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Journal of Biological Inorganic... Aug 2023Two chiral ruthenium(II) polypyridyl complexes, Λ-[Ru(bpy)(dppx)] (bpy = 2,2'-bipyridine, dppx = 7,8-dimethyldipyridophenazine; Λ-1) and Δ-[Ru(bpy)(dppx)]...
Two chiral ruthenium(II) polypyridyl complexes, Λ-[Ru(bpy)(dppx)] (bpy = 2,2'-bipyridine, dppx = 7,8-dimethyldipyridophenazine; Λ-1) and Δ-[Ru(bpy)(dppx)] (Δ-1) have been synthesized and characterized in this work. Interactions of Λ-1 and Δ-1 with the RNA triplex poly(U)⋅poly(A)*poly(U) have been investigated by various biophysical techniques. Spectrophotometric titrations and viscosity measurements suggested that enantiomers Λ-1 and Δ-1 bind with the triplex through intercalation, while the binding strengths of the two enantiomers toward the triplex differed only slightly from each other. Fluorescence titrations showed that although enantiomers Λ-1 and Δ-1 exhibited molecular "light switch" effects toward the triplex, the effect of Δ-1 was more marked. Furthermore, Furthermore, thermal denaturation showed that the two enantiomers have significantly different stabilizing effects on the triplex. The obtained results indicate that the racemic complex [Ru(bpy)(dppx)] is similar to a non-specific metallointercalator for the triplex investigated in this study, and chiralities of Ru(II) polypyridine complexes have an important influence on the binding and stabilizing effects of enantiomers toward the triplex. Two chiral ruthenium(II) polypyridyl complexes, Λ-[Ru(bpy)(dppx)] (bpy = 2,2'-bipyridine, dppx = 7,8-dimethyldipyridophenazine; Λ-1) and Δ-[Ru(bpy)(dppx)] (Δ-1) have been synthesized and characterized in this work. Interactions of Λ-1 and Δ-1 with the RNA triplex poly(U)⋅poly(A)*poly(U) have been investigated by various biophysical techniques. The obtained results indicate that the racemic complex [Ru(bpy)(dppx)] is similar as a non-specific metallointercalator for the triplex investigated in this study, and chiralities of Ru(II) polypyridine complexes have an important influence on the binding and stabilizing effects of enantiomers toward the triplex.
Topics: Poly A; Ruthenium; Poly U; 2,2'-Dipyridyl; RNA
PubMed: 37452869
DOI: 10.1007/s00775-023-02004-2 -
Nature Aug 2017A fundamental principle in biology is that the program for early development is established during oogenesis in the form of the maternal transcriptome. How the maternal...
A fundamental principle in biology is that the program for early development is established during oogenesis in the form of the maternal transcriptome. How the maternal transcriptome acquires the appropriate content and dosage of transcripts is not fully understood. Here we show that 3' terminal uridylation of mRNA mediated by TUT4 and TUT7 sculpts the mouse maternal transcriptome by eliminating transcripts during oocyte growth. Uridylation mediated by TUT4 and TUT7 is essential for both oocyte maturation and fertility. In comparison to somatic cells, the oocyte transcriptome has a shorter poly(A) tail and a higher relative proportion of terminal oligo-uridylation. Deletion of TUT4 and TUT7 leads to the accumulation of a cohort of transcripts with a high frequency of very short poly(A) tails, and a loss of 3' oligo-uridylation. By contrast, deficiency of TUT4 and TUT7 does not alter gene expression in a variety of somatic cells. In summary, we show that poly(A) tail length and 3' terminal uridylation have essential and specific functions in shaping a functional maternal transcriptome.
Topics: Animals; Cell Line; DNA-Binding Proteins; Female; Infertility, Female; Male; Maternal Inheritance; Mice; Mice, Knockout; Mothers; Nucleotidyltransferases; Oocytes; Organ Specificity; Poly A; RNA Stability; RNA, Messenger; Transcriptome; Uridine Monophosphate
PubMed: 28792939
DOI: 10.1038/nature23318 -
Journal of Biomolecular Structure &... 2023To reveal the effect of DNA- or RNA-specific low-molecular compounds on cellular processes on the molecular level, we have carried out the studies with the application...
To reveal the effect of DNA- or RNA-specific low-molecular compounds on cellular processes on the molecular level, we have carried out the studies with the application of spectroscopic methods. It is necessary for the understanding of structural-functional properties of nucleic acids in cell. In this work the interaction of DNA-specific thiazine dye methylene blue (MB) with synthetic polynucleotides poly(rA) and poly(rU) was studied. The interaction of MB with synthetic polyribonucleotides poly(rA) and poly(rU) was examined in the solution with high ionic strength in a wide phosphate-to-dye (P/D) range, using the absorption and fluorescence spectroscopies, as well as the fluorescence 2D spectra and 3D spectra analyses were given. Values of the fluorescence quenching constants for the complexes of MB with poly(rA) and poly(rU) were calculated (K is the Stern-Volmer quenching constant). Two different modes of MB binding to single-stranded (ss-) poly(rA) and poly(rU) and to their hybrid double-stranded (ds-) structure - poly(rA)-poly(rU) were identified. This ligand binds to ss-poly(rA) and poly(rA)-poly(rU) by semi-intercalation and electrostatic modes, but to ss-poly(rU) the prevailing mode is the electrostatic interaction.Communicated by Ramaswamy H. Sarma.
Topics: Methylene Blue; Poly A-U; Nucleic Acid Conformation; Poly A; DNA
PubMed: 36919567
DOI: 10.1080/07391102.2023.2189475 -
Scientific Reports Jun 2022The full-length double-strand cDNA sequencing, one of the RNA-Seq methods, is a powerful method used to investigate the transcriptome status of a gene of interest, such...
The full-length double-strand cDNA sequencing, one of the RNA-Seq methods, is a powerful method used to investigate the transcriptome status of a gene of interest, such as its transcription level and alternative splicing variants. Furthermore, full-length double-strand cDNA sequencing has the advantage that it can create a library from a small amount of sample and the library can be applied to long-read sequencers in addition to short-read sequencers. Nevertheless, one of our previous studies indicated that the full-length double-strand cDNA sequencing yields non-specific genomic DNA amplification, affecting transcriptome analysis, such as transcript quantification and alternative splicing analysis. In this study, it was confirmed that it is possible to produce the RNA-Seq library from only genomic DNA and that the full-length double-strand cDNA sequencing of genomic DNA yielded non-specific genomic DNA amplification. To avoid non-specific genomic DNA amplification, two methods were examined, which are the DNase I-treated full-length double-strand cDNA sequencing and poly(A) capture full-length double-strand cDNA sequencing. Contrary to expectations, the non-specific genomic DNA amplification was increased and the number of the detected expressing genes was reduced in DNase I-treated full-length double-strand cDNA sequencing. On the other hand, in the poly(A) capture full-length double-strand cDNA sequencing, the non-specific genomic DNA amplification was significantly reduced, accordingly the accuracy and the number of detected expressing genes and splicing events were increased. The expression pattern and percentage spliced in index of splicing events were highly correlated. Our results indicate that the poly(A) capture full-length double-strand cDNA sequencing improves transcript quantification accuracy and the detection ability of alternative splicing events. It is also expected to contribute to the determination of the significance of DNA variants to splicing events.
Topics: Alternative Splicing; DNA, Complementary; Deoxyribonuclease I; High-Throughput Nucleotide Sequencing; Poly A; Sequence Analysis, RNA; Transcriptome
PubMed: 35732903
DOI: 10.1038/s41598-022-14902-7 -
Journal of Biological Inorganic... Sep 2023Two arene ruthenium(II) complexes [η-(CH)Ru(pprip)Cl]PF (Ru1; pprip = 2-(3-phenyl-1H-pyrazol-4-yl)-imidazolo[4,5-f][1,10]phenanthroline) and [η-(CH)Ru(HiiP)Cl]PF...
Two arene ruthenium(II) complexes [η-(CH)Ru(pprip)Cl]PF (Ru1; pprip = 2-(3-phenyl-1H-pyrazol-4-yl)-imidazolo[4,5-f][1,10]phenanthroline) and [η-(CH)Ru(HiiP)Cl]PF (Ru2; HiiP = 2-(indole-3-yl)-imidazolo[4,5-f][1,10]phenanthroline) have been synthesized and characterized in this work. Binding properties of Ru1 and Ru2 with the triplex RNA poly(U)•poly(A)*poly(U) were investigated by spectrophotometry and spectrofluorometry as well as viscosimetry. Analysis of spectroscopic titrations and viscosity measurements show that the two complexes bind with the triplex through intercalation, while the binding affinity for Ru2 toward the triplex is stronger than that for Ru1. Melting experiments indicate that the stabilizing effects of Ru1 and Ru2 toward the triplex differ from each other. Under the conditions used herein, Ru1 only stabilizes the Hoogsteen base-paired strand (third strand) without affecting stabilization of the Watson-Crick base-paired strand (the template duplex) of the triplex, while Ru2 stabilizes both the template duplex and the third strand. Although the two complexes prefer to stabilizing the third strand rather than the template duplex, the third-strand stabilization effect of Ru2 is stronger than that of Ru1. The obtained results of this work reveal that the planarity of the intercalative ligands plays an important role in the triplex stabilization by arene Ru(II) complexes.
Topics: Poly A; Ruthenium; Poly U; RNA; Phenanthrolines; Nucleic Acid Conformation; Spectrometry, Fluorescence
PubMed: 37477757
DOI: 10.1007/s00775-023-02008-y -
Biochimie Jun 2021Eukaryotic mRNA deadenylation is generally considered as a two-step process in which the PAN2-PAN3 complex initiates the poly(A) tail degradation while, in the second...
Eukaryotic mRNA deadenylation is generally considered as a two-step process in which the PAN2-PAN3 complex initiates the poly(A) tail degradation while, in the second step, the CCR4-NOT complex completes deadenylation, leading to decapping and degradation of the mRNA body. However, the mechanism of the biphasic poly(A) tail deadenylation remains enigmatic in several points such as the timing of the switch between the two steps, the role of translation termination and the mRNAs population involved. Here, we have studied the deadenylation of endogenous mRNAs in human cells depleted in either PAN3 or translation termination factor eRF3. Among the mRNAs tested, we found that only the endogenous ATF4 mRNA meets the biphasic model for deadenylation and that eRF3 prevents the shortening of its poly(A) tail. For the other mRNAs, the poor effect of PAN3 depletion on their poly(A) tail shortening questions the mode of their deadenylation. It is possible that these mRNAs experience a single step deadenylation process. Alternatively, we propose that a very short initial deadenylation by PAN2-PAN3 is followed by a rapid transition to the second phase involving CCR4-NOT complex. These differences in the timing of the transition from one deadenylation step to the other could explain the difficulties encountered in the generalization of the biphasic deadenylation model.
Topics: Activating Transcription Factor 4; Carrier Proteins; Exoribonucleases; HCT116 Cells; Humans; Peptide Termination Factors; Poly A; RNA Stability; RNA, Messenger
PubMed: 33775689
DOI: 10.1016/j.biochi.2021.03.013 -
The International Journal of... Sep 2014While most long noncoding RNAs (lncRNAs) appear indistinguishable from mRNAs, having 5' cap structures and 3' poly(A) tails, recent work has revealed new formats. Rather... (Review)
Review
While most long noncoding RNAs (lncRNAs) appear indistinguishable from mRNAs, having 5' cap structures and 3' poly(A) tails, recent work has revealed new formats. Rather than taking advantage of the canonical cleavage and polyadenylation for their 3' end maturation, such lncRNAs are processed and stablized by a number of other mechanisms, including the RNase P cleavage to generate a mature 3' end, or capped by snoRNP complexes at both ends, or by forming circular structures. Importantly, such lncRNAs have also been implicated in gene expression regulation in mammalian cells. Here, we highlight recent progress in our understanding of the biogenesis and function of lncRNAs without a poly(A) tail. This paper is part of a directed issue entitled: The Non-coding RNA Revolution.
Topics: Animals; Gene Expression Regulation; Humans; Poly A; RNA 3' End Processing; RNA, Long Noncoding; RNA, Messenger
PubMed: 24513732
DOI: 10.1016/j.biocel.2013.10.009 -
PLoS Computational Biology Apr 2020The mammalian circadian clock is deeply rooted in rhythmic regulation of gene expression. Rhythmic transcriptional control mediated by the circadian transcription...
The mammalian circadian clock is deeply rooted in rhythmic regulation of gene expression. Rhythmic transcriptional control mediated by the circadian transcription factors is thought to be the main driver of mammalian circadian gene expression. However, mounting evidence has demonstrated the importance of rhythmic post-transcriptional controls, and it remains unclear how the transcriptional and post-transcriptional mechanisms collectively control rhythmic gene expression. In mouse liver, hundreds of genes were found to exhibit rhythmicity in poly(A) tail length, and the poly(A) rhythms are strongly correlated with the protein expression rhythms. To understand the role of rhythmic poly(A) regulation in circadian gene expression, we constructed a parsimonious model that depicts rhythmic control imposed upon basic mRNA expression and poly(A) regulation processes, including transcription, deadenylation, polyadenylation, and degradation. The model results reveal the rhythmicity in deadenylation as the strongest contributor to the rhythmicity in poly(A) tail length and the rhythmicity in the abundance of the mRNA subpopulation with long poly(A) tails (a rough proxy for mRNA translatability). In line with this finding, the model further shows that the experimentally observed distinct peak phases in the expression of deadenylases, regardless of other rhythmic controls, can robustly cluster the rhythmic mRNAs by their peak phases in poly(A) tail length and abundance of the long-tailed subpopulation. This provides a potential mechanism to synchronize the phases of target gene expression regulated by the same deadenylases. Our findings highlight the critical role of rhythmic deadenylation in regulating poly(A) rhythms and circadian gene expression.
Topics: Adenine; Animals; Circadian Clocks; Computational Biology; Computer Simulation; Gene Expression Regulation; Liver; Mice; Models, Genetic; Poly A; Polyadenylation; RNA, Messenger
PubMed: 32339166
DOI: 10.1371/journal.pcbi.1007842 -
Chemical Communications (Cambridge,... Mar 2022A series of amphipathic poly-β-peptides are designed for intracellular protein delivery. The poly-β-peptides with higher molecular weight and hydrophobic contents...
A series of amphipathic poly-β-peptides are designed for intracellular protein delivery. The poly-β-peptides with higher molecular weight and hydrophobic contents exhibit higher protein loading and superior delivery efficiency. The lead material efficiently delivers proteins into cells with reserved bioactivity.
Topics: Hydrophobic and Hydrophilic Interactions; Peptides; Poly A; Proteins
PubMed: 35293911
DOI: 10.1039/d2cc00453d -
G3 (Bethesda, Md.) Jun 2019Poly(A)-tail targeted RNAseq approaches, such as 3'READS, PAS-Seq and Poly(A)-ClickSeq, are becoming popular alternatives to random-primed RNAseq to focus sequencing...
Poly(A)-tail targeted RNAseq approaches, such as 3'READS, PAS-Seq and Poly(A)-ClickSeq, are becoming popular alternatives to random-primed RNAseq to focus sequencing reads just to the 3' ends of polyadenylated RNAs to identify poly(A)-sites and characterize changes in their usage. Additionally, we and others have demonstrated that these approaches perform similarly to other RNAseq strategies for differential gene expression analysis, while saving on the volume of sequencing data required and providing a simpler library synthesis strategy. Here, we present DPAC ( ifferential oly( )- lustering); a streamlined pipeline for the preprocessing of poly(A)-tail targeted RNAseq data, mapping of poly(A)-sites, poly(A)-site clustering and annotation, and determination of differential poly(A)-cluster usage using DESeq2. Changes in poly(A)-cluster usage is simultaneously used to report differential gene expression, differential terminal exon usage and alternative polyadenylation (APA).
Topics: Biomarkers; Cluster Analysis; Computational Biology; Exons; Gene Expression Regulation; Gene Knockdown Techniques; HeLa Cells; Humans; Molecular Sequence Annotation; Poly A; Polyadenylation; RNA, Messenger; Sequence Analysis, RNA; Software
PubMed: 31023725
DOI: 10.1534/g3.119.400273