Transcription inhibition induced by modified triple helix-forming oligonucleotides: a quantitative assay for evaluation in cells.

Abstract:

:Oligonucleotides can bind to double-stranded DNA in a sequence-specific manner to form triple helices. Uniformly modified, pyrimidine-rich oligodeoxyribonuclotides containing internucleosidic N3'-P5' phosphoramidate linkages are known to form very stable triplexes with their DNA target. Psoralen-conjugated triple helix-forming oligonucleotides (Pso-TFOs) can additionally be photo-induced to become irreversibly bound to their targeted DNA sequence. Here, we have examined the ability of various 15-mer phosphoramidate TFOs targeted to the HIV-1 polypurine tract (PPT) sequence to prevent transcription elongation in cell cultures; the PPT sequence has been cloned in the transcribed region of a reporter firefly luciferase gene (luc) and transient expression experiments performed. We show that the level of transcription inhibition of the reporter gene in cells perfectly correlates with the amount of covalent triplex at the PPT site. The efficacy of non-covalent triplexes (either omitting the irradiation step with the psoralen conjugate, or using the unsubstituted oligonucleotide) has been studied in our expression system; the oligonucleotides were introduced into living cells by cationic lipid-mediated delivery or directly into the cell nucleus by microinjection. This experimental approach allowed us to evaluate the intrinsic activity of triplexes as transcriptional inhibitors; transcription elongation was inhibited in cells in a sequence-dependent and concentration-dependent manner. This experimental system is convenient for quantitative and fast evaluation of new chemistries of antigene oligonucleotides as inhibitors of gene expression in cells and in vivo.

journal_name

J Mol Biol

authors

Faria M,Wood CD,White MR,Hélène C,Giovannangeli C

doi

10.1006/jmbi.2000.4386

keywords:

subject

Has Abstract

pub_date

2001-02-09 00:00:00

pages

15-24

issue

1

eissn

0022-2836

issn

1089-8638

pii

S0022-2836(00)94386-9

journal_volume

306

pub_type

杂志文章
  • Programming microbes using pulse width modulation of optical signals.

    abstract::Cells transmit and receive information via signalling pathways. A number of studies have revealed that information is encoded in the temporal dynamics of these pathways and has highlighted how pathway architecture can influence the propagation of signals in time and space. The functional properties of pathway architec...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1016/j.jmb.2013.07.036

    authors: Davidson EA,Basu AS,Bayer TS

    更新日期:2013-11-15 00:00:00

  • Dominant mutations affecting muscle structure in Caenorhabditis elegans that map near the actin gene cluster.

    abstract::By examining F1 progeny of mutagenized Caenorhabditis elegans larvae, we recovered several dominant mutations which affect muscle structure. Five of these new mutations resulted in phenotypes unlike the previously recognized unc-54 and unc-15 dominant alleles. Mapping studies placed all five mutations in the same smal...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1016/0022-2836(84)90023-8

    authors: Waterston RH,Hirsh D,Lane TR

    更新日期:1984-12-15 00:00:00

  • Minimum-energy path for a u6 RNA conformational change involving protonation, base-pair rearrangement and base flipping.

    abstract::The U6 RNA internal stem-loop (U6 ISL) is a highly conserved domain of the spliceosome that is important for pre-mRNA splicing. The U6 ISL contains an internal loop that is in equilibrium between two conformations controlled by the protonation state of an adenine (pK(a)=6.5). Lower pH favors formation of a protonated ...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1016/j.jmb.2009.07.003

    authors: Venditti V,Clos L 2nd,Niccolai N,Butcher SE

    更新日期:2009-09-04 00:00:00

  • Structure and ligand binding of carbohydrate-binding module CsCBM6-3 reveals similarities with fucose-specific lectins and "galactose-binding" domains.

    abstract::Carbohydrate-binding polypeptides, including carbohydrate-binding modules (CBMs) from polysaccharidases, and lectins, are widespread in nature. Whilst CBMs are classically considered distinct from lectins, in that they are found appended to polysaccharide-degrading enzymes, this distinction is blurring. The crystal st...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1016/s0022-2836(03)00152-9

    authors: Boraston AB,Notenboom V,Warren RA,Kilburn DG,Rose DR,Davies G

    更新日期:2003-03-28 00:00:00

  • DNA-directed DNA polymerase and strand displacement activity of the reverse transcriptase encoded by the R2 retrotransposon.

    abstract::R2 elements are non-long terminal repeat (non-LTR) retrotransposons with a single open reading-frame encoding reverse transcriptase, DNA endonuclease and nucleic acid-binding domains. The elements are specialized for insertion into the 28 S rRNA genes of many animal phyla. The R2-encoded activities initiate retrotrans...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1016/j.jmb.2007.09.047

    authors: Kurzynska-Kokorniak A,Jamburuthugoda VK,Bibillo A,Eickbush TH

    更新日期:2007-11-23 00:00:00

  • Crystal structure and novel recognition motif of rho ADP-ribosylating C3 exoenzyme from Clostridium botulinum: structural insights for recognition specificity and catalysis.

    abstract::Clostridium botulinum C3 exoenzyme inactivates the small GTP-binding protein family Rho by ADP-ribosylating asparagine 41, which depolymerizes the actin cytoskeleton. C3 thus represents a major family of the bacterial toxins that transfer the ADP-ribose moiety of NAD to specific amino acids in acceptor proteins to mod...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1006/jmbi.2000.4292

    authors: Han S,Arvai AS,Clancy SB,Tainer JA

    更新日期:2001-01-05 00:00:00

  • Membrane Targeting and Insertion of the C-Tail Protein SciP.

    abstract::C-tailed membrane proteins insert into the bilayer post-translationally because the hydrophobic anchor segment leaves the ribosome at the end of translation. Nevertheless, we find here evidence that the targeting of SciP to the membrane of Escherichia coli occurs co-translationally since signal elements in the N-termi...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1016/j.jmb.2016.09.001

    authors: Pross E,Soussoula L,Seitl I,Lupo D,Kuhn A

    更新日期:2016-10-09 00:00:00

  • Effects of chloride ion binding on the photochemical properties of salinibacter sensory rhodopsin I.

    abstract::Microbial organisms utilize light not only as energy sources but also as signals by which rhodopsins (containing retinal as a chromophore) work as photoreceptors. Sensory rhodopsin I (SRI) is a dual photoreceptor that regulates both negative and positive phototaxis in microbial organisms, such as the archaeon Halobact...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1016/j.jmb.2009.06.050

    authors: Suzuki D,Furutani Y,Inoue K,Kikukawa T,Sakai M,Fujii M,Kandori H,Homma M,Sudo Y

    更新日期:2009-09-11 00:00:00

  • Accomodation of S-cis-tamoxifen-N(2)-guanine adduct within a bent and widened DNA minor groove.

    abstract::The non-steroidal anti-estrogen tamoxifen [TAM] has been in clinical use over the last two decades as a potent adjunct chemotherapeutic agent for treatment of breast cancer. It has also been given prophylactically to women with a strong family history of breast cancer. However, tamoxifen treatment has also been associ...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1006/jmbi.2000.4071

    authors: Shimotakahara S,Gorin A,Kolbanovskiy A,Kettani A,Hingerty BE,Amin S,Broyde S,Geacintov N,Patel DJ

    更新日期:2000-09-15 00:00:00

  • Involvement of the C-terminal region of yeast proteinase B inhibitor 2 in its inhibitory action.

    abstract::Yeast proteinase B inhibitor 2 (YIB2), which is composed of 74 amino acid residues, is an unusual serine protease inhibitor, since it lacks disulfide bonds. To identify its reactive site for proteases, we constructed an expression system for a synthetic YIB2 gene and then attempted to change the inhibitory properties ...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1006/jmbi.1998.2498

    authors: Kojima S,Deguchi M,Miura K

    更新日期:1999-02-26 00:00:00

  • DNA curvature in native and modified EcoRI recognition sites and possible influence upon the endonuclease cleavage reaction.

    abstract::The ligation of a decadeoxynucleotide containing the EcoRI recognition site forms a series of multimers which appear to be curved based on observed anomalous gel migration in polyacrylamide gels. The degree of DNA curvature present in the recognition sequence, based upon the observed migration anomaly, can be altered ...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1016/0022-2836(88)90561-x

    authors: Diekmann S,McLaughlin LW

    更新日期:1988-08-20 00:00:00

  • Interaction of E. coli Hsp90 with DnaK Involves the DnaJ Binding Region of DnaK.

    abstract::The 90-kDa heat shock protein (Hsp90) is a widely conserved and ubiquitous molecular chaperone that participates in ATP-dependent protein remodeling in both eukaryotes and prokaryotes. It functions in conjunction with Hsp70 and the Hsp70 cochaperones, an Hsp40 (J-protein) and a nucleotide exchange factor. In Escherich...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1016/j.jmb.2016.12.014

    authors: Kravats AN,Doyle SM,Hoskins JR,Genest O,Doody E,Wickner S

    更新日期:2017-03-24 00:00:00

  • Functional interactions between the subunits of the lactose transporter from Streptococcus thermophilus.

    abstract::Although the quaternary state has been assessed in detail for only a few members of the major facilitator superfamily (MFS), it is clear that multiple oligomeric states are represented within the MFS. One of its members, the lactose transporter LacS from Streptococcus thermophilus assumes a dimeric structure in the me...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1016/j.jmb.2005.04.047

    authors: Geertsma ER,Duurkens RH,Poolman B

    更新日期:2005-07-01 00:00:00

  • Effect of organic co-solvents on the dimer/tetramer equilibrium of human haemoglobin.

    abstract::We have studied the effect of organic co-solvents (monohydric alcohols and formamide) on the dimer-tetramer equilibrium of human haemoglobin by measuring the dependence of oxygen affinity upon haemoglobin concentration over a 100-fold concentration range and analysing the data with a modified Monod-Wyman-Changeux mode...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1016/0022-2836(86)90514-0

    authors: Cupane A,Vitrano E,Cordone L

    更新日期:1986-05-20 00:00:00

  • Reversible and irreversible changes in nucleosome structure along the c-fos and c-myc oncogenes following inhibition of transcription.

    abstract::A new affinity chromatographic procedure for the separation of transcriptionally active nucleosomes has been used to study the changes that take place in chromatin structure along the c-fos and c-myc genes when RNA synthesis is inhibited. Mercury-affinity chromatography separates the sulfhydryl-reactive nucleosomes of...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1016/0022-2836(90)90327-I

    authors: Chen TA,Sterner R,Cozzolino A,Allfrey VG

    更新日期:1990-04-05 00:00:00

  • The three-dimensional structures of two complexes between recombinant MS2 capsids and RNA operator fragments reveal sequence-specific protein-RNA interactions.

    abstract::Crystal structures of two complexes between recombinant MS2 capsids and RNA operator fragments have been determined at 2.7 A resolution. The coat protein of the RNA bacteriophage MS2 is bifunctional; it forms the icosahedral virus shell to protect the viral nucleic acid and it acts as a translational repressor by bind...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1006/jmbi.1997.1144

    authors: Valegârd K,Murray JB,Stonehouse NJ,van den Worm S,Stockley PG,Liljas L

    更新日期:1997-08-01 00:00:00

  • Preliminary crystallographic study of glycosylated recombinant human renin.

    abstract::Single crystals of glycosylated recombinant human renin have been obtained using the hanging-drop vapor diffusion method with polyethylene glycol and sodium chloride as coprecipitants. The crystals belong to the cubic space group P2(1)3 with a = 143.0 A and contain two molecules of renin in the asymmetric unit. A self...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1016/0022-2836(89)90305-7

    authors: Lim LW,Stegeman RA,Leimgruber NK,Gierse JK,Abdel-Meguid SS

    更新日期:1989-11-05 00:00:00

  • A unique FGF23 with the ability to activate FGFR signaling through both αKlotho and βKlotho.

    abstract::Three fibroblast growth factor (FGF) molecules, FGF19, FGF21, and FGF23, form a unique subfamily that functions as endocrine hormones. FGF19 and FGF21 can regulate glucose, lipid, and energy metabolism, while FGF23 regulates phosphate homeostasis. The FGF receptors and co-receptors for these three FGF molecules have b...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1016/j.jmb.2012.02.027

    authors: Wu X,Weiszmann J,Ge H,Baribault H,Stevens J,Hawkins N,Vonderfecht S,Gardner J,Gupte J,Sheng J,Wang M,Li Y

    更新日期:2012-04-20 00:00:00

  • INT131: a selective modulator of PPAR gamma.

    abstract::The nuclear hormone receptor peroxisome proliferator-activated receptor gamma (PPAR gamma; NR1C3) plays a central role in adipogenesis and is the molecular target of the thiazolidinedione class of antidiabetic drugs. To overcome the well-known shortcomings of thiazolidinediones, we have identified INT131 (formerly T13...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1016/j.jmb.2009.01.025

    authors: Motani A,Wang Z,Weiszmann J,McGee LR,Lee G,Liu Q,Staunton J,Fang Z,Fuentes H,Lindstrom M,Liu J,Biermann DH,Jaen J,Walker NP,Learned RM,Chen JL,Li Y

    更新日期:2009-03-13 00:00:00

  • A pentameric form of human serum amyloid P component. Crystallization, X-ray diffraction and neutron scattering studies.

    abstract::Human serum amyloid P component crystallizes from sodium acetate buffer (pH 5.5) in the presence of calcium and polyethylene glycol 6000, at 4 degrees C. The space group is P2(1) and the cell parameters are a = 69.0 A (1 A = 0.1 nm), b = 99.3 A, c = 96.8 A, beta = 96.1. Density considerations supported by neutron scat...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1016/0022-2836(88)90529-3

    authors: Wood SP,Oliva G,O'Hara BP,White HE,Blundell TL,Perkins SJ,Sardharwalla I,Pepys MB

    更新日期:1988-07-05 00:00:00

  • Specific mode of interaction between components of model pulmonary surfactants using computer simulations.

    abstract::Atomistic molecular dynamics simulations and structural bioinformatics tools enable the identification of the exact mode of interaction between model pulmonary surfactant components. Two nanosecond long simulations of the N-terminal region of human surfactant protein-B (SP-B(1-25)) in dipalmitoylphosphatidylcholine (D...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1016/s0022-2836(02)00774-x

    authors: Kaznessis YN,Kim S,Larson RG

    更新日期:2002-09-20 00:00:00

  • Functional interactions among the His40, Glu58 and His92 catalysts of ribonuclease T1 as studied by double and triple mutants.

    abstract::We report on the functional interplay between the His40, Glu58 and His92 catalysts of ribonuclease T1. The kinetic properties of the single His40Ala, Glu58Ala and His92Gln mutants have been compared with those of the corresponding double and triple mutants. When His40, Glu58 and His92 are mutated separately or togethe...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1006/jmbi.1993.1078

    authors: Steyaert J,Wyns L

    更新日期:1993-02-05 00:00:00

  • Ensemble Modeling and Intracellular Aggregation of an Engineered Immunoglobulin-Like Domain.

    abstract::The production of recombinant proteins in Escherichia coli frequently results in the formation of insoluble protein aggregates called inclusion bodies (IBs). The determinants of IB formation remain poorly understood and are of much interest for biotechnological and research applications, as well as offering insight in...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1016/j.jmb.2016.02.016

    authors: Trainor K,Gingras Z,Shillingford C,Malakian H,Gosselin M,Lipovšek D,Meiering EM

    更新日期:2016-03-27 00:00:00

  • Constraining enzyme conformational change by an antibody leads to hyperbolic inhibition.

    abstract::Although it has been known for many years that antibodies display properties characteristic of allosteric effectors, the molecular mechanisms responsible for these effects remain poorly understood. Here, we describe a single-domain antibody fragment (nanobody) that modulates protein function by constraining conformati...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1016/j.jmb.2011.01.017

    authors: Oyen D,Srinivasan V,Steyaert J,Barlow JN

    更新日期:2011-03-18 00:00:00

  • A ligand-centric analysis of the diversity and evolution of protein-ligand relationships in E.coli.

    abstract::As enzymes evolve and diverge from common ancestor sequences, they often keep their overall reaction chemistry but specialize in the binding of different cognate ligands. This study borrows methods for the computational assessment of 2D similarity of small molecules from the field of chemoinformatics, to examine the e...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1016/j.jmb.2005.01.061

    authors: Nobeli I,Spriggs RV,George RA,Thornton JM

    更新日期:2005-03-25 00:00:00

  • Accommodation of amino acid insertions in an alpha-helix of T4 lysozyme. Structural and thermodynamic analysis.

    abstract::One to four alanines were inserted by site-directed mutagenesis at three different locations within the alpha-helix comprising residues 39 to 50 in bacteriophage T4 lysozyme. All insertion mutants were correctly folded and catalytically active although the insertions led to a thermal destabilization by 1.1 to 4.2 kcal...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1006/jmbi.1994.1195

    authors: Heinz DW,Baase WA,Zhang XJ,Blaber M,Dahlquist FW,Matthews BW

    更新日期:1994-02-25 00:00:00

  • Single amino acid changes in the Bacillus thuringiensis var. israelensis delta-endotoxin affect the toxicity and expression of the protein.

    abstract::Site-directed mutagenesis has been used to change individual amino acids of the larvicidal 27,000 Mr delta-endotoxin of Bacillus thuringiensis var. israelensis. Basic and acidic residues have been systematically replaced by alanine, and the resulting mutant polypeptides analysed for cytolytic and larvicidal activity, ...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1016/0022-2836(88)90283-5

    authors: Ward ES,Ellar DJ,Chilcott CN

    更新日期:1988-08-05 00:00:00

  • Cooperative unfolding of a metastable serpin to a molten globule suggests a link between functional and folding energy landscapes.

    abstract::Alpha-1 antitrypsin (alpha(1)-AT) is a member of the serpin class of protease inhibitors, and folds to a metastable state rather than its thermodynamically most stable native state. Upon cleavage by a target protease, alpha(1)-AT undergoes a dramatic conformational change to a stable form, translocating the bound prot...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1016/j.jmb.2007.05.039

    authors: Tsutsui Y,Wintrode PL

    更新日期:2007-08-03 00:00:00

  • An on-pathway hidden intermediate and the early rate-limiting transition state of Rd-apocytochrome b562 characterized by protein engineering.

    abstract::The folding pathway of Rd-apocytochrome b562, a four-helix bundle protein, was characterized using Trp and Ala/Gly pair mutations. We found that the Trp mutants (F65W) of both the fully folded Rd-apocytochrome b562 and a partially unfolded intermediate with the N-terminal helix (helix I) unfolded, fold with identical ...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1016/j.jmb.2005.07.057

    authors: Zhou Z,Huang Y,Bai Y

    更新日期:2005-09-30 00:00:00

  • RNase E and polyadenyl polymerase I are involved in maturation of CI RNA, the P4 phage immunity factor.

    abstract::Bacteriophage P4 immunity is controlled by a small stable RNA (CI RNA) that derives from the processing of primary transcripts. In previous works, we observed that the endonuclease RNase P is required for the maturation of CI RNA 5'-end; moreover, we found that polynucleotide phosphorylase (PNPase), a 3' to 5' RNA-deg...

    journal_title:Journal of molecular biology

    pub_type: 杂志文章

    doi:10.1016/S0022-2836(02)00085-2

    authors: Briani F,Del Vecchio E,Migliorini D,Hajnsdorf E,Régnier P,Ghisotti D,Dehò G

    更新日期:2002-04-26 00:00:00