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Nucleic Acids Research
IRL Press Ltd
Nucleic Acids Research

IRL Press Ltd

0305-1048

Nucleic Acids Research/Journal Nucleic Acids ResearchSCICCRISTP
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    Prp43/DHX15 exemplify RNA helicase multifunctionality in the gene expression network

    Bohnsack, Katherine E.Kanwal, NidhiBohnsack, Markus T.
    11页
    查看更多>>摘要：Dynamic regulation of RNA folding and structure is critical for the biogenesis and function of RNAs and ribonucleoprotein (RNP) complexes. Through their nucleotide triphosphate-dependent remodelling functions, RNA helicases are key modulators of RNA/RNP structure. While some RNA helicases are dedicated to a specific target RNA, others are multifunctional and engage numerous substrate RNAs in different aspects of RNA metabolism. The discovery of such multitasking RNA helicases raises the intriguing question of how these enzymes can act on diverse RNAs but also maintain specificity for their particular targets within the RNA-dense cellular environment. Furthermore, the identification of RNA helicases that sit at the nexus between different aspects of RNA metabolism raises the possibility that they mediate cross-regulation of different cellular processes. Prominent and extensively characterized multifunctional DEAH/RHA-box RNA helicases are DHX15 and its Saccharomyces cerevisiae (yeast) homologue Prp43. Due to their central roles in key cellular processes, these enzymes have also served as prototypes for mechanistic studies elucidating the mode of action of this type of enzyme. Here, we summarize the current knowledge on the structure, regulation and cellular functions of Prp43/DHX15, and discuss the general concept and implications of RNA helicase multifunctionality.
      原文链接:
    • NSTL
    • Oxford Univ Press

    The influenza A virus genome packaging network - complex, flexible and yet unsolved

    Jakob, CeliaPaul-Stansilaus, RithuSchwemmle, MartinMarquet, Roland...
    16页
    查看更多>>摘要：The genome of influenza A virus (IAV) consists of eight unique viral RNA segments. This genome organization allows genetic reassortment between co-infecting IAV strains, whereby new IAVs with altered genome segment compositions emerge. While it is known that reassortment events can create pandemic IAVs, it remains impossible to anticipate reassortment outcomes with pandemic prospects. Recent research indicates that reassortment is promoted by a viral genome packaging mechanism that delivers the eight genome segments as a supramolecular complex into the virus particle. This finding holds promise of predicting pandemic IAVs by understanding the intermolecular interactions governing this genome packaging mechanism. Here, we critically review the prevailing mechanistic model postulating that IAV genome packaging is orchestrated by a network of intersegmental RNA-RNA interactions. Although we find supporting evidence, including segment-specific packaging signals and experimentally proposed RNA-RNA interaction networks, this mechanistic model remains debatable due to a current shortage of functionally validated intersegmental RNA-RNA interactions. We speculate that identifying such functional intersegmental RNA-RNA contacts might be hampered by limitations of the utilized probing techniques and the inherent complexity of the genome packaging mechanism. Nevertheless, we anticipate that improved probing strategies combined with a mutagenesis-based validation could facilitate their discovery.
      原文链接:
    • NSTL
    • Oxford Univ Press

    2 '-O-Methylation of the second transcribed nucleotide within the mRNA 5 ' cap impacts the protein production level in a cell-specific manner and contributes to RNA immune evasion

    Drazkowska, KarolinaTomecki, RafalWarminski, MarcinBaran, Natalia...
    21页
    查看更多>>摘要：In mammals, m(7)G-adjacent nucleotides undergo extensive modifications. Ribose of the first or first and second transcribed nucleotides can be subjected to 2 '-O-methylation to form cap1 or cap2, respectively. When the first transcribed nucleotide is 2 '-O-methylated adenosine, it can be additionally modified to N6,2 '-O-dimethyladenosine (m(6)A(m)). Recently, the crucial role of cap1 in distinguishing between 'self' and 'non-self' in mammalian cells during viral infection was revealed. Here, we attempted to understand the impact of cap methylations on RNA-related processes. Therefore, we synthesized tetranucleotide cap analogues and used them for RNA capping during in vitro transcription. Using this tool, we found that 2 '-O-methylation of the second transcribed nucleotide within the mRNA 5 ' cap influences protein production levels in a cell-specific manner. This modification can strongly hamper protein biosynthesis or have no influence on protein production levels, depending on the cell line. Interestingly, 2 '-O-methylation of the second transcribed nucleotide and the presence of m(6)A(m) as the first transcribed nucleotide serve as determinants that define transcripts as 'self' and contribute to transcript escape from the host innate immune response. Additionally, cap methylation status does not influence transcript affinity towards translation initiation factor eIF4E or in vitro susceptibility to decapping by DCP2; however, we observe the resistance of cap2-RNA to DXO (decapping exoribonuclease)-mediated decapping and degradation.
      原文链接:
    • NSTL
    • Oxford Univ Press

    Markov state models elucidate the stability of DNA influenced by the chiral 5S-Tg base

    Wang, Shu-DongZhang, Ru-BoEriksson, Leif A.
    11页
    查看更多>>摘要：The static and dynamic structures of DNA duplexes affected by 5S-Tg (Tg, Thymine glycol) epimers were studied using MD simulations and Markov State Models (MSMs) analysis. The results show that the 5S,6S-Tg base caused little perturbation to the helix, and the base-flipping barrier was determined to be 4.4 kcal mol(-1) through the use of enhanced sampling meta-eABF calculations, comparable to 5.4 kcal mol(-1) of the corresponding thymine flipping. Two conformations with the different hydrogen bond structures between 5S,6R-Tg and A19 were identified in several independent MD trajectories. The 5S,6R-Tg:O6H(O6)center dot center dot center dot N1:A19 hydrogen bond is present in the high-energy conformation displaying a clear helical distortion, and near barrier-free Tg base flipping. The low-energy conformation always maintains Watson-Crick base pairing between 5S,6R-Tg and A19, and 5S-Tg base flipping is accompanied by a small barrier of ca. 2.0 KBT (T = 298 K). The same conformations are observed in the MSMs analysis. Moreover, the transition path and metastable structures of the damaged base flipping are for the first time verified through MSMs analysis. The data clearly show that the epimers have completely different influence on the stability of the DNA duplex, thus implying different enzymatic mechanisms for DNA repair.
      原文链接:
    • NSTL
    • Oxford Univ Press

    Painters in chromatin: a unified quantitative framework to systematically characterize epigenome regulation and memory

    Abdulla, Amith Z.Vaillant, CedricJost, Daniel
    22页
    查看更多>>摘要：In eukaryotes, many stable and heritable phenotypes arise from the same DNA sequence, owing to epigenetic regulatory mechanisms relying on the molecular cooperativity of 'reader-writer' enzymes. In this work, we focus on the fundamental, generic mechanisms behind the epigenome memory encoded by post-translational modifications of histone tails. Based on experimental knowledge, we introduce a unified modeling framework, the painter model, describing the mechanistic interplay between sequence-specific recruitment of chromatin regulators, chromatin-state-specific reader-writer processes and long-range spreading mechanisms. A systematic analysis of the model building blocks highlights the crucial impact of tridimensional chromatin organization and state-specific recruitment of enzymes on the stability of epigenomic domains and on gene expression. In particular, we show that enhanced 3D compaction of the genome and enzyme limitation facilitate the formation of ultra-stable, confined chromatin domains. The model also captures how chromatin state dynamics impact the intrinsic transcriptional properties of the region, slower kinetics leading to noisier expression. We finally apply our framework to analyze experimental data, from the propagation of gamma H2AX around DNA breaks in human cells to the maintenance of heterochromatin in fission yeast, illustrating how the painter model can be used to extract quantitative information on epigenomic molecular processes.
      原文链接:
    • NSTL
    • Oxford Univ Press

    DNAffinity: a machine-learning approach to predict DNA binding affinities of transcription factors

    Barissi, SandroSala, AlbaWieczor, MiloszBattistini, Federica...
    10页
    查看更多>>摘要：We present a physics-based machine learning approach to predict in vitro transcription factor binding affinities from structural and mechanical DNA properties directly derived from atomistic molecular dynamics simulations. The method is able to predict affinities obtained with techniques as different as uPBM, gcPBM and HT-SELEX with an excellent performance, much better than existing algorithms. Due to its nature, the method can be extended to epigenetic variants, mismatches, mutations, or any non-coding nucleobases. When complemented with chromatin structure information, our in vitro trained method provides also good estimates of in vivo binding sites in yeast.
      原文链接:
    • NSTL
    • Oxford Univ Press

    Performance evaluation of differential splicing analysis methods and splicing analytics platform construction

    Li, KuokuoLuo, TengfeiZhu, YanWang, An...
    12页
    查看更多>>摘要：A proportion of previously defined benign variants or variants of uncertain significance in humans, which are challenging to identify, may induce an abnormal splicing process. An increasing number of methods have been developed to predict splicing variants, but their performance has not been completely evaluated using independent benchmarks. Here, we manually sourced similar to 50 000 positive/negative splicing variants from > 8000 studies and selected the independent splicing variants to evaluate the performance of prediction methods. These methods showed different performances in recognizing splicing variants in donor and acceptor regions, reminiscent of different weight coefficient applications to predict novel splicing variants. Of these methods, 66.67% exhibited higher specificities than sensitivities, suggesting that more moderate cut-off values are necessary to distinguish splicing variants. Moreover, the high correlation and consistent prediction ratio validated the feasibility of integration of the splicing prediction method in identifying splicing variants. We developed a splicing analytics platform called SPCards, which curates splicing variants from publications and predicts splicing scores of variants in genomes. SPCards also offers variant-level and gene-level annotation information, including allele frequency, non-synonymous prediction and comprehensive functional information. SPCards is suitable for high-throughput genetic identification of splicing variants, particularly those located in non-canonical splicing regions.
      原文链接:
    • NSTL
    • Oxford Univ Press

    Differential dependencies of human RNA polymerase II promoters on TBP, TAF1, TFIIB and XPB

    Santana, Juan F.Collins, Geoffrey S.Parida, MrutyunjayaLuse, Donal S....
    22页
    查看更多>>摘要：The effects of rapid acute depletion of components of RNA polymerase II (Pol II) general transcription factors (GTFs) that are thought to be critical for formation of preinitiation complexes (PICs) and initiation in vitro were quantified in HAP1 cells using precision nuclear run-on sequencing (PRO-Seq). The average dependencies for each factor across >70 000 promoters varied widely even though levels of depletions were similar. Some of the effects could be attributed to the presence or absence of core promoter elements such as the upstream TBP-specificity motif or downstream G-rich sequences, but some dependencies anti-correlated with such sequences. While depletion of TBP had a large effect on most Pol III promoters only a small fraction of Pol II promoters were similarly affected. TFIIB depletion had the largest general effect on Pol II and also correlated with apparent termination defects downstream of genes. Our results demonstrate that promoter activity is combinatorially influenced by recruitment of TFIID and sequence-specific transcription factors. They also suggest that interaction of the preinitiation complex (PIC) with nucleosomes can affect activity and that recruitment of TFIID containing TBP only plays a positive role at a subset of promoters.
      原文链接:
    • NSTL
    • Oxford Univ Press

    What is a supercoiling-sensitive gene? Insights from topoisomerase I inhibition in the Gram-negative bacterium Dickeya dadantii

    Pineau, MaiwennMartis, Shiny B.Forquet, RaphaelBaude, Jessica...
    13页
    查看更多>>摘要：DNA supercoiling is an essential mechanism of bacterial chromosome compaction, whose level is mainly regulated by topoisomerase I and DNA gyrase. Inhibiting either of these enzymes with antibiotics leads to global supercoiling modifications and subsequent changes in global gene expression. In previous studies, genes responding to DNA relaxation induced by DNA gyrase inhibition were categorised as 'supercoiling-sensitive'. Here, we studied the opposite variation of DNA supercoiling in the phytopathogen Dickeya dadantii using the non-marketed antibiotic seconeolitsine. We showed that the drug is active against topoisomerase I from this species, and analysed the first transcriptomic response of a Gram-negative bacterium to topoisomerase I inhibition. We find that the responding genes essentially differ from those observed after DNA relaxation, and further depend on the growth phase. We characterised these genes at the functional level, and also detected distinct patterns in terms of expression level, spatial and orientational organisation along the chromosome. Altogether, these results highlight that the supercoiling-sensitivity is a complex feature, which depends on the action of specific topoisomerases, on the physiological conditions, and on their genomic context. Based on previous in vitro expression data of several promoters, we propose a qualitative model of SC-dependent regulation that accounts for many of the contrasting transcriptomic features observed after DNA gyrase or topoisomerase I inhibition.
      原文链接:
    • NSTL
    • Oxford Univ Press

    The DEAD-box helicase Hlc regulates basal transcription and chromatin opening of stress-responsive genes

    Jia, RuiruiLin, JiameiYou, JinLi, Shi...
    15页
    查看更多>>摘要：Stress-responsive genes are lowly transcribed under normal conditions and robustly induced in response to stress. The significant difference between basal and induced transcription indicates that the general transcriptional machinery requires a mechanism to distinguish each transcription state. However, what factors specifically function in basal transcription remains poorly understood. Using a classic model stress-responsive gene (Drosophila MtnA), we found that knockdown of the DEAD-box helicase Hlc resulted in a significant transcription attenuation of MtnA under normal, but not stressed, conditions. Mechanistically, Hlc directly binds to the MtnA locus to maintain the accessibility of chromatin near the transcriptional start site, which allows the recruitment of RNA polymerase II and subsequent MtnA transcription. Using RNA-seq, we then identified plenty of additional stress-responsive genes whose basal transcription was reduced upon knockdown of Hlc. Taken together, these data suggest that Hlc-mediated basal transcription regulation is an essential and widespread mechanism for precise control of stress-responsive genes.
      原文链接:
    • NSTL
    • Oxford Univ Press