-
Accounts of Chemical Research Sep 2019Nanoparticles (NPs) have enormous potential to improve disease diagnosis and treatment due to their intrinsic electronic, optical, magnetic, mechanical, and... (Review)
Review
Nanoparticles (NPs) have enormous potential to improve disease diagnosis and treatment due to their intrinsic electronic, optical, magnetic, mechanical, and physiological properties. To realize their full potential for nanomedicine, NPs must be biocompatible and targetable toward specific biomolecules to ensure selective sensing, imaging, and drug delivery in complex environments such as living cells, tissues, animals, and human bodies. In this Account, we summarize our efforts to impart specific biocompatibility and biorecognition functionality to NPs by developing strategies to integrate inorganic and organic NPs with functional DNA (fDNA), including aptamers, DNAzymes, and aptazymes to create fDNA-NPs. These hybrid NPs take advantage of fDNA's ability to either bind targets or catalyze reactions in the presence of targets selectively and utilize their unique physicochemical properties including small size, low immunogenicity, and ease of synthesis and chemical modification in comparison with other molecules such as antibodies. By integrating inorganic NPs such as gold NPs, quantum dots, and iron oxide nanoparticles with fDNA, we designed stimuli-responsive fDNA-NPs that exhibit target induced assembly and disassembly of NPs, resulting in a variety of colorimetric, fluorescent, and magnetic resonance imaging (MRI)-based sensors for diagnostic of a broad range of analytes. To impart both biocompatibility and selectivity on inorganic NPs for targeted bioimaging, we have demonstrated DNA-mediated surface functionalization, shape-controlled synthesis, and coordinative assembly of such NPs as specific bioprobes. A highlight is provided on the construction of fDNA-based nanoprobes with light-activatable sensing and imaging functions, which provides precise control of recognition properties of fDNA with high spatiotemporal resolution. To explore the potential of organic NPs for biosensing applications, we have developed an enzyme-responsive fDNA-liposome as a universal sensing platform compatible with diverse biological targets as well as different detection methods including fluorescence, MRI, or temperature, making possible point-of-care diagnostics. To expand the application regime of organic NPs, we collaborated with the Zimmerman group to prepare single-chain block copolymer-based NPs and incorporated it with a variety of functions, including monovalent DNA for assembly, tunable surface chemistry for cellular imaging, and coordinative Cu(II) sites for catalyzing intracellular click reactions, demonstrating the potential of using organic NPs to create promising fDNA-NP systems with programmable functionalities. Furthermore, we survey our recent endeavor in integration of cell-specific aptamers with different NPs for targeted drug delivery, showing that introducing stimuli-responsive properties into NPs that target tumor microenvironments would enable safer and more effective therapy for cancers. Finally, current challenges and future perspectives in fDNA-mediated engineering of NPs for biomedical applications are discussed.
Topics: Animals; Biomedical Research; Biosensing Techniques; Colorimetry; DNA; Humans; Magnetic Resonance Imaging; Microscopy, Fluorescence; Nanoparticles
PubMed: 31411853
DOI: 10.1021/acs.accounts.9b00167 -
Sheng Wu Gong Cheng Xue Bao = Chinese... Mar 2023The modern bio-fermentation industry requires design and creation of efficient microbial cell factories for directed conversion of raw materials to target products. The... (Review)
Review
The modern bio-fermentation industry requires design and creation of efficient microbial cell factories for directed conversion of raw materials to target products. The main criteria for assessing the performance of microbial cell factories are their product synthesis capacity and stability. Due to the deficiencies of plasmids in gene expression such as instability and being easy to lose, integration of genes into chromosome is often a better choice for stable expression in microbial hosts. To this end, chromosomal gene integration technology has received much attention and has developed rapidly. In this review, we summarize the recent research progresses of chromosomal integration of large DNA fragments in microorganisms, illustrate the principles and features of various technologies, highlight the opportunity brought by the CRISPR-associated transposon systems, and prospect future research direction of this technology.
Topics: Chromosomes; Plasmids; DNA; Cloning, Molecular; Fermentation
PubMed: 36994558
DOI: 10.13345/j.cjb.220737 -
Proceedings. Biological Sciences Aug 2021Ancient DNA (aDNA) has played a major role in our understanding of the past. Important advances in the sequencing and analysis of aDNA from a range of organisms have... (Review)
Review
Ancient DNA (aDNA) has played a major role in our understanding of the past. Important advances in the sequencing and analysis of aDNA from a range of organisms have enabled a detailed understanding of processes such as past demography, introgression, domestication, adaptation and speciation. However, to date and with the notable exception of microbiomes and sediments, most aDNA studies have focused on single taxa or taxonomic groups, making the study of changes at the community level challenging. This is rather surprising because current sequencing and analytical approaches allow us to obtain and analyse aDNA from multiple source materials. When combined, these data can enable the simultaneous study of multiple taxa through space and time, and could thus provide a more comprehensive understanding of ecosystem-wide changes. It is therefore timely to develop an integrative approach to aDNA studies by combining data from multiple taxa and substrates. In this review, we discuss the various applications, associated challenges and future prospects of such an approach.
Topics: DNA, Ancient; Ecosystem; Fossils; Geologic Sediments
PubMed: 34428961
DOI: 10.1098/rspb.2021.1252 -
Current Biology : CB Aug 2021CRISPR-associated proteins (Cas1 and Cas2) integrate foreign DNA at the "leader" end of CRISPR loci. Several CRISPR leader sequences are reported to contain a binding...
CRISPR-associated proteins (Cas1 and Cas2) integrate foreign DNA at the "leader" end of CRISPR loci. Several CRISPR leader sequences are reported to contain a binding site for a DNA-bending protein called integration host factor (IHF). IHF-induced DNA bending kinks the leader of type I-E CRISPRs, recruiting an upstream sequence motif that helps dock Cas1-2 onto the first repeat of the CRISPR locus. To determine the prevalence of IHF-directed CRISPR adaptation, we analyzed 15,274 bacterial and archaeal CRISPR leaders. These experiments reveal multiple IHF binding sites and diverse upstream sequence motifs in a subset of the I-C, I-E, I-F, and II-C CRISPR leaders. We identify subtype-specific motifs and show that the phase of these motifs is critical for CRISPR adaptation. Collectively, this work clarifies the prevalence and mechanism(s) of IHF-dependent CRISPR adaptation and suggests that leader sequences and adaptation proteins may coevolve under the selective pressures of foreign genetic elements like plasmids or phages.
Topics: CRISPR-Associated Proteins; CRISPR-Cas Systems; Clustered Regularly Interspaced Short Palindromic Repeats; DNA; Endonucleases
PubMed: 34174210
DOI: 10.1016/j.cub.2021.05.068 -
Science Advances Jul 2023Recent advances in structural DNA nanotechnology have been facilitated by design tools that continue to push the limits of structural complexity while simplifying an...
Recent advances in structural DNA nanotechnology have been facilitated by design tools that continue to push the limits of structural complexity while simplifying an often-tedious design process. We recently introduced the software MagicDNA, which enables design of complex 3D DNA assemblies with many components; however, the design of structures with free-form features like vertices or curvature still required iterative design guided by simulation feedback and user intuition. Here, we present an updated design tool, MagicDNA 2.0, that automates the design of free-form 3D geometries, leveraging design models informed by coarse-grained molecular dynamics simulations. Our GUI-based, stepwise design approach integrates a high level of automation with versatile control over assembly and subcomponent design parameters. We experimentally validated this approach by fabricating a range of DNA origami assemblies with complex free-form geometries, including a 3D Nozzle, G-clef, and Hilbert and Trifolium curves, confirming excellent agreement between design input, simulation, and structure formation.
Topics: Nucleic Acid Conformation; Nanostructures; Nanotechnology; DNA; Computer-Aided Design; Molecular Dynamics Simulation
PubMed: 37494445
DOI: 10.1126/sciadv.adi0697 -
Nature Communications Oct 2022Dynamic regulation of nucleic acid hybridization is fundamental for switchable nanostructures and controllable functionalities of nucleic acids in both material...
Dynamic regulation of nucleic acid hybridization is fundamental for switchable nanostructures and controllable functionalities of nucleic acids in both material developments and biological regulations. In this work, we report a ligand-invasion pathway to regulate DNA hybridization based on host-guest interactions. We propose a concept of recognition handle as the ligand binding site to disrupt Watson-Crick base pairs and induce the direct dissociation of DNA duplex structures. Taking cucurbit[7]uril as the invading ligand and its guest molecules that are integrated into the nucleobase as recognition handles, we successfully achieve orthogonal and reversible manipulation of DNA duplex dissociation and recovery. Moreover, we further apply this approach of ligand-controlled nucleic acid hybridization for functional regulations of both the RNA-cleaving DNAzyme in test tubes and the antisense oligonucleotide in living cells. This ligand-invasion strategy establishes a general pathway toward dynamic control of nucleic acid structures and functionalities by supramolecular interactions.
Topics: DNA; DNA, Catalytic; Ligands; Nucleic Acid Hybridization; Oligonucleotides, Antisense; RNA
PubMed: 36209265
DOI: 10.1038/s41467-022-33738-3 -
Microbiology Spectrum Jun 2023A large number of transcriptome studies generate important data and information for the study of pathogenic mechanisms of pathogens, including Vibrio cholerae. V....
A large number of transcriptome studies generate important data and information for the study of pathogenic mechanisms of pathogens, including Vibrio cholerae. V. cholerae transcriptome data include RNA-seq and microarray: microarray data mainly include clinical human and environmental samples, and RNA-seq data mainly focus on laboratory processing conditions, including different stresses and experimental animals . In this study, we integrated the data sets of both platforms using Rank-in and the Limma R package normalized Between Arrays function, achieving the first cross-platform transcriptome data integration of V. cholerae. By integrating the entire transcriptome data, we obtained the profiles of the most active or silent genes. By transferring the integrated expression profiles into the weighted correlation network analysis (WGCNA) pipeline, we identified the important functional modules of V. cholerae stress treatment, gene manipulation, and culture as DNA transposon, chemotaxis and signaling, signal transduction, and secondary metabolic pathways, respectively. The analysis of functional module hub genes revealed the uniqueness of clinical human samples; however, under specific expression patterning, the Δ, Δ strains, and tobramycin treatment group showed high expression profile similarity with human samples. By constructing a protein-protein interaction (PPI) interaction network, we discovered several unreported novel protein interactions within transposon functional modules. We used two techniques to integrate RNA-seq data for laboratory studies with clinical microarray data for the first time. The interactions between V. cholerae genes were obtained from a global perspective, as well as comparing the similarity between clinical human samples and the current experimental conditions, and uncovering the functional modules that play a major role under different conditions. We believe that this data integration can provide us with some insight and basis for elucidating the pathogenesis and clinical control of V. cholerae.
Topics: Animals; Humans; Vibrio cholerae; Transcriptome; Gene Expression Profiling; RNA-Seq; DNA Transposable Elements
PubMed: 37191528
DOI: 10.1128/spectrum.05369-22 -
Nature Cell Biology Apr 2021R-loops are non-B DNA structures with intriguing dual consequences for gene expression and genome stability. In addition to their recognized roles in triggering DNA... (Review)
Review
R-loops are non-B DNA structures with intriguing dual consequences for gene expression and genome stability. In addition to their recognized roles in triggering DNA double-strand breaks (DSBs), R-loops have recently been demonstrated to accumulate in cis to DSBs, especially those induced in transcriptionally active loci. In this Review, we discuss whether R-loops actively participate in DSB repair or are detrimental by-products that must be removed to avoid genome instability.
Topics: DNA; DNA Breaks, Double-Stranded; DNA Repair; DNA Replication; Genomic Instability; Humans; R-Loop Structures; RNA; Saccharomyces cerevisiae
PubMed: 33837288
DOI: 10.1038/s41556-021-00663-4 -
ACS Nano Mar 2022Transient dissipative dimerization and transient gated dimerization of DNA tetrahedra nanostructures are introduced as functional modules to emulate transient and gated...
Transient dissipative dimerization and transient gated dimerization of DNA tetrahedra nanostructures are introduced as functional modules to emulate transient and gated protein-protein interactions and emergent protein-protein guided transient catalytic functions, operating in nature. Four tetrahedra are engineered to yield functional modules that, in the presence of pre-engineered auxiliary nucleic acids and the nicking enzyme Nt.BbvCI, lead to the fueled transient dimerization of two pairs of tetrahedra. The dynamic transient formation and depletion of DNA tetrahedra are followed by transient FRET signals generated by fluorophore-labeled tetrahedra. The integration of two inhibitors within the mixture of the four tetrahedra and two auxiliary modules, fueling the transient dimerization, results in selective inhibitor-guided gated transient dimerization of two different DNA tetrahedra dimers. Kinetic models for the dynamic transient dimerization and gated transient dimerization of the DNA tetrahedra are formulated and computationally simulated. The derived rate-constants allow the prediction and subsequent experimental validation of the performance of the systems under different auxiliary conditions. In addition, by appropriate modification of the four tetrahedra structures, the triggered gated emergence of selective transient catalytic functions driven by the two pairs of DNA tetrahedra dimers is demonstrated.
Topics: Catalysis; DNA; DNA, Catalytic; Dimerization; Nanostructures
PubMed: 35184545
DOI: 10.1021/acsnano.1c06117 -
Emerging Microbes & Infections Dec 2024The persistence of HBV covalently closed circular DNA (cccDNA) and HBV integration into the host genome in infected hepatocytes pose significant challenges to the cure...
The persistence of HBV covalently closed circular DNA (cccDNA) and HBV integration into the host genome in infected hepatocytes pose significant challenges to the cure of chronic HBV infection. Although CRISPR/Cas9-mediated genome editing shows promise for targeted clearance of viral genomes, a safe and efficient delivery method is currently lacking. Here, we developed a novel approach by combining light-induced heterodimerization and protein acylation to enhance the loading efficiency of Cas9 protein into extracellular vesicles (EVs). Moreover, vesicular stomatitis virus-glycoprotein (VSV-G) was incorporated onto the EVs membrane, significantly facilitating the endosomal escape of Cas9 protein and increasing its gene editing activity in recipient cells. Our results demonstrated that engineered EVs containing Cas9/gRNA and VSV-G can effectively reduce viral antigens and cccDNA levels in the HBV-replicating and infected cell models. Notably, we also confirmed the antiviral activity and high safety of the engineered EVs in the HBV-replicating mouse model generated by hydrodynamic injection and the HBV transgenic mouse model. In conclusion, engineered EVs could successfully mediate functional CRISPR/Cas9 delivery both and , leading to the clearance of episomal cccDNA and integrated viral DNA fragments, and providing a novel therapeutic approach for curing chronic HBV infection.
Topics: Animals; Mice; Hepatitis B virus; CRISPR-Associated Protein 9; DNA, Circular; CRISPR-Cas Systems; RNA, Guide, CRISPR-Cas Systems; DNA, Viral; Hepatitis B; Virus Replication
PubMed: 37982370
DOI: 10.1080/22221751.2023.2284286