Yeast tRNA Research Articles

Quantitative detection of pseudouridine in RNA by mass spectrometry.

Pseudouridine (Ψ) is one of the most prevalent and dynamic modification in RNA, and was shown to evade the host immune response in mRNA vaccines. Despite its significance, the biological role of Ψ remains poorly understood as certain key limitations and challenges in the detection of Ψ are yet to be overcome. In account of this, we report the usage of a chemical labelling strategy for the first quantitative detection of Ψ by mass spectrometry. We demonstrate a labelling efficiency exceeding 99% in isolated yeast tRNAs hosting multiple Ψs. LC-MS/MS analysis enables precise mapping of Ψ at single-base resolution, while simultaneously capturing a wide array of additional post-transcriptional modifications, which is not achieved with current sequencing technologies. This advancement may help unravel the dynamics and biological implications of Ψ, shedding light on its interplay with other modifications and deepening our understanding of its functional role.

Combining Nanopore direct RNA sequencing with genetics and mass spectrometry for analysis of T-loop base modifications across 42 yeast tRNA isoacceptors.

Transfer RNAs (tRNAs) contain dozens of chemical modifications. These modifications are critical for maintaining tRNA tertiary structure and optimizing protein synthesis. Here we advance the use of Nanopore direct RNA-sequencing (DRS) to investigate the synergy between modifications that are known to stabilize tRNA structure. We sequenced the 42 cytosolic tRNA isoacceptors from wild-type yeast and five tRNA-modifying enzyme knockout mutants. These data permitted comprehensive analysis of three neighboring and conserved modifications in T-loops: 5-methyluridine (m5U54), pseudouridine (Ψ55),and 1-methyladenosine (m1A58). Our results were validated using direct measurements of chemical modifications by mass spectrometry. We observed concerted T-loop modification circuits-the potent influence of Ψ55 for subsequent m1A58 modification on more tRNA isoacceptors than previously observed. Growing cells under nutrient depleted conditions also revealed a novel condition-specific increase in m1A58 modification on some tRNAs. A global and isoacceptor-specific classification strategy was developed to predict the status of T-loop modifications from a user-input tRNA DRS dataset, applicable to other conditions and tRNAs in other organisms. These advancements demonstrate how orthogonal technologies combined with genetics enable precise detection of modification landscapes of individual, full-length tRNAs, at transcriptome-scale.

Adapting nanopore sequencing basecalling models for modification detection via incremental learning and anomaly detection

We leverage machine learning approaches to adapt nanopore sequencing basecallers for nucleotide modification detection. We first apply the incremental learning (IL) technique to improve the basecalling of modification-rich sequences, which are usually of high biological interest. With sequence backbones resolved, we further run anomaly detection (AD) on individual nucleotides to determine their modification status. By this means, our pipeline promises the single-molecule, single-nucleotide, and sequence context-free detection of modifications. We benchmark the pipeline using control oligos, further apply it in the basecalling of densely-modified yeast tRNAs and E.coli genomic DNAs, the cross-species detection of N6-methyladenosine (m6A) in mammalian mRNAs, and the simultaneous detection of N1-methyladenosine (m1A) and m6A in human mRNAs. Our IL-AD workflow is available at: https://github.com/wangziyuan66/IL-AD.

The non-negligible non-specific adsorption of oligonucleotides in target-immobilized Mag-SELEX

Target-immobilized magnetic beads-based Systematic Evolution of Ligands by Exponential Enrichment (target-immobilized Mag-SELEX) has emerged as a powerful tool for aptamer selection owing to its convenience, efficiency, and versatility. However, in this study we systematically investigated non-specific adsorption in target-immobilized Mag-SELEX and found that the non-specific adsorption of the oligonucleotides to target-labeled magnetic beads was comparable to that of the screening libraries, indicating a substantial portion of captured sequences likely stem from non-specific adsorption. Longer nucleic acid sequences (80 nt and above, such as polyA80 and yeast tRNA) were found to attenuate this non-specific adsorption, with more complex higher-order structures demonstrating greater efficacy, while dNTP and short sequences such as primer sequences (20 nt), polyT(59), or polyA(59), did not possess this capability. Various evidence suggested that hydrophobic interactions and other weak interactions may be the primary underlying cause of non-specific adsorption. Additionally, surface modification of magnetic beads with polar molecule polyethylene glycol (PEG) also yielded a significant reduction in non-specific adsorption. In conclusion, our research underscores the critical importance of closely monitoring non-specific adsorption in target-immobilized Mag-SELEX.

Palladium(ii), platinum(ii), and silver(i) complexes with 3-acetylcoumarin benzoylhydrazone Schiff base: Synthesis, characterization, biomolecular interactions, cytotoxic activity, and computational studies.

New Pd(ii) (C1), Pt(ii) (C2), and Ag(i) (C3) complexes derived from 3-acetylcoumarin benzoylhydrazone (HL) Schiff base were synthesized and characterized by FTIR, 1H NMR, UV-visible spectroscopies along with elemental analysis (C, H, N), magnetic, molar conductivity measurements, and DFT calculations. The obtained results suggested that the ligand had different behaviors in the complexes: mono-negative tridentate (C1) and neutral tridentate (C2) as an ONO-donor and neutral bidentate (C3) as an ON-donor. Quantum chemistry calculations were performed to validate the stability of the suggested geometries and indicated that all the complexes possess tetra-coordinated metal ions. The binding affinity of all the compounds toward calf thymus (ctDNA), yeast (tRNA), and bovine serum albumin (BSA) was evaluated by absorption/emission spectral titration studies, which revealed the intercalative binding to ctDNA and tRNA and static binding upon complex formation with BSA. Molecular insights into the binding affinity of the characterized complexes were provided through conducting molecular docking analysis. Moreover, the cytotoxic activity (in vitro) of the compounds was screened against human cancerous cell lines and a non-cancerous lung fibroblast (WI38) one using cis-platin as a reference drug. The IC50 and selective index (SI) values indicated the higher cytotoxic activity of all the metal complexes compared to their parent ligand. Among all the compounds, the complex C2 showed the highest activity. These results confirmed the improvement of the anticancer activity of the ligand by incorporating the metal ions. In addition, flow cytometry results showed that complexes C1 and C2 induced cell cycle arrest at S and G1/S, respectively.

Non-covalent binding interaction of bioactive coumarin esculetin with calf thymus DNA and yeast transfer RNA: A detailed investigation to decipher the binding affinities, binding location, interacting forces and structural alterations at a molecular level

Esculetin is a well-known coumarin derivative found abundantly in nature possessing an extensive array of pharmacological and therapeutic properties. Consequently, to comprehend its molecular recognition mechanism, our objective is to conduct a complete investigation of its interactions with the nucleic acid, specifically ct-DNA, and t-RNA, using spectroscopic and computational techniques. The intrinsic fluorescence of esculetin is quenched when it interacts with ct-DNA and t-RNA, and this occurs through a static quenching mechanism. The thermodynamic parameters demonstrated that the interaction is influenced by hydrogen bonding and weak van der Waals forces. CD and FT-IR results revealed no conformational changes in ct-DNA and t-RNA structure on binding with esculetin. Furthermore, competitive displacement assay with ethidium bromide, melting temperature, viscosity measurement, and potassium iodide quenching experiments, reflected that esculetin probably binds to the minor groove of ct-DNA. The molecular docking results provided further confirmation for the spectroscopic findings, including the binding location of esculetin and binding energies of esculetin complexes with ct-DNA and t-RNA. Molecular dynamics simulation studies demonstrated the conformational stability and flexibility of nucleic acids.

TRNA m1G9 modification depends on substrate-specific RNA conformational changes induced by the methyltransferase Trm10

The methyltransferase Trm10 modifies a subset of tRNAs on the base N1 position of the ninth nucleotide in the tRNA core. Trm10 is conserved throughout Eukarya and Archaea, and mutations in the human gene (TRMT10A) have been linked to neurological disorders such as microcephaly and intellectual disability, as well as defects in glucose metabolism. Of the 26 tRNAs in yeast with guanosine at position 9, only 13 are substrates for Trm10. However, no common sequence or other posttranscriptional modifications have been identified among these substrates, suggesting the presence of some other tRNA feature(s) that allow Trm10 to distinguish substrate from nonsubstrate tRNAs. Here, we show that substrate recognition by Saccharomyces cerevisiae Trm10 is dependent on both intrinsic tRNA flexibility and the ability of the enzyme to induce specific tRNA conformational changes upon binding. Using the sensitive RNA structure-probing method SHAPE, conformational changes upon binding to Trm10 in tRNA substrates, but not nonsubstrates, were identified and mapped onto a model of Trm10-bound tRNA. These changes may play an important role in substrate recognition by allowing Trm10 to gain access to the target nucleotide. Our results highlight a novel mechanism of substrate recognition by a conserved tRNA modifying enzyme. Further, these studies reveal a strategy for substrate recognition that may be broadly employed by tRNA-modifying enzymes which must distinguish between structurally similar tRNA species.

Crosstalk between the tRNA methyltransferase Trm1 and RNA chaperone La influences eukaryotic tRNA maturation

tRNAs undergo an extensive maturation process involving posttranscriptional modifications often associated with tRNA structural stability and promoting the native fold. Impaired posttranscriptional modification has been linked to human disease, likely through defects in translation, mitochondrial function, and increased susceptibility to degradation by various tRNA decay pathways. More recently, evidence has emerged that bacterial tRNA modification enzymes can act as tRNA chaperones to guide tRNA folding in a manner independent from catalytic activity. Here, we provide evidence that the fission yeast tRNA methyltransferase Trm1, which dimethylates nuclear- and mitochondrial-encoded tRNAs at G26, can also promote tRNA functionality in the absence of catalysis. We show that WT and catalytic-dead Trm1 are active in an invivo tRNA-mediated suppression assay and possess RNA strand annealing and dissociation activity invitro, similar to previously characterized RNA chaperones. Trm1 and the RNA chaperone La have previously been proposed to function synergistically in promoting tRNA maturation, yet we surprisingly demonstrate that La binding to nascent pre-tRNAs decreases Trm1 tRNA dimethylation invivo and invitro. Collectively, these results support the hypothesis for tRNA modification enzymes that combine catalytic and noncatalytic activities to promote tRNA maturation, as well as expand our understanding of how La function can influence tRNA modification.

Expanded tRNA methyltransferase family member TRMT9B regulates synaptic growth and function.

Nervous system function rests on the formation of functional synapses between neurons. We have identified TRMT9B as a new regulator of synapse formation and function in Drosophila. TRMT9B has been studied for its role as a tumor suppressor and is one of two metazoan homologs of yeast tRNA methyltransferase 9 (Trm9), which methylates tRNA wobble uridines. Whereas Trm9 homolog ALKBH8 is ubiquitously expressed, TRMT9B is enriched in the nervous system. However, in the absence of animal models, TRMT9B's role in the nervous system has remained unstudied. Here, we generate null alleles of TRMT9B and find it acts postsynaptically to regulate synaptogenesis and promote neurotransmission. Through liquid chromatography-mass spectrometry, we find that ALKBH8 catalyzes canonical tRNA wobble uridine methylation, raising the question of whether TRMT9B is a methyltransferase. Structural modeling studies suggest TRMT9B retains methyltransferase function and, in vivo, disruption of key methyltransferase residues blocks TRMT9B's ability to rescue synaptic overgrowth, but not neurotransmitter release. These findings reveal distinct roles for TRMT9B in the nervous system and highlight the significance of tRNA methyltransferase family diversification in metazoans.

Reproducible evaluation of transposable element detectors with McClintock 2 guides accurate inference of Ty insertion patterns in yeast

BackgroundMany computational methods have been developed to detect non-reference transposable element (TE) insertions using short-read whole genome sequencing data. The diversity and complexity of such methods often present challenges to new users seeking to reproducibly install, execute, or evaluate multiple TE insertion detectors.ResultsWe previously developed the McClintock meta-pipeline to facilitate the installation, execution, and evaluation of six first-generation short-read TE detectors. Here, we report a completely re-implemented version of McClintock written in Python using Snakemake and Conda that improves its installation, error handling, speed, stability, and extensibility. McClintock 2 now includes 12 short-read TE detectors, auxiliary pre-processing and analysis modules, interactive HTML reports, and a simulation framework to reproducibly evaluate the accuracy of component TE detectors. When applied to the model microbial eukaryote Saccharomyces cerevisiae, we find substantial variation in the ability of McClintock 2 components to identify the precise locations of non-reference TE insertions, with RelocaTE2 showing the highest recall and precision in simulated data. We find that RelocaTE2, TEMP, TEMP2 and TEBreak provide consistent estimates of sim50 non-reference TE insertions per strain and that Ty2 has the highest number of non-reference TE insertions in a species-wide panel of sim1000 yeast genomes. Finally, we show that best-in-class predictors for yeast applied to resequencing data have sufficient resolution to reveal a dyad pattern of integration in nucleosome-bound regions upstream of yeast tRNA genes for Ty1, Ty2, and Ty4, allowing us to extend knowledge about fine-scale target preferences revealed previously for experimentally-induced Ty1 insertions to spontaneous insertions for other copia-superfamily retrotransposons in yeast.ConclusionMcClintock (https://github.com/bergmanlab/mcclintock/) provides a user-friendly pipeline for the identification of TEs in short-read WGS data using multiple TE detectors, which should benefit researchers studying TE insertion variation in a wide range of different organisms. Application of the improved McClintock system to simulated and empirical yeast genome data reveals best-in-class methods and novel biological insights for one of the most widely-studied model eukaryotes and provides a paradigm for evaluating and selecting non-reference TE detectors in other species.

Effect of hydrophobicity and size of the ligands on the intercalative binding interactions of some metallo-surfactants containing π-conjugated systems with yeast tRNA

The intercalative yeast t-RNA binding behavior of some metallo-surfactant complexes, Co(ip)2(TA)2](ClO4)3 (1) and [Co(dpq)2(TA)2](ClO4)3 (2) where TA = Tetradecylamine (Myristylamine), ip = imidazo[4,5-f][1,10]phenanthroline and dpq = dipyrido[3,2-d:2′-3′-f]quinoxaline containing π-conjugated systems (both below and above critical micelle concentration) have been investigated by means of absorption spectral titration, competitive binding, circular dichroism, cyclic voltammetry, and viscometry measurements. Absorption spectral titration results implicate yeast tRNA has significant effects on the binding behaviors of two surfactant complexes via intercalative mode showed a significant absorption band of hypochromicity with red shift. The intrinsic binding constant values below and above CMC were determined as K b = 6.12 × 105 M−1, 2.31 × 106 M−1, for complex (1) and 7.23 × 105 M−1, 3.57 × 106 M−1, for complex (2). In both sets of complexes (1) and (2), the complexes bind more strongly to yeast tRNA in the above critical micelle concentration can be hydrophobic and confirm intercalation. Competitive displacement studies confirmed that complexes bind to yeast tRNA via intercalative mode. Cyclic voltammetry studies suggest the increasing amounts of yeast tRNA, the cathodic potential E pc for the two complexes shows a positive shift in peak potential indicated the process of binding via intercalation. These observations were further validated by CD, and hydrodynamic measurements. All these studies suggesting that a surfactant complex binds to yeast tRNA appear to be mainly intercalative because of hydrophobicity due to extending aromaticity of the π system of the ligand and planarity of the complex has a significant effect on tRNA binding affinity increasing in the order of complexes containing ligands ip < dpq. Communicated by Ramaswamy H. Sarma

Structural insights into interactions of new polymeric (μ–oxo) bridged Cu(II) complexes of taurine with yeast tRNA by spectroscopic and computational approaches and its application towards chemoresistant cancer lines

RNA-targeted drugs are considered as safe treatment option for the cure of many chronic diseases preventing off-targeted delivery and acute toxic manifestations. FDA has approved many such RNA therapies in different phases of clinical trials, validating their use for the treatment of various chronic diseases. We report herein, new water-soluble (μ–oxo) bridged polymeric Cu(II) complexes of taurine (2–aminoethane sulfonic acid) complexes 1 and 2. The therapeutic potency of 1 and 2 was ascertained by studying biophysical interactions with tRNA/ct-DNA. The experimental results demonstrated that the complexes interacted avidly to nucleic acids through intercalation mode depicting a specific preference for tRNA in comparison to ct–DNA and, moreover 2 showed higher binding propensity than 1. The electrophoretic behaviour of the complexes with plasmid pBR322 DNA and tRNA were examined by gel mobility assay that revealed a concentration-dependent activity with complex 2 performing more efficient cleavage as compared to complex 1. Cytotoxicity results on cancer cell strains displayed higher cytotoxicity than complex 1 against treated cancer cells. The synthesized copper(II) taurine complexes have met the basic criteria of anticancer drug design as they are structurally well-characterized, exhibiting good solubility in water, lipophilic in nature with superior intercalating propensity towards tRNA and cytotoxic in nature.

Enhancement of the activity of the antimicrobial peptides HNP1 and LL-37 by bovine pancreatic ribonuclease A.

Background: HNP1, LL-37, and HBD1 are antimicrobial against Escherichia coli ATCC 25922 at the standard inoculum but less active at higher inocula. Methods: The virtual colony count (VCC) microbiological assay was adapted for high inocula and the addition of yeast tRNA and bovine pancreatic ribonuclease A (RNase). 96-well plates were read for 12 hours in a Tecan Infinite M1000 plate reader and photographed under 10x magnification. Results: Adding tRNA 1:1 wt/wt to HNP1 at the standard inoculum almost completely abrogated activity. Adding RNase 1:1 to HNP1 at the standard inoculum of 5x10 5 CFU/mL did not enhance activity. Increasing the inoculum to 6.25x10 7 CFU/mL almost abrogated HNP1 activity. However, adding RNase 25:1 to HNP1 enhanced activity at the highest tested concentration of HNP1. Adding both tRNA and RNase resulted in enhanced activity, indicating that the enhancement effect of RNase overwhelms the inhibiting effect of tRNA when both are present. HBD1 activity at the standard inoculum was almost completely abrogated by the addition of tRNA, but LL-37 activity was only slightly inhibited by tRNA. At the high inoculum, LL-37 activity was enhanced by RNase. HBD1 activity was not enhanced by RNase. RNase was not antimicrobial in the absence of antimicrobial peptides. Cell clumps were observed at the high inoculum in the presence of all three antimicrobial peptides and at the standard inoculum in the presence of HNP1+tRNA and HBD1+tRNA. Conclusions: Antimicrobial peptide-ribonuclease combinations have the potential to be active against high cell concentrations, conditions where the antimicrobial agent alone is relatively ineffective.

An Optimized Quantitative Pull-Down Analysis of RNA-Binding Proteins Using Short Biotinylated RNA.

Protein-RNA interactions regulate gene expression and cellular functions at transcriptional and post-transcriptional levels. For this reason, identifying the binding partners of an RNA of interest remains of high importance to unveil the mechanisms behind many cellular processes. However, RNA molecules might interact transiently and dynamically with some RNA-binding proteins (RBPs), especially with non-canonical ones. Hence, improved methods to isolate and identify such RBPs are greatly needed. To identify the protein partners of a known RNA sequence efficiently and quantitatively, we developed a method based on the pull-down and characterization of all interacting proteins, starting from cellular total protein extract. We optimized the protein pull-down using biotinylated RNA pre-loaded on streptavidin-coated beads. As a proof of concept, we employed a short RNA sequence known to bind the neurodegeneration-associated protein TDP-43 and a negative control of a different nucleotide composition but the same length. After blocking the beads with yeast tRNA, we loaded the biotinylated RNA sequences on the streptavidin beads and incubated them with the total protein extract from HEK 293T cells. After incubation and several washing steps to remove nonspecific binders, we eluted the interacting proteins with a high-salt solution, compatible with the most commonly used protein quantification reagents and with sample preparation for mass spectrometry. We quantified the enrichment of TDP-43 in the pull-down performed with the known RNA binder compared to the negative control by mass spectrometry. We used the same technique to verify the selective interactions of other proteins computationally predicted to be unique binders of our RNA of interest or of the control. Finally, we validated the protocol by western blot via the detection of TDP-43 with an appropriate antibody. This protocol will allow the study of the protein partners of an RNA of interest in near-to-physiological conditions, helping uncover unique and unpredicted protein-RNA interactions.

Reconstitution of Mammalian Mitochondrial Translation System Capable of Long Polypeptide Synthesis.

Mammalian mitochondria have their own dedicated protein synthesis system, which produces 13 essential subunits of the oxidative phosphorylation complexes. Here, we describe the in vitro reconstitution of the mammalian mitochondrial translation system, utilizing purified recombinant mitochondrial translation factors, 55S ribosomes from pig liver mitochondria, and a heterologous yeast tRNA mixture. The system is capable of translating leaderless mRNAs encoding model proteins, such as nanoluciferase with a molecular weight of 19kDa, and is readily applicable for in vitro evaluations of mRNAs and nascent peptide chain sequences, as well as factors and small molecules that affect mitochondrial translation.

Enhancement of the activity of the antimicrobial peptides HNP1 and LL-37 by bovine pancreatic ribonuclease A

Background: HNP1, LL-37, and HBD1 are antimicrobial against Escherichia coli ATCC 25922 at the standard inoculum but less active at higher inocula. Methods: The virtual colony count (VCC) microbiological assay was adapted for high inocula and the addition of yeast tRNA and bovine pancreatic ribonuclease A (RNase). 96-well plates were read for 12 hours in a Tecan Infinite M1000 plate reader and photographed under 10x magnification. Results: Adding tRNA 1:1 wt/wt to HNP1 at the standard inoculum almost completely abrogated activity. Adding RNase 1:1 to HNP1 at the standard inoculum of 5x105 CFU/mL did not enhance activity. Increasing the inoculum to 6.25x107 CFU/mL almost abrogated HNP1 activity. However, adding RNase 25:1 to HNP1 enhanced activity at the highest tested concentration of HNP1. Adding both tRNA and RNase resulted in enhanced activity, indicating that the enhancement effect of RNase overwhelms the inhibiting effect of tRNA when both are present. HBD1 activity at the standard inoculum was almost completely abrogated by the addition of tRNA, but LL-37 activity was only slightly inhibited by tRNA. At the high inoculum, LL-37 activity was enhanced by RNase. HBD1 activity was not enhanced by RNase. RNase was not antimicrobial in the absence of antimicrobial peptides. Cell clumps were observed at the high inoculum in the presence of all three antimicrobial peptides and at the standard inoculum in the presence of HNP1+tRNA and HBD1+tRNA. Conclusions: Antimicrobial peptide-ribonuclease combinations have the potential to be active against high cell concentrations and biofilms, conditions where the antimicrobial agent alone is relatively ineffective.

Fitness Landscape Analysis of a tRNA Gene Reveals that the Wild Type Allele is Sub-optimal, Yet Mutationally Robust.

Fitness landscape mapping and the prediction of evolutionary trajectories on these landscapes are major tasks in evolutionary biology research. Evolutionary dynamics is tightly linked to the landscape topography, but this relation is not straightforward. Here, we analyze a fitness landscape of a yeast tRNA gene, previously measured under four different conditions. We find that the wild type allele is sub-optimal, and 8–10% of its variants are fitter. We rule out the possibilities that the wild type is fittest on average on these four conditions or located on a local fitness maximum. Notwithstanding, we cannot exclude the possibility that the wild type might be fittest in some of the many conditions in the complex ecology that yeast lives at. Instead, we find that the wild type is mutationally robust (“flat”), while more fit variants are typically mutationally fragile. Similar observations of mutational robustness or flatness have been so far made in very few cases, predominantly in viral genomes.

Enhancement of the activity of the antimicrobial peptides HNP1 and LL-37 by bovine pancreatic ribonuclease A

Background: HNP1, LL-37, and HBD1 are antimicrobial against Escherichia coli ATCC 25922 at the standard inoculum but less active at higher inocula. Methods: The virtual colony count (VCC) microbiological assay was adapted for high inocula and the addition of yeast tRNA and bovine pancreatic ribonuclease A (RNase). 96-well plates were read for 12 hours in a Tecan Infinite M1000 plate reader and photographed under 10x magnification. Results: Adding tRNA 1:1 to HNP1 at the standard inoculum almost completely abrogated activity. Adding RNase 1:1 to HNP1 at the standard inoculum of 5x105 CFU/mL did not enhance activity. Increasing the inoculum to 6.25x107 CFU/mL almost abrogated HNP1 activity. However, adding RNase 25:1 to HNP1 enhanced activity. Adding both tRNA and RNase resulted in enhanced activity, indicating that the enhancement effect of RNase overwhelms the inhibiting effect of tRNA when both are present. HBD1 activity at the standard inoculum was almost completely abrogated by the addition of tRNA, but LL-37 activity was only slightly inhibited by tRNA. At the high inoculum, LL-37 activity was enhanced by RNase. HBD1 activity was not enhanced by RNase. RNase was not antimicrobial in the absence of antimicrobial peptides. Cell clumps were observed at the high inoculum in the presence of all three antimicrobial peptides and at the standard inoculum in the presence of HNP1+tRNA. Conclusions: Antimicrobial peptide-ribonuclease combinations have the potential to be active against high cell concentrations and biofilms, conditions where the antimicrobial agent alone is relatively ineffective.

Terpyridyl ruthenium complexes as visible spectral probe for poly(A) RNA and bifunctional TAR RNA binders and HIV-1 reverse transcriptase inhibitors

HIV-1 reverse transcriptase (RT) inhibitors play essential role in anti-HIV therapy. The vast majority of them target the enzymes, while very few are able to bind to the viral RNA. Here we designed and synthesized two new terpyridyl Ru(II) complexes with HIV-1 TAR RNA binding groups. The complex RuTz2 exhibited a remarkable selectivity for poly(A) RNA over calf thymus DNA, total RNA and yeast transfer RNA, generated significant visible spectral response and inhibited the reverse transcription of poly(A) RNA to poly(dT) cDNA by M-MuLV RT. Moreover, RuTz2 was found to target the HIV-1 TAR RNA tightly and selectively by molecular recognition of hydrogen bonds, further stabilize the Ru(II)-RNA binding complex by electrostatic attraction, and efficiently inhibit the HIV-1 RT. These terpyridyl Ru(II) complexes also showed low toxicity to normal cells, which would greatly reduce its harmful side-effect on normal cells in drug application. This work also provides valuable drug design strategies for AIDS and other RT related diseases researches, such as HCV, EBOV and SARS-CoV-2.

Review of: "Deep sequencing of yeast and mouse tRNAs and tRNA fragments using OTTR"

Review of: "Deep sequencing of yeast and mouse tRNAs and tRNA fragments using OTTR"