scholarly journals Oligonucleotides DNA containing 8-trifluoromethyl-2′-deoxyguanosine for observing Z-DNA structure

2020 ◽  
Author(s):  
Hong-Liang Bao ◽  
Tatsuki Masuzawa ◽  
Takanori Oyoshi ◽  
Yan Xu
Keyword(s):  
Genetic Diseases ◽  
Dna Structure ◽  
2D Nmr ◽  
Living Cells ◽  
Molecular Probes ◽  
Alternative Form ◽  
Left Handed ◽  
Z Dna ◽  
And Function

Abstract Z-DNA is known to be a left-handed alternative form of DNA and has important biological roles as well as being related to cancer and other genetic diseases. It is therefore important to investigate Z-DNA structure and related biological events in living cells. However, the development of molecular probes for the observation of Z-DNA structures inside living cells has not yet been realized. Here, we have succeeded in developing site-specific trifluoromethyl oligonucleotide DNA by incorporation of 8-trifluoromethyl-2′-deoxyguanosine (FG). 2D NMR strongly suggested that FG adopted a syn conformation. Trifluoromethyl oligonucleotides dramatically stabilized Z-DNA, even under physiological salt concentrations. Furthermore, the trifluoromethyl DNA can be used to directly observe Z-form DNA structure and interaction of DNA with proteins in vitro, as well as in living human cells by19F NMR spectroscopy for the first time. These results provide valuable information to allow understanding of the structure and function of Z-DNA.

scholarly journals Characterization of Flowering Genes of Arabidopsis thaliana for Mirror Repeats

2021 ◽  
Vol 12 (3) ◽  
pp. 2852-2861
Keyword(s):  
Arabidopsis Thaliana ◽  
Genetic Diseases ◽  
Dna Repeats ◽  
Quick Method ◽  
Flowering Genes ◽  
Repeat Sequences ◽  
Mutational Hotspots ◽  
And Function

A variety of simple DNA repeats are enriched in the eukaryotic genomes. Recent studies have proven their importance in understanding genome organization and function, especially how genomes evolve using them as mutational hotspots during DNA replication. Mirror repeat sequences, the most underrated subset of this class of repeats, are now gaining importance because of their probable involvement in developing several genetic diseases in humans. These repeats typically adopt H-DNA conformations in both in-vitro and in-vivo conditions. On the other end, plants were still not analyzed for their presence or distribution and whether they are responsible for causing diseases in them or not. The present study aims to extract mirror repeats in the flowering genes of Arabidopsis thaliana. To this end, we have deployed FPCB (FASTA-PARALLEL COMPLEMENT-BLAST), an efficacious and quick method to extract perfect and degenerate mirror repeat sequences through pattern matching of alignments with user-defined algorithmic parameters. All the analyzed genes were reported to have quite high densities of mirror sequences. A total of 93 unique mirror repeats of significant lengths were extracted in the analyzed genes.

Localization of Z-RNA in Normal Lens Epithelium: Middle Fibers

2000 ◽  
Vol 6 (S2) ◽  
pp. 900-901
Author(s):  
C.E. Gagna ◽  
H.R. Kuo ◽  
R. Schulz ◽  
R. Cordova ◽  
G. Crippen ◽  
...  
Keyword(s):  
Dna Sequences ◽  
Cellular Localization ◽  
Cultured Cells ◽  
Polyclonal Antibodies ◽  
Image Analyzer ◽  
Left Handed ◽  
Z Dna ◽  
Normal Lens

The goal of this project was to analyze the cellular localization of Z-RNA, within middle fibers (MF) of the adult dog ocular lens (1.5 yr) (Fig. 1), using anti-Z-RNA IgG polyclonal antibodies. B-DNA can adopt the left-handed Z-DNA conformation in vitro (1). Right-handed A-RNA can be transformed into left-handed Z-RNA (2). Z-RNA has been studied in cultured cells (3). Evidence supports the presence of Z-DNA in vivo (1). Removal of DNA binding proteins by fixatives can initiate supercoiling which stabilizes Z-DNA sequences (1).Anti-Z-RNA polyclonal antibody probes were developed in rabbits immunized with multiple injections of Z-RNA: Br-poly[ribosomal(G-C)]. Regarding immunohistochemistry, lens tissues were fixed in Carnoy's, embedded in paraffin and sectioned (2.5 μm) (Fig. 2). Computerized image analysis was performed using a Leitz DM-RB microscope and Leica Quantiment 500 + image analyzer.

Disruption of Golgi structure and function in mammalian cells expressing a mutant dynamin

2000 ◽  
Vol 113 (11) ◽  
pp. 1993-2002 ◽  
Author(s):  
H. Cao ◽  
H.M. Thompson ◽  
E.W. Krueger ◽  
M.A. McNiven
Keyword(s):  
Mammalian Cells ◽  
Living Cells ◽  
Dominant Negative ◽  
Coated Vesicle ◽  
Large Numbers ◽  
Golgi Structure ◽  
And Function ◽  
Golgi Network ◽  
Mutant Cells

The large GTPase dynamin is a mechanoenzyme that participates in the scission of nascent vesicles from the plasma membrane. Recently, dynamin has been demonstrated to associate with the Golgi apparatus in mammalian cells by morphological and biochemical methods. Additional studies using a well characterized, cell-free assay have supported these findings by demonstrating a requirement for dynamin function in the formation of clathrin-coated, and non-clathrin-coated vesicles from the trans-Golgi network (TGN). In this study, we tested if dynamin participates in Golgi function in living cells through the expression of a dominant negative dynamin construct (K44A). Cells co-transfected to express this mutant dynamin and a GFP-tagged Golgi resident protein (TGN38) exhibit Golgi structures that are either compacted, vesiculated, or tubulated. Electron microscopy of these mutant cells revealed large numbers of Golgi stacks comprised of highly tubulated cisternae and an extraordinary number of coated vesicle buds. Cells expressing mutant dynamin and GFP-tagged VSVG demonstrated a marked retention (8- to 11-fold) of the nascent viral G-protein in the Golgi compared to control cells. These observations in living cells are consistent with previous morphological and in vitro studies demonstrating a role for dynamin in the formation of secretory vesicles from the TGN.

CGG repeats associated with fragile X chromosome form left-handed Z-DNA structure

Biopolymers ◽  
2010 ◽  
Vol 95 (3) ◽  
pp. 174-181 ◽  
Author(s):  
Daniel Renčiuk ◽  
Jaroslav Kypr ◽  
Michaela Vorlíčková
Keyword(s):  
X Chromosome ◽  
Fragile X ◽  
Dna Structure ◽  
Chromosome Form ◽  
Left Handed ◽  
Cgg Repeats ◽  
Z Dna

The Z-DNA motif d(TG)30 promotes reception of information during gene conversion events while stimulating homologous recombination in human cells in culture

1990 ◽  
Vol 10 (2) ◽  
pp. 785-793
Author(s):  
W P Wahls ◽  
L J Wallace ◽  
P D Moore
Keyword(s):  
Homologous Recombination ◽  
Gene Conversion ◽  
Dna Sequences ◽  
Human Cells ◽  
Base Pairs ◽  
Cells In Culture ◽  
Left Handed ◽  
Dna Motif ◽  
Z Dna

Tracts of the alternating dinucleotide polydeoxythymidylic-guanylic [d(TG)].polydeoxyadenylic-cytidylic acid [d(AC)], present throughout the human genome, are capable of readily forming left-handed Z-DNA in vitro. We have analyzed the effects of the Z-DNA motif d(TG)30 upon homologous recombination between two nonreplicating plasmid substrates cotransfected into human cells in culture. In this study, the sequence d(TG)30 is shown to stimulate homologous recombination up to 20-fold. Enhancement is specific to the Z-DNA motif; a control DNA fragment of similar size does not alter the recombination frequency. The stimulation of recombination is observed at a distance (237 to 1,269 base pairs away from the Z-DNA motif) and involves both gene conversion and reciprocal exchange events. Maximum stimulation is observed when the sequence is present in both substrates, but it is capable of stimulating when present in only one substrate. Analysis of recombination products indicates that the Z-DNA motif increases the frequency and alters the distribution of multiple, unselected recombination events. Specifically designed crosses indicate that the substrate containing the Z-DNA motif preferentially acts as the recipient of genetic information during gene conversion events. Models describing how left-handed Z-DNA sequences might promote the initiation of homologous recombination are presented.

scholarly journals The Z-DNA motif d(TG)30 promotes reception of information during gene conversion events while stimulating homologous recombination in human cells in culture.

1990 ◽  
Vol 10 (2) ◽  
pp. 785-793 ◽  
Author(s):  
W P Wahls ◽  
L J Wallace ◽  
P D Moore
Keyword(s):  
Homologous Recombination ◽  
Gene Conversion ◽  
Dna Sequences ◽  
Human Cells ◽  
Base Pairs ◽  
Cells In Culture ◽  
Left Handed ◽  
Dna Motif ◽  
Z Dna

Tracts of the alternating dinucleotide polydeoxythymidylic-guanylic [d(TG)].polydeoxyadenylic-cytidylic acid [d(AC)], present throughout the human genome, are capable of readily forming left-handed Z-DNA in vitro. We have analyzed the effects of the Z-DNA motif d(TG)30 upon homologous recombination between two nonreplicating plasmid substrates cotransfected into human cells in culture. In this study, the sequence d(TG)30 is shown to stimulate homologous recombination up to 20-fold. Enhancement is specific to the Z-DNA motif; a control DNA fragment of similar size does not alter the recombination frequency. The stimulation of recombination is observed at a distance (237 to 1,269 base pairs away from the Z-DNA motif) and involves both gene conversion and reciprocal exchange events. Maximum stimulation is observed when the sequence is present in both substrates, but it is capable of stimulating when present in only one substrate. Analysis of recombination products indicates that the Z-DNA motif increases the frequency and alters the distribution of multiple, unselected recombination events. Specifically designed crosses indicate that the substrate containing the Z-DNA motif preferentially acts as the recipient of genetic information during gene conversion events. Models describing how left-handed Z-DNA sequences might promote the initiation of homologous recombination are presented.

scholarly journals Structure and Function of the Saccharomyces cerevisiae Sir3 BAH Domain

2006 ◽  
Vol 26 (8) ◽  
pp. 3256-3265 ◽  
Author(s):  
Jessica J. Connelly ◽  
Peihua Yuan ◽  
Hao-Chi Hsu ◽  
Zhizhong Li ◽  
Rui-Ming Xu ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Protein A ◽  
Transcriptional Silencing ◽  
Structure And Function ◽  
Full Length ◽  
Left Handed ◽  
N Terminus ◽  
Bah Domain ◽  
And Function

ABSTRACT Previous work has shown that the N terminus of the Saccharomyces cerevisiae Sir3 protein is crucial for the function of Sir3 in transcriptional silencing. Here, we show that overexpression of N-terminal fragments of Sir3 in strains lacking the full-length protein can lead to some silencing of HML and HMR. Sir3 contains a BAH (bromo-adjacent homology) domain at its N terminus. Overexpression of this domain alone can lead to silencing as long as Sir1 is overexpressed and Sir2 and Sir4 are present. Overexpression of the closely related Orc1 BAH domain can also silence in the absence of any Sir3 protein. A previously characterized hypermorphic sir3 mutation, D205N, greatly improves silencing by the Sir3 BAH domain and allows it to bind to DNA and oligonucleosomes in vitro. A previously uncharacterized region in the Sir1 N terminus is required for silencing by both the Sir3 and Orc1 BAH domains. The structure of the Sir3 BAH domain has been determined. In the crystal, the molecule multimerizes in the form of a left-handed superhelix. This superhelix may be relevant to the function of the BAH domain of Sir3 in silencing.

scholarly journals The role of the metabolite cargo of extracellular vesicles in tumor progression

2021 ◽  
Author(s):  
Mária Harmati ◽  
Mátyás Bukva ◽  
Tímea Böröczky ◽  
Krisztina Buzás ◽  
Edina Gyukity-Sebestyén
Keyword(s):  
Tumor Progression ◽  
Extracellular Vesicles ◽  
Biomarker Discovery ◽  
Tumor Development ◽  
Living Cells ◽  
Pathological Conditions ◽  
Membrane Bound ◽  
And Function

AbstractMetabolomic reprogramming in tumor and stroma cells is a hallmark of cancer but understanding its effects on the metabolite composition and function of tumor-derived extracellular vesicles (EVs) is still in its infancy. EVs are membrane-bound sacs with a complex molecular composition secreted by all living cells. They are key mediators of intercellular communication both in normal and pathological conditions and play a crucial role in tumor development. Although lipids are major components of EVs, most of the EV cargo studies have targeted proteins and nucleic acids. The potential of the EV metabolome as a source for biomarker discovery has gained recognition recently, but knowledge on the biological activity of tumor EV metabolites still remains limited. Therefore, we aimed (i) to compile the list of metabolites identified in tumor EVs isolated from either clinical specimens or in vitro samples and (ii) describe their role in tumor progression through literature search and pathway analysis.

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