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Nature Communications Aug 2023N-methyladenosine (mA) is an abundant, dynamic mRNA modification that regulates key steps of cellular mRNA metabolism. mA in the mRNA coding regions inhibits translation...
N-methyladenosine (mA) is an abundant, dynamic mRNA modification that regulates key steps of cellular mRNA metabolism. mA in the mRNA coding regions inhibits translation elongation. Here, we show how mA modulates decoding in the bacterial translation system using a combination of rapid kinetics, smFRET and single-particle cryo-EM. We show that, while the modification does not impair the initial binding of aminoacyl-tRNA to the ribosome, in the presence of mA fewer ribosomes complete the decoding process due to the lower stability of the complexes and enhanced tRNA drop-off. The mRNA codon adopts a π-stacked codon conformation that is remodeled upon aminoacyl-tRNA binding. mA does not exclude canonical codon-anticodon geometry, but favors alternative more dynamic conformations that are rejected by the ribosome. These results highlight how modifications outside the Watson-Crick edge can still interfere with codon-anticodon base pairing and complex recognition by the ribosome, thereby modulating the translational efficiency of modified mRNAs.
Topics: Anticodon; Codon; Models, Molecular; Nucleic Acid Conformation; Protein Biosynthesis; RNA, Messenger; RNA, Transfer
PubMed: 37553384
DOI: 10.1038/s41467-023-40422-7 -
Bioinformatics (Oxford, England) Nov 2023The two strands of the DNA double helix locally and spontaneously separate and recombine in living cells due to the inherent thermal DNA motion. This dynamics results in...
MOTIVATION
The two strands of the DNA double helix locally and spontaneously separate and recombine in living cells due to the inherent thermal DNA motion. This dynamics results in transient openings in the double helix and is referred to as "DNA breathing" or "DNA bubbles." The propensity to form local transient openings is important in a wide range of biological processes, such as transcription, replication, and transcription factors binding. However, the modeling and computer simulation of these phenomena, have remained a challenge due to the complex interplay of numerous factors, such as, temperature, salt content, DNA sequence, hydrogen bonding, base stacking, and others.
RESULTS
We present pyDNA-EPBD, a parallel software implementation of the Extended Peyrard-Bishop-Dauxois (EPBD) nonlinear DNA model that allows us to describe some features of DNA dynamics in detail. The pyDNA-EPBD generates genomic scale profiles of average base-pair openings, base flipping probability, DNA bubble probability, and calculations of the characteristically dynamic length indicating the number of base pairs statistically significantly affected by a single point mutation using the Markov Chain Monte Carlo algorithm.
AVAILABILITY AND IMPLEMENTATION
pyDNA-EPBD is supported across most operating systems and is freely available at https://github.com/lanl/pyDNA_EPBD. Extensive documentation can be found at https://lanl.github.io/pyDNA_EPBD/.
Topics: Computer Simulation; Models, Chemical; DNA; Software; Base Pairing; Nucleic Acid Conformation
PubMed: 37991847
DOI: 10.1093/bioinformatics/btad699 -
IScience Jun 2024We show that the non-canonical nucleobase 2,6-diaminopurine (D) spontaneously base pairs with uracil (U) in water and the solid state without the need to be attached...
We show that the non-canonical nucleobase 2,6-diaminopurine (D) spontaneously base pairs with uracil (U) in water and the solid state without the need to be attached to the ribose-phosphate backbone. Depending on the reaction conditions, D and U assemble in thermodynamically stable hydrated and anhydrated D-U base-paired cocrystals. Under UV irradiation, an aqueous solution of D-U base-pair undergoes photochemical degradation, while a pure aqueous solution of U does not. Our simulations suggest that D may trigger the U photodimerization and show that complementary base-pairing modifies the photochemical properties of nucleobases, which might have implications for prebiotic chemistry.
PubMed: 38783999
DOI: 10.1016/j.isci.2024.109894 -
Biochimie Nov 2023Long non-coding RNAs (lncRNAs) are recently-discovered transcripts involved in gene expression regulation and associated with diseases. Despite the unprecedented... (Review)
Review
Long non-coding RNAs (lncRNAs) are recently-discovered transcripts involved in gene expression regulation and associated with diseases. Despite the unprecedented molecular complexity of these transcripts, recent studies of the secondary and tertiary structure of lncRNAs are starting to reveal the principles of lncRNA structural organization, with important functional implications. It therefore starts to be possible to analyze lncRNA structures systematically. Here, using a set of prototypical and medically-relevant lncRNAs of known secondary structure, we specifically catalogue the distribution and structural environment of one of the first-identified and most frequently occurring non-canonical Watson-Crick interactions, the G·U base pair. We compare the properties of G·U base pairs in our set of lncRNAs to those of the G·U base pairs in other well-characterized transcripts, like rRNAs, tRNAs, ribozymes, and riboswitches. Furthermore, we discuss how G·U base pairs in these targets participate in establishing interactions with proteins or miRNAs, and how they enable lncRNA tertiary folding by forming intramolecular or metal-ion interactions. Finally, by identifying highly-G·U-enriched regions of yet unknown function in our target lncRNAs, we provide a new rationale for future experimental investigation of these motifs, which will help obtain a more comprehensive understanding of lncRNA functions and molecular mechanisms in the future.
Topics: Base Pairing; RNA, Long Noncoding; Nucleic Acid Conformation; RNA, Ribosomal; RNA, Transfer
PubMed: 37353139
DOI: 10.1016/j.biochi.2023.06.003 -
ACS Omega Jul 2023Hydrogen bonding between nucleobases is a crucial noncovalent interaction for life on Earth. Canonical nucleobases form base pairs according to two main geometries:...
Hydrogen bonding between nucleobases is a crucial noncovalent interaction for life on Earth. Canonical nucleobases form base pairs according to two main geometries: Watson-Crick pairing, which enables the static functions of nucleic acids, such as the storing of genetic information; and Hoogsteen pairing, which facilitates the dynamic functions of these biomacromolecules. This precisely tuned system can be affected by oxidation or substitution of nucleobases, leading to changes in their hydrogen-bonding patterns. This paper presents an investigation into the intermolecular interactions of various 8-substituted purine derivatives with their hydrogen-bonding partners. The systems were analyzed using nuclear magnetic resonance spectroscopy and density functional theory calculations. Our results demonstrate that the stability of hydrogen-bonded complexes, or base pairs, depends primarily on the number of intermolecular H-bonds and their donor-acceptor alternation. No strong preferences for a particular geometry, either Watson-Crick or Hoogsteen, were found.
PubMed: 37483191
DOI: 10.1021/acsomega.3c03244 -
Proceedings of the National Academy of... Oct 2023The emergence of functional oligonucleotides on early Earth required a molecular selection mechanism to screen for specific sequences with prebiotic functions. Cyclic...
The emergence of functional oligonucleotides on early Earth required a molecular selection mechanism to screen for specific sequences with prebiotic functions. Cyclic processes such as daily temperature oscillations were ubiquitous in this environment and could trigger oligonucleotide phase separation. Here, we propose sequence selection based on phase separation cycles realized through sedimentation in a system subjected to the feeding of oligonucleotides. Using theory and experiments with DNA, we show sequence-specific enrichment in the sedimented dense phase, in particular of short 22-mer DNA sequences. The underlying mechanism selects for complementarity, as it enriches sequences that tightly interact in the dense phase through base-pairing. Our mechanism also enables initially weakly biased pools to enhance their sequence bias or to replace the previously most abundant sequences as the cycles progress. Our findings provide an example of a selection mechanism that may have eased screening for auto-catalytic self-replicating oligonucleotides.
Topics: Oligonucleotides; DNA; Temperature; Base Pairing
PubMed: 37847736
DOI: 10.1073/pnas.2218876120 -
Nucleic Acids Research Nov 2023There are >170 naturally occurring RNA chemical modifications, with both known and unknown biological functions. Analytical methods for detecting chemical modifications...
There are >170 naturally occurring RNA chemical modifications, with both known and unknown biological functions. Analytical methods for detecting chemical modifications and for analyzing their effects are relatively limited and have had difficulty keeping pace with the demand for RNA chemical biology and biochemistry research. Some modifications can affect the ability of RNA to hybridize with its complementary sequence or change the selectivity of base pairing. Here, we investigate the use of affinity-based DNA nanoswitches to resolve energetic differences in hybridization. We found that a single m3C modification can sufficiently destabilize hybridization to abolish a detection signal, while an s4U modification can selectively hybridize with G over A. These results establish proof of concept for using DNA nanoswitches to detect certain RNA modifications and analyzing their effects in base pairing stability and specificity.
Topics: Base Pairing; Base Sequence; DNA; Nucleic Acid Hybridization; RNA
PubMed: 37811879
DOI: 10.1093/nar/gkad802 -
Heliyon Sep 2023The vast majority of human transcriptome is represented by various types of small RNAs with little or no protein-coding capability referred to as non-coding RNAs... (Review)
Review
The vast majority of human transcriptome is represented by various types of small RNAs with little or no protein-coding capability referred to as non-coding RNAs (ncRNAs). Functional ncRNAs include microRNAs (miRNAs), long non-coding RNAs (lncRNAs), and circular RNAs (circRNAs), which are expressed at very low, but stable and reproducible levels in a variety of cell types. ncRNAs regulate gene expression due to miRNA capability of complementary base pairing with mRNAs, whereas lncRNAs and circRNAs can sponge miRNAs off their target mRNAs to act as competitive endogenous RNAs (ceRNAs). Each miRNA can target multiple mRNAs and a single mRNA can interact with several miRNAs, thereby creating miRNA-mRNA, lncRNA-miRNA-mRNA, and circRNA-miRNA-mRNA regulatory networks. Over the past few years, a variety of differentially expressed miRNAs, lncRNAs, and circRNAs (DEMs, DELs, and DECs, respectively) have been linked to cancer pathogenesis. They can exert both oncogenic and tumor suppressor roles. In this review, we discuss the recent advancements in uncovering the roles of DEMs, DELs, and DECs and their networks in aberrant cell signaling, cell cycle, transcription, angiogenesis, and apoptosis, as well as tumor microenvironment remodeling and metabolic reprogramming during hepatocarcinogenesis. We highlight the potential and challenges in the use of differentially expressed ncRNAs as biomarkers for liver cancer diagnosis and prognosis.
PubMed: 37662778
DOI: 10.1016/j.heliyon.2023.e19223 -
Cell Research Sep 2023Nicotinamide adenine dinucleotide (NAD) is a central metabolite in cellular processes. Depletion of NAD has been demonstrated to be a prevalent theme in both prokaryotic...
Nicotinamide adenine dinucleotide (NAD) is a central metabolite in cellular processes. Depletion of NAD has been demonstrated to be a prevalent theme in both prokaryotic and eukaryotic immune responses. Short prokaryotic Argonaute proteins (Agos) are associated with NADase domain-containing proteins (TIR-APAZ or SIR2-APAZ) encoded in the same operon. They confer immunity against mobile genetic elements, such as bacteriophages and plasmids, by inducing NAD depletion upon recognition of target nucleic acids. However, the molecular mechanisms underlying the activation of such prokaryotic NADase/Ago immune systems remain unknown. Here, we report multiple cryo-EM structures of NADase/Ago complexes from two distinct systems (TIR-APAZ/Ago and SIR2-APAZ/Ago). Target DNA binding triggers tetramerization of the TIR-APAZ/Ago complex by a cooperative self-assembly mechanism, while the heterodimeric SIR2-APAZ/Ago complex does not assemble into higher-order oligomers upon target DNA binding. However, the NADase activities of these two systems are unleashed via a similar closed-to-open transition of the catalytic pocket, albeit by different mechanisms. Furthermore, a functionally conserved sensor loop is employed to inspect the guide RNA-target DNA base pairing and facilitate the conformational rearrangement of Ago proteins required for the activation of these two systems. Overall, our study reveals the mechanistic diversity and similarity of Ago protein-associated NADase systems in prokaryotic immune response.
Topics: Argonaute Proteins; NAD+ Nucleosidase; NAD; Bacteria; DNA
PubMed: 37311833
DOI: 10.1038/s41422-023-00839-7 -
Nature Nanotechnology Dec 2023The DNA double helix structure is stabilized by base-pairing and base-stacking interactions. However, a comprehensive understanding of dinucleotide base-stacking...
The DNA double helix structure is stabilized by base-pairing and base-stacking interactions. However, a comprehensive understanding of dinucleotide base-stacking energetics is lacking. Here we combined multiplexed DNA-based point accumulation in nanoscale topography (DNA-PAINT) imaging with designer DNA nanostructures and measured the free energy of dinucleotide base stacking at the single-molecule level. Multiplexed imaging enabled us to extract the binding kinetics of an imager strand with and without additional dinucleotide stacking interactions. The DNA-PAINT data showed that a single additional dinucleotide base stacking results in up to 250-fold stabilization for the DNA duplex nanostructure. We found that the dinucleotide base-stacking energies vary from -0.95 ± 0.12 kcal mol to -3.22 ± 0.04 kcal mol for C|T and A|C base-stackings, respectively. We demonstrate the application of base-stacking energetics in designing DNA-PAINT probes for multiplexed super-resolution imaging, and efficient assembly of higher-order DNA nanostructures. Our results will aid in designing functional DNA nanostructures, and DNA and RNA aptamers, and facilitate better predictions of the local DNA structure.
Topics: DNA; Base Pairing; Nanostructures; Nanotechnology; Nucleic Acid Conformation; Thermodynamics
PubMed: 37591937
DOI: 10.1038/s41565-023-01485-1