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European Biophysics Journal : EBJ Nov 2023Nucleic acids are highly deformable helical molecules constantly stretched, twisted and bent in their biological functioning. Single molecule experiments have shown that...
Nucleic acids are highly deformable helical molecules constantly stretched, twisted and bent in their biological functioning. Single molecule experiments have shown that double stranded (ds)-RNA and standard ds-DNA have opposite twist-stretch patterns and stretching properties when overwound under a constant applied load. The key structural features of the A-form RNA and B-form DNA helices are here incorporated in a three-dimensional mesoscopic Hamiltonian model which accounts for the radial, bending and twisting fluctuations of the base pairs. Using path integral techniques which sum over the ensemble of the base pair fluctuations, I compute the average helical repeat of the molecules as a function of the load. The obtained twist-stretch relations and stretching properties, for short A- and B-helical fragments, are consistent with the opposite behaviors observed in kilo-base long molecules.
Topics: Nucleic Acids; Nucleic Acid Conformation; DNA; Base Pairing; RNA
PubMed: 37357224
DOI: 10.1007/s00249-023-01669-6 -
Theranostics 2020Liquid biopsy has emerged in the last ten years as an appealing noninvasive strategy to support early cancer diagnosis and follow-up interventions. However, conventional... (Review)
Review
Liquid biopsy has emerged in the last ten years as an appealing noninvasive strategy to support early cancer diagnosis and follow-up interventions. However, conventional liquid biopsy strategies involving specified biomarkers have encountered unexpected inconsistencies stemming from the use of different analytical methodologies. Recent reports have repeatedly demonstrated that integrated detection of multiple liquid biopsy biomarkers can significantly improve diagnostic performance by eliminating the influence of intratumoral heterogeneity. Herein, we review the progress in the field of liquid biopsy and propose a novel integrated liquid biopsy framework consisting of three categories: elementary, intermediate, and advanced integration. We also summarize the merits of the integration strategy and propose a roadmap toward refining cancer diagnosis, metastasis surveillance, and prognostication.
Topics: Biomarkers, Tumor; Circulating Tumor DNA; Early Detection of Cancer; Extracellular Vesicles; Humans; Liquid Biopsy; Neoplasms; Neoplastic Cells, Circulating
PubMed: 32089746
DOI: 10.7150/thno.40677 -
Journal of the American Chemical Society May 2022This manuscript introduces geometry as a means to program the tile-based DNA self-assembly in two and three dimensions. This strategy complements the sequence-focused...
This manuscript introduces geometry as a means to program the tile-based DNA self-assembly in two and three dimensions. This strategy complements the sequence-focused programmable assembly. DNA crystal assembly critically relies on intermotif, sticky-end cohesion, which requires complementarity not only in sequence but also in geometry. For DNA motifs to assemble into crystals, they must be associated with each other in the proper geometry and orientation to ensure that geometric hindrance does not prevent sticky ends from associating. For DNA motifs with exactly the same pair of sticky-end sequences, by adjusting the length (thus, helical twisting phase) of the motif branches, it is possible to program the assembly of these distinct motifs to either mix with one another, to self-sort and consequently separate from one another, or to be alternatingly arranged. We demonstrate the ability to program homogeneous crystals, DNA "alloy" crystals, and definable grain boundaries through self-assembly. We believe that the integration of this strategy and conventional sequence-focused assembly strategy could further expand the programming versatility of DNA self-assembly.
Topics: DNA; Nucleic Acid Conformation; Nucleotide Motifs
PubMed: 35507317
DOI: 10.1021/jacs.2c02456 -
Nucleic Acids Research May 2023Prokaryotic Argonautes (pAgos) are programmable nucleases involved in cell defense against invading DNA. In vitro, pAgos can bind small single-stranded guide DNAs to...
Prokaryotic Argonautes (pAgos) are programmable nucleases involved in cell defense against invading DNA. In vitro, pAgos can bind small single-stranded guide DNAs to recognize and cleave complementary DNA. In vivo, pAgos preferentially target plasmids, phages and multicopy genetic elements. Here, we show that CbAgo nuclease from Clostridium butyricum can be used for genomic DNA engineering in bacteria. We demonstrate that CbAgo loaded with plasmid-derived guide DNAs can recognize and cleave homologous chromosomal loci, and define the minimal length of homology required for this targeting. Cleavage of plasmid DNA at an engineered site of the I-SceI meganuclease increases guide DNA loading into CbAgo and enhances processing of homologous chromosomal loci. Analysis of guide DNA loading into CbAgo also reveals off-target sites of I-SceI in the Escherichia coli genome, demonstrating that pAgos can be used for highly sensitive detection of double-stranded breaks in genomic DNA. Finally, we show that CbAgo-dependent targeting of genomic loci with plasmid-derived guide DNAs promotes homologous recombination between plasmid and chromosomal DNA, depending on the catalytic activity of CbAgo. Specific targeting of plasmids with Argonautes can be used to integrate plasmid-encoded sequences into the chromosome thus enabling genome editing.
Topics: Gene Editing; Plasmids; DNA; Bacteria; DNA, Single-Stranded; Endonucleases
PubMed: 36987855
DOI: 10.1093/nar/gkad191 -
Trends in Genetics : TIG Aug 2021DNA is fundamentally important for all cellular organisms due to its role as a store of hereditary genetic information. The precise and accurate regulation of gene... (Review)
Review
DNA is fundamentally important for all cellular organisms due to its role as a store of hereditary genetic information. The precise and accurate regulation of gene transcription depends primarily on promoters, which vary significantly within and between genomes. Some promoters are rich in specific types of bases, while others have more varied, complex sequence characteristics. However, it is not only base sequence but also epigenetic modifications and altered DNA structure that regulate promoter activity. Significantly, many promoters across all organisms contain sequences that can form intrastrand hairpins (cruciforms) or four-stranded structures (G-quadruplex or i-motif). In this review we integrate recent studies on promoter regulation that highlight the importance of DNA structure in the evolutionary adaptation of promoter sequences.
Topics: DNA; Evolution, Molecular; G-Quadruplexes; Nucleic Acid Conformation; Promoter Regions, Genetic; Transcription, Genetic
PubMed: 33931265
DOI: 10.1016/j.tig.2021.04.003 -
Current Opinion in Structural Biology Aug 2022The structural unit of eukaryotic chromatin is a nucleosome, comprising two histone H2A/H2B heterodimers and one histone (H3/H4) tetramer, wrapped around by ∼146-bp... (Review)
Review
The structural unit of eukaryotic chromatin is a nucleosome, comprising two histone H2A/H2B heterodimers and one histone (H3/H4) tetramer, wrapped around by ∼146-bp core DNA and linker DNA. Flexible histone tails sticking out from the core undergo posttranslational modifications that are responsible for various epigenetic functions. Recently, the functional dynamics of histone tails and their modulation within the nucleosome and nucleosomal complexes have been investigated by integrating NMR, molecular dynamics simulations, and cryo-electron microscopy approaches. In particular, recent NMR studies have revealed correlations in the structures of histone N-terminal tails between H2A and H2B, as well as between H3 and H4 depending on linker DNA, suggesting that histone tail networks exist even within the nucleosome.
Topics: Chromatin; Cryoelectron Microscopy; DNA; Histones; Nucleosomes
PubMed: 35863166
DOI: 10.1016/j.sbi.2022.102436 -
Chemistry (Weinheim An Der Bergstrasse,... Apr 2023Eukaryotic transcription factors (TFs) are the final integrators of a complex molecular feedback mechanism that interfaces with the genome, consolidating information for... (Review)
Review
Eukaryotic transcription factors (TFs) are the final integrators of a complex molecular feedback mechanism that interfaces with the genome, consolidating information for transcriptional regulation. TFs consist of both structured DNA-binding domains and long intrinsically disordered regions (IDRs) embedded with motifs linked to transcriptional control. It is now well established that the dynamic multifunctionality of IDRs is the basis for a wide spectrum of TF functions necessary to navigate and regulate the human genome. This review dissects the chemical features of TF IDRs that endow them with structural plasticity that is central to their functions in the nucleus. Sequence analysis of a set of over 1600 human TFs through AlphaFold was used to identify key features of their IDRs. Recent studies were then highlighted to illustrate IDR involvement in processes such as protein interactions, DNA binding and specificity, chromatin opening, and phase separation. To expand our understanding of TF functions, future directions are suggested for integrating experiments and simulations, from in vitro to living systems.
Topics: Humans; Transcription Factors; Intrinsically Disordered Proteins; Eukaryota; DNA
PubMed: 36648282
DOI: 10.1002/chem.202203369 -
Experimental & Molecular Medicine Dec 2022PARPs play fundamental roles in multiple DNA damage recognition and repair pathways. Persistent nuclear PARP activation causes cellular NAD depletion and exacerbates...
PARPs play fundamental roles in multiple DNA damage recognition and repair pathways. Persistent nuclear PARP activation causes cellular NAD depletion and exacerbates cellular aging. However, very little is known about mitochondrial PARP (mtPARP) and poly ADP-ribosylation (PARylation). The existence of mtPARP is controversial, and the biological roles of mtPARP-induced mitochondrial PARylation are unclear. Here, we demonstrate the presence of PARP1 and PARylation in purified mitochondria. The addition of the PARP1 substrate NAD to isolated mitochondria induced PARylation, which was suppressed by treatment with the inhibitor olaparib. Mitochondrial PARylation was also evaluated by enzymatic labeling of terminal ADP-ribose (ELTA). To further confirm the presence of mtPARP1, we evaluated mitochondrial nucleoid PARylation by ADP ribose-chromatin affinity purification (ADPr-ChAP) and PARP1 chromatin immunoprecipitation (ChIP). We observed that NAD stimulated PARylation and TFAM occupancy on the mtDNA regulatory region D-loop, inducing mtDNA transcription. These findings suggest that PARP1 is integrally involved in mitochondrial PARylation and that NAD-dependent mtPARP1 activity contributes to mtDNA transcriptional regulation.
Topics: Poly ADP Ribosylation; NAD; Poly(ADP-ribose) Polymerase Inhibitors; Mitochondria; DNA, Mitochondrial
PubMed: 36473936
DOI: 10.1038/s12276-022-00894-x -
Viruses Jan 2021Human hepatitis B virus (HBV) can cause chronic, lifelong infection of the liver that may lead to persistent or episodic immune-mediated inflammation against... (Review)
Review
Human hepatitis B virus (HBV) can cause chronic, lifelong infection of the liver that may lead to persistent or episodic immune-mediated inflammation against virus-infected hepatocytes. This immune response results in elevated rates of killing of virus-infected hepatocytes, which may extend over many years or decades, lead to fibrosis and cirrhosis, and play a role in the high incidence of hepatocellular carcinoma (HCC) in HBV carriers. Immune-mediated inflammation appears to cause oxidative DNA damage to hepatocytes, which may also play a major role in hepatocarcinogenesis. An additional DNA damaging feature of chronic infections is random integration of HBV DNA into the chromosomal DNA of hepatocytes. While HBV DNA integration does not have a role in virus replication it may alter gene expression of the host cell. Indeed, most HCCs that arise in HBV carriers contain integrated HBV DNA and, in many, the integrant appears to have played a role in hepatocarcinogenesis. Clonal expansion of hepatocytes, which is a natural feature of liver biology, occurs because the hepatocyte population is self-renewing and therefore loses complexity due to random hepatocyte death and replacement by proliferation of surviving hepatocytes. This process may also represent a risk factor for the development of HCC. Interestingly, during chronic HBV infection, hepatocyte clones detected using integrated HBV DNA as lineage-specific markers, emerge that are larger than those expected to occur by random death and proliferation of hepatocytes. The emergence of these larger hepatocyte clones may reflect a survival advantage that could be explained by an ability to avoid the host immune response. While most of these larger hepatocyte clones are probably not preneoplastic, some may have already acquired preneoplastic changes. Thus, chronic inflammation in the HBV-infected liver may be responsible, at least in part, for both initiation of HCC via oxidative DNA damage and promotion of HCC via stimulation of hepatocyte proliferation through immune-mediated killing and compensatory division.
Topics: Animals; DNA, Viral; Hepatitis B virus; Hepatitis B, Chronic; Hepatocytes; Humans; Liver; Virus Integration
PubMed: 33573130
DOI: 10.3390/v13020210 -
Biochemical Society Transactions Aug 2020Synthetic gene circuits allow programming in DNA the expression of a phenotype at a given environmental condition. The recent integration of memory systems with gene... (Review)
Review
Synthetic gene circuits allow programming in DNA the expression of a phenotype at a given environmental condition. The recent integration of memory systems with gene circuits opens the door to their adaptation to new conditions and their re-programming. This lays the foundation to emulate neuromorphic behaviour and solve complex problems similarly to artificial neural networks. Cellular products such as DNA or proteins can be used to store memory in both digital and analog formats, allowing cells to be turned into living computing devices able to record information regarding their previous states. In particular, synthetic gene circuits with memory can be engineered into living systems to allow their adaptation through reinforcement learning. The development of gene circuits able to adapt through reinforcement learning moves Sciences towards the ambitious goal: the bottom-up creation of a fully fledged living artificial intelligence.
Topics: Artificial Intelligence; DNA; Gene Regulatory Networks; Genes, Synthetic
PubMed: 32756895
DOI: 10.1042/BST20200008