be reused after thoroughly cleaning and the PDMS membrane can be replaced conveniently, which could effectively decrease the time and cost of chip manufacturing. The normally closed microvalves/pumps were used to automatically control all processes of the on-chip DNA SPE without cross-contamination and leakage, enabling the processing of multiple samples in parallel without changing the microvalve control module. Using the microchip device with integrated microvalves/pumps, automated, programmable, and simultaneous λ-DNA extractions from different samples could be attained, even from complex solutions such as human blood, and the silica-deposited open-channel columns could be reused stably and reliably. Results have demonstrated that most of the eluted λ-DNA was recovered in the second 2 µL of elution buffer with high-purity suitable for successful polymerase chain reaction amplification, making it possible for further integration into microfluidic devices for fully functional and high-throughput genetic analysis.
Topics: Humans; Microfluidics; Lab-On-A-Chip Devices; Polymerase Chain Reaction; DNA; Solid Phase Extraction; Microfluidic Analytical Techniques
PubMed: 36694428
DOI: 10.1002/elps.202200185 -
Nanotechnology Feb 2023Atom manufacturing has become a blooming frontier direction in the field of material and chemical science in recent years, focusing on the fabrication of functional... (Review)
Review
Atom manufacturing has become a blooming frontier direction in the field of material and chemical science in recent years, focusing on the fabrication of functional materials and devices with individual atoms or with atomic precision. Framework nucleic acids (FNAs) refer to nanoscale nucleic acid framework structures with novel properties distinct from those of conventional nucleic acids. Due to their ability to be precisely positioned and assembled at the nanometer or even atomic scale, FNAs are ideal materials for atom manufacturing. They hold great promise for the bottom-up construction of electronic devices by precisely arranging and integrating building blocks with atomic or near-atomic precision. In this review, we summarize the progress of atom manufacturing based on FNAs. We begin by introducing the atomic-precision construction of FNAs and the intrinsic electrical properties of DNA molecules. Then, we describe various approaches for the fabrication of FNAs templated materials and devices, which are classified as conducting, insulating, or semiconducting based on their electrical properties. We highlight the role of FNAs in the fabrication of functional electronic devices with atomic precision, as well as the challenges and opportunities for atom manufacturing with FNAs.
Topics: Nucleic Acids; DNA; Electronics
PubMed: 36669170
DOI: 10.1088/1361-6528/acb4f2 -
Journal of the American Chemical Society Mar 2022Chemistry is in a powerful position to synthetically replicate biomolecular structures. Adding functional complexity is key to increase the biomimetics' value for...
Chemistry is in a powerful position to synthetically replicate biomolecular structures. Adding functional complexity is key to increase the biomimetics' value for science and technology yet is difficult to achieve with poorly controlled building materials. Here, we use defined DNA blocks to rationally design a triggerable synthetic nanopore that integrates multiple functions of biological membrane proteins. Soluble triggers bind via molecular recognition to the nanopore components changing their structure and membrane position, which controls the assembly into a defined channel for efficient transmembrane cargo transport. Using ensemble, single-molecule, and simulation analysis, our activatable pore provides insight into the kinetics and structural dynamics of DNA assembly at the membrane interface. The triggered channel advances functional DNA nanotechnology and synthetic biology and will guide the design of controlled nanodevices for sensing, cell biological research, and drug delivery.
Topics: Biomimetics; DNA; Ion Channels; Nanopores; Nanotechnology
PubMed: 35253434
DOI: 10.1021/jacs.1c06598 -
Accounts of Chemical Research Aug 2023Excitons are the molecular-scale currency of electronic energy. Control over excitons enables energy to be directed and harnessed for light harvesting, electronics, and...
Excitons are the molecular-scale currency of electronic energy. Control over excitons enables energy to be directed and harnessed for light harvesting, electronics, and sensing. Excitonic circuits achieve such control by arranging electronically active molecules to prescribe desired spatiotemporal dynamics. Photosynthetic solar energy conversion is a canonical example of the power of excitonic circuits, where chromophores are positioned in a protein scaffold to perform efficient light capture, energy transport, and charge separation. Synthetic systems that aim to emulate this functionality include self-assembled aggregates, molecular crystals, and chromophore-modified proteins. While the potential of this approach is clear, these systems lack the structural precision to control excitons or even test the limits of their power. In recent years, DNA origami has emerged as a designer material that exploits biological building blocks to construct nanoscale architectures. The structural precision afforded by DNA origami has enabled the pursuit of naturally inspired organizational principles in a highly precise and scalable manner. In this Account, we describe recent developments in DNA-based platforms that spatially organize chromophores to construct tunable excitonic systems. The high fidelity of DNA base pairing enables the formation of programmable nanoscale architectures, and sequence-specific placement allows for the precise positioning of chromophores within the DNA structure. The integration of a wide range of chromophores across the visible spectrum introduces spectral tunability. These excitonic DNA-chromophore assemblies not only serve as model systems for light harvesting, solar conversion, and sensing but also lay the groundwork for the integration of coupled chromophores into larger-scale nucleic acid architectures.We have used this approach to generate DNA-chromophore assemblies of strongly coupled delocalized excited states through both sequence-specific self-assembly and the covalent attachment of chromophores. These strategies have been leveraged to independently control excitonic coupling and system-bath interaction, which together control energy transfer. We then extended this framework to identify how scaffold configurations can steer the formation of symmetry-breaking charge transfer states, paving the way toward the design of dual light-harvesting and charge separation DNA machinery. In an orthogonal application, we used the programmability of DNA chromophore assemblies to change the optical emission properties of strongly coupled dimers, generating a series of fluorophore-modified constructs with separable emission properties for fluorescence assays. Upcoming advances in the chemical modification of nucleotides, design of large-scale DNA origami, and predictive computational methods will aid in constructing excitonic assemblies for optical and computing applications. Collectively, the development of DNA-chromophore assemblies as a platform for excitonic circuitry offers a pathway to identifying and applying design principles for light harvesting and molecular electronics.
Topics: Photosynthesis; Energy Transfer; Fluorescent Dyes; DNA
PubMed: 37345736
DOI: 10.1021/acs.accounts.3c00086 -
Nature Aug 2022Biological processes depend on the differential expression of genes over time, but methods to make physical recordings of these processes are limited. Here we report a...
Biological processes depend on the differential expression of genes over time, but methods to make physical recordings of these processes are limited. Here we report a molecular system for making time-ordered recordings of transcriptional events into living genomes. We do this through engineered RNA barcodes, based on prokaryotic retrons, that are reverse transcribed into DNA and integrated into the genome using the CRISPR-Cas system. The unidirectional integration of barcodes by CRISPR integrases enables reconstruction of transcriptional event timing based on a physical record through simple, logical rules rather than relying on pretrained classifiers or post hoc inferential methods. For disambiguation in the field, we will refer to this system as a Retro-Cascorder.
Topics: CRISPR-Cas Systems; DNA; Gene Editing; Gene Expression; Genome; Information Storage and Retrieval; Integrases; Prokaryotic Cells; RNA; Reverse Transcription; Time Factors
PubMed: 35896746
DOI: 10.1038/s41586-022-04994-6