Characteristic effect of hydroxyurea on the higher-order structure of DNA and gene expression.

Hydroxyurea (HU; hydroxycarbamide) is a chemotherapy medication used to treat various types of cancer and other diseases such as sickle cell anemia. HU inhibits DNA synthesis by targeting ribonucleotide reductase (RNR). Recent studies have suggested that HU also causes oxidative stress in living systems.

The Anticancer Drug Daunomycin Directly Affects Gene Expression and DNA Structure.

Daunomycin (DM), an anthracycline antibiotic, is frequently used to treat various cancers, but the direct effects of DM on gene expression and DNA structure are unclear. We used an in vitro cell-free system, optimized with spermine (SP), to study the effect of DM on gene expression. A bimodal effect of DM on gene expression, weak promotion followed by inhibition, was observed with increasing concentration of DM.

Activation/Inhibition of Gene Expression Caused by Alcohols: Relationship with the Viscoelastic Property of a DNA Molecule.

Alcohols are used in the life sciences because they can condense and precipitate DNA. Alcohol consumption has been linked to many diseases and can alter genetic activity. In the present report, we carried out experiments to make clear how alcohols affect the efficiency of transcription-translation (TX-TL) and translation (TL) by adapting cell-free gene expression systems with plasmid DNA and RNA templates, respectively.

Damages of DNA in tritiated water.

Tritium is a radioisotope of hydrogen emitting low energy β-rays in disintegration to He. DNA molecules are damaged mainly by β-ray irradiation, and additional damages can be induced by break of chemical bond by nuclear transmutation to inert He. Deep knowledges of the mechanisms underlying DNA damages lead to better understanding of biological effects of tritium.

Markedly Different Effects of Monovalent Cations on the Efficiency of Gene Expression.

The effect of monovalent cations on a cell-free transcription-translation (TX-TL) system is examined using a luciferase assay. It is found that the potency for all ions analyzed here is in the order Rb  > K  > Cs  > Na  ≈ Li  > (CH ) N , where Rb is most efficient at promoting TX-TL and the ions of Li , Na , and (CH ) N exhibit an inhibitory effect. Similar promotion/inhibition effects are observed for cell-free TL alone with an mRNA template.

Effects of Structural Isomers of Spermine on the Higher-Order Structure of DNA and Gene Expression.

Polyamines are involved in various biological functions, including cell proliferation, differentiation, gene regulation, etc. Recently, it was found that polyamines exhibit biphasic effects on gene expression: promotion and inhibition at low and high concentrations, respectively. Here, we compared the effects of three naturally occurring tetravalent polyamines, spermine (SPM), thermospermine (TSPM), and -aminopropylspermidine (BSPD).

Construction of 3D Cellular Composites with Stem Cells Derived from Adipose Tissue and Endothelial Cells by Use of Optical Tweezers in a Natural Polymer Solution.

To better understand the regulation and function of cellular interactions, three-dimensional (3D) assemblies of single cells and subsequent functional analysis are gaining popularity in many research fields. While we have developed strategies to build stable cellular structures using optical tweezers in a minimally invasive state, methods for manipulating a wide range of cell types have yet to be established. To mimic organ-like structures, the construction of 3D cellular assemblies with variety of cell types is essential.

Manipulating Living Cells to Construct Stable 3D Cellular Assembly Without Artificial Scaffold.

Regenerative medicine and tissue engineering offer several advantages for the treatment of intractable diseases, and several studies have demonstrated the importance of 3-dimensional (3D) cellular assemblies in these fields. Artificial scaffolds have often been used to construct 3D cellular assemblies. However, the scaffolds used to construct cellular assemblies are sometimes toxic and may change the properties of the cells.

Optical Fluid Pump: Generation of Directional Flow via Microphase Segregation/Homogenization.

We report the successful generation of directional liquid-flow under stationary laser irradiation at a fixed position in a chamber. We adopt a homogeneous solution consisting of a mixture of water and triethylamine (TEA), with a composition near the critical point for phase segregation. When geometrical asymmetry is introduced around the laser focus in the chamber, continuous directional flow is generated, accompanied by the emergence of water-rich microdroplets at the laser focus.

Cracking pattern of tissue slices induced by external extension provides useful diagnostic information.

Although biopsy is one of the most important methods for diagnosis in diseases, there is ambiguity based on the information obtained from the visual inspection of tissue slices. Here, we studied the effect of external extension on tissue slices from mouse liver with different stages of disease: Healthy normal state, Simple steatosis, Non-alcoholic steatohepatitis and Hepatocellular carcinoma. We found that the cracking pattern of a tissue slice caused by extension can provide useful information for distinguishing among the disease states.

Opposite effect of polyamines on In vitro gene expression: Enhancement at low concentrations but inhibition at high concentrations.

Background: Polyamines have various biological functions including marked effects on the structure and function of genomic DNA molecules. Changes in the higher-order structure of DNA caused by polyamines are expected to be closely related to genetic activity. To clarify this issue, we examined the relationship between gene expression and the higher-order structure of DNA under different polyamine concentrations.

Manipulating Living Cells to Construct a 3D Single-Cell Assembly without an Artificial Scaffold.

Artificial scaffolds such as synthetic gels or chemically-modified glass surfaces that have often been used to achieve cell adhesion are xenobiotic and may harm cells. To enhance the value of cell studies in the fields of regenerative medicine and tissue engineering, it is becoming increasingly important to create a cell-friendly technique to promote cell⁻cell contact. In the present study, we developed a novel method for constructing stable cellular assemblies by using optical tweezers in a solution of a natural hydrophilic polymer, dextran.

Double-Strand Breaks in Genome-Sized DNA Caused by Ultrasound.

DNA double-strand breaks (DSBs) caused by ultrasound were evaluated in a quantitative manner by single-molecule fluorescence microscopy. We compared the effect of time-interval (or pulse) sonication to that of continuous wave (CW) sonication at a fixed frequency of 30 kHz. Pulses caused fewer DSBs than CW sonication under the same total input ultrasound energy when the pulse repetition period was above the order of a second.

Divalent cation shrinks DNA but inhibits its compaction with trivalent cation.

Our observation reveals the effects of divalent and trivalent cations on the higher-order structure of giant DNA (T4 DNA 166 kbp) by fluorescence microscopy. It was found that divalent cations, Mg(2+) and Ca(2+), inhibit DNA compaction induced by a trivalent cation, spermidine (SPD(3+)). On the other hand, in the absence of SPD(3+), divalent cations cause the shrinkage of DNA.

Highly Concentrated Ethanol Solutions: Good Solvents for DNA as Revealed by Single-Molecule Observation.

We observed single DNA molecules at different ethanol concentrations by using fluorescence microscopy. Large single DNA molecules undergo reentrant conformational transitions from elongated coil into folded globule and then into elongated coil state, accompanied by the increase of the concentration of ethanol in a low-salt aqueous environment. The second transition from globule into the coil state occurs at around 70 % (v/v) ethanol.

Marked difference in conformational fluctuation between giant DNA molecules in circular and linear forms.

We performed monomolecular observations on linear and circular giant DNAs (208 kbp) in an aqueous solution by the use of fluorescence microscopy. The results showed that the degree of conformational fluctuation in circular DNA was ca. 40% less than that in linear DNA, although the long-axis length of circular DNA was only 10% smaller than that of linear DNA.

Structural change of DNA induced by nucleoid proteins: growth phase-specific Fis and stationary phase-specific Dps.

The effects of nucleoid proteins Fis and Dps of Escherichia coli on the higher order structure of a giant DNA were studied, in which Fis and Dps are known to be expressed mainly in the exponential growth phase and stationary phase, respectively. Fis causes loose shrinking of the higher order structure of a genome-sized DNA, T4 DNA (166 kbp), in a cooperative manner, that is, the DNA conformational transition proceeds through the appearance of a bimodal size distribution or the coexistence of elongated coil and shrunken globular states. The effective volume of the loosely shrunken state induced by Fis is 30-60 times larger than that of the compact state induced by spermidine, suggesting that cellular enzymes can access for DNA with the shrunken state but cannot for the compact state.