Simple Elastic Network Models for Exhaustive Analysis of Long Double-Stranded DNA Dynamics with Sequence Geometry Dependence

Simple elastic network models of DNA were developed to reveal the structure-dynamics relationships for several nucleotide sequences. First, we propose a simple all-atom elastic network model of DNA that can explain the profiles of temperature factors for several crystal structures of DNA. Second, we propose a coarse-grained elastic network model of DNA, where each nucleotide is described only by one node. This model could effectively reproduce the detailed dynamics obtained with the all-atom elastic network model according to the sequence-dependent geometry. Through normal-mode analysis for the coarse-grained elastic network model, we exhaustively analyzed the dynamic features of a large number of long DNA sequences, approximately ∼150 bp in length. These analyses revealed positive correlations between the nucleosome-forming abilities and the inter-strand fluctuation strength of double-stranded DNA for several DNA sequences.     

Impact of symptoms by gender and age in Japanese subjects with irritable bowel syndrome with constipation (IBS-C): a large population-based internet survey

Background

Irritable bowel syndrome with constipation (IBS-C) is a representative psychosomatic disorder. Several pathophysiological factors have been linked to IBS symptoms such as the modulation of gastrointestinal motility, visceral hypersensitivity, dysregulation of the gut-brain axis, genetic and environmental factors, sequelae of infection, and psychosocial disorders. It is likely that biopsychosocial aspects of IBS-C underlie its gender and age effects. However, the influence of each symptom of IBS-C by gender and age is not well understood. We hypothesized that the expression rate of each IBS-C symptom in females and in subjects aged 20–49 years was higher than that of subjects who were male and aged 50–79 years.

Methods

We conducted an internet survey of 30,000 adults from the general Japanese population. IBS-C subjects were asked to answer a questionnaire on the degree of anxiety, thoughts about bowel habits, and their dominant gastrointestinal symptoms together with exacerbation factors. The correlation between gender and age and IBS-C symptoms was analyzed.

Results

When analyzed by gender, the expression rate of abdominal discomfort, abdominal distention, and abdominal fullness was significantly higher in female than male IBS-C subjects (66.5% vs. 58.7%, p?<?0.05; 54.7% vs. 43.6%, p?<?0.01; 18.9% vs. 9.6%, p?<?0.01, respectively). When analyzed by age, the expression rate of abdominal distention and abdominal pain was significantly higher among IBS-C subjects aged 20–49 years than those aged 50–79 years (55.7% vs. 46.8%, p?<?0.05; 36.6% vs. 20.6%, p?<?0.001, respectively). In contrast, there was no gender or age differences with regard to the most common and bothersome symptom (abdominal bloating) among IBS-C subjects.

Conclusions

The expression rate of some IBS-C symptoms was higher among females and those aged 20–49 years than males and those aged 50–79 years, respectively. It is important to understand the impact of symptoms by gender and age to evaluate the pathology of IBS-C from a biopsychosocial perspective.

Trial registration

Although this survey was an anonymous internet survey, we obtained informed consent for the study as an online response. The disclosure of this study was approved by the Ethics Committee of Tohoku University Graduate School of Medicine (approval number: 2015–1-405).

     

Time course of gene expression during Porphyromonas gingivalis strain ATCC 33277 biofilm formation

Chronological gene expression patterns of biofilm-forming cells are important to understand bioactivity and pathogenicity of biofilms. For Porphyromonas gingivalis ATCC 33277 biofilm formation, the number of genes differentially regulated by more than 1.5-fold was highest during the growth stage (312/2,090 genes), and some pathogen-associated genes were time-dependently controlled.     

Peptidyl-prolyl cis/trans isomerase NIMA-interacting 1 associates with insulin receptor substrate-1 and enhances insulin actions and adipogenesis

Peptidyl-prolyl cis/trans isomerase NIMA-interacting 1 (Pin1) is a unique enzyme that associates with the pSer/Thr-Pro motif and catalyzes cis-trans isomerization. We identified Pin1 in the immunoprecipitates of overexpressed IRS-1 with myc and FLAG tags in mouse livers and confirmed the association between IRS-1 and Pin1 by not only overexpression experiments but also endogenously in the mouse liver. The analysis using deletion- and point-mutated Pin1 and IRS-1 constructs revealed the WW domain located in the N terminus of Pin1 and Ser-434 in the SAIN (Shc and IRS-1 NPXY binding) domain of IRS-1 to be involved in their association. Subsequently, we investigated the role of Pin1 in IRS-1 mediation of insulin signaling. The overexpression of Pin1 in HepG2 cells markedly enhanced insulin-induced IRS-1 phosphorylation and its downstream events: phosphatidylinositol 3-kinase binding with IRS-1 and Akt phosphorylation. In contrast, the treatment of HepG2 cells with Pin1 siRNA or the Pin1 inhibitor Juglone suppressed these events. In good agreement with these in vitro data, Pin1 knock-out mice exhibited impaired insulin signaling with glucose intolerance, whereas adenoviral gene transfer of Pin1 into the ob/ob mouse liver mostly normalized insulin signaling and restored glucose tolerance. In addition, it was also demonstrated that Pin1 plays a critical role in adipose differentiation, making Pin1 knock-out mice resistant to diet-induced obesity. Importantly, Pin1 expression was shown to be up-regulated in accordance with nutrient conditions such as food intake or a high-fat diet. Taken together, these observations indicate that Pin1 binds to IRS-1 and thereby markedly enhances insulin action, essential for adipogenesis.     

The role of metals in site-specific DNA damage with reference to carcinogenesis

We reviewed the mechanism of oxidative DNA damage with reference to metal carcinogenesis and metal-mediated chemical carcinogenesis. On the basis of the finding that chromium (VI) induced oxidative DNA damage in the presence of hydrogen peroxide (H2O2), we proposed the hypothesis that endogenous reactive oxygen species play a role in metal carcinogenesis. Since then, we have reported that various metal compounds, such as cobalt, nickel, and ferric nitrilotriacetate, directly cause site-specific DNA damage in the presence of H2O2. We also found that carcinogenic metals could cause DNA damage through indirect mechanisms. Certain nickel compounds induced oxidative DNA damage in rat lungs through inflammation. Endogenous metals, copper and iron, catalyzed ROS generation from various organic carcinogens, resulting in oxidative DNA damage. Polynuclear compounds, such as 4-aminobiphenyl and heterocyclic amines, appear to induce cancer mainly through DNA adduct formation, although their N-hydroxy and nitroso metabolites can also cause oxidative DNA damage. On the other hand, mononuclear compounds, such as benzene metabolites, caffeic acid, and o-toluidine, should express their carcionogenicity through oxidative DNA damage. Metabolites of certain carcinogens efficiently caused oxidative DNA damage by forming NADH-dependent redox cycles. These findings suggest that metal-mediated oxidative DNA damage plays important roles in chemical carcinogenesis.     

Detection of MPLW515L/K Mutations and Determination of Allele Frequencies with a Single-Tube PCR Assay

A gain-of-function mutation in the myeloproliferative leukemia virus (MPL) gene, which encodes the thrombopoietin receptor, has been identified in patients with essential thrombocythemia and primary myelofibrosis, subgroups of classic myeloproliferative neoplasms (MPNs). The presence of MPL gene mutations is a critical diagnostic criterion for these diseases. Here, we developed a rapid, simple, and cost-effective method of detecting two major MPL mutations, MPLW515L/K, in a single PCR assay; we termed this method DARMS (dual amplification refractory mutation system)-PCR. DARMS-PCR is designed to produce three different PCR products corresponding to MPLW515L, MPLW515K, and all MPL alleles. The amplicons are later detected and quantified using a capillary sequencer to determine the relative frequencies of the mutant and wild-type alleles. Applying DARMS-PCR to human specimens, we successfully identified MPL mutations in MPN patients, with the exception of patients bearing mutant allele frequencies below the detection limit (5%) of this method. The MPL mutant allele frequencies determined using DARMS-PCR correlated strongly with the values determined using deep sequencing. Thus, we demonstrated the potential of DARMS-PCR to detect MPL mutations and determine the allele frequencies in a timely and cost-effective manner.     

Mechanism for generation of hydrogen peroxide and change of mitochondrial membrane potential during rotenone-induced apoptosis

Rotenone, an inhibitor of NADH dehydrogenase complex, is a naturally occurring insecticide, which is capable of inducing apoptosis. Rotenone-induced apoptosis is considered to contribute to its anticancer effect and the etiology of Parkinson's disease (PD). We demonstrated that rotenone induced internucleosomal DNA fragmentation, DNA ladder formation, in human cultured cells, HL-60 (promyelocytic leukemia) and BJAB cells (B-cell lymphoma). Flow cytometry showed that rotenone induced H2O2 generation, followed by significant changes in the mitochondrial membrane potential (DeltaPsim). Caspase-3 activity increased in HL-60 cells in a time-dependent manner. These apoptotic events were delayed in HP100 cells, an H2O2-resistant clone of HL-60, confirming the involvement of H2O2 in apoptosis. Expression of anti-apoptotic protein, Bcl-2, in BJAB cells drastically inhibited DeltaPsim change and DNA ladder formation but not H2O2 generation, confirming the participation of mitochondrial dysfunction in apoptosis. NAD(P)H oxidase inhibitors prevented H2O2 generation and DNA ladder formation. These results suggest that rotenone induces O2(-)-derived H2O2 generation through inhibition of NADH dehydrogenase complex and/or activation of NAD(P)H oxidase, and H2O2 generation causes the disruption of mitochondrial membrane in rotenone-induced apoptosis.     

Inhibition of DNA polymerases of sea urchin by palmitoyl coenzyme A

Palmitoyl CoA noncompetitively inhibited the activities of DNA polymerase α and γ, prepared from sea urchin germ cells, with Ki values of 28 μM and 116 μM, respectively. Myristoyl CoA also inhibited DNA polymerse α and γ, while coenzyme A, short chain fatty acyl CoA's, Na-myristate and Na-palmitate failed to inhibit the enzymes. It was concluded that both the long hydrocarbon chain and CoA moiety of long chain fatty acyl CoA's are necessary for inhibition of DNA polymerase activity. DNA polymerse β was not inhibited by long chain fatty acyl CoA's.     

cDNA cloning and sequencing of tobacco chloroplast ribosomal protein L12

Tobacco chloroplast ribosomal protein L12 was isolated as a ssDNA-cellulose-binding protein from a chloroplast soluble protein fraction. Based on the N-terminal amino acid sequence of chloroplast L12, a cDNA clone was isolated and characterized. The precursor protein deduced from the DNA sequence consists of a transit peptide of 53 amino acid residues and a mature L12 protein of 133 amino acid residues. The chloroplast L12 protein was synthesized with a reticulocyte lysate and subjected to nucleic acid-binding assays. L12 synthesized in vitro does not bind to ssDNA, dsDNA nor ribonucleotide homopolymers, but it binds to cellulose matrix.