Genome sequence of Soos: a siphovirus of the CP cluster infecting .

Novel actinobacteriophage Soos was isolated and purified from Southern Indiana soil using host NRRL B-16540. Sequencing revealed a 57,509 bp circularly permuted genome encoding 87 predicted protein-coding genes. Soos is only the third phage in cluster CP, along with phages Clawz and Sting.

Genome Sequence of Gordonia rubripertincta Phage Survivors, a Cluster CT Siphovirus.

Bacteriophage Survivors is a siphovirus isolated from Gordonia rubripertincta NRRL B-16540. Survivors has a 45,436-bp genome encoding 69 predicted protein-coding genes, of which 32 have assigned functions. Based on gene content similarity to sequenced actinobacteriophages, Survivors is assigned to phage cluster CT.

Linear external skeletal fixation applied in minimally invasive fashion for stabilization of nonarticular tibial fractures in dogs and cats.

Objective: The objective of this study was to evaluate the use of linear external skeletal fixation (ESF) applied using minimally invasive techniques in dogs and cats.

Study Design: Retrospective study.

Animals: Forty-nine dogs and 6 cats.

Elimination of rNMPs from mitochondrial DNA has no effect on its stability.

Ribonucleotides (rNMPs) incorporated in the nuclear genome are a well-established threat to genome stability and can result in DNA strand breaks when not removed in a timely manner. However, the presence of a certain level of rNMPs is tolerated in mitochondrial DNA (mtDNA) although aberrant mtDNA rNMP content has been identified in disease models. We investigated the effect of incorporated rNMPs on mtDNA stability over the mouse life span and found that the mtDNA rNMP content increased during early life.

Heterozygous colon cancer-associated mutations of SAMHD1 have functional significance.

Even small variations in dNTP concentrations decrease DNA replication fidelity, and this observation prompted us to analyze genomic cancer data for mutations in enzymes involved in dNTP metabolism. We found that sterile alpha motif and histidine-aspartate domain-containing protein 1 (SAMHD1), a deoxyribonucleoside triphosphate triphosphohydrolase that decreases dNTP pools, is frequently mutated in colon cancers, that these mutations negatively affect SAMHD1 activity, and that several SAMHD1 mutations are found in tumors with defective mismatch repair. We show that minor changes in dNTP pools in combination with inactivated mismatch repair dramatically increase mutation rates.

Genome-wide analysis of the specificity and mechanisms of replication infidelity driven by imbalanced dNTP pools.

The absolute and relative concentrations of the four dNTPs are key determinants of DNA replication fidelity, yet the consequences of altered dNTP pools on replication fidelity have not previously been investigated on a genome-wide scale. Here, we use deep sequencing to determine the types, rates and locations of uncorrected replication errors that accumulate in the nuclear genome of a mismatch repair-deficient diploid yeast strain with elevated dCTP and dTTP concentrations. These imbalanced dNTP pools promote replication errors in specific DNA sequence motifs suggesting increased misinsertion and increased mismatch extension at the expense of proofreading.

Increased and imbalanced dNTP pools symmetrically promote both leading and lagging strand replication infidelity.

The fidelity of DNA replication requires an appropriate balance of dNTPs, yet the nascent leading and lagging strands of the nuclear genome are primarily synthesized by replicases that differ in subunit composition, protein partnerships and biochemical properties, including fidelity. These facts pose the question of whether imbalanced dNTP pools differentially influence leading and lagging strand replication fidelity. Here we test this possibility by examining strand-specific replication infidelity driven by a mutation in yeast ribonucleotide reductase, rnr1-Y285A, that leads to elevated dTTP and dCTP concentrations.

Altered Ig hypermutation pattern and frequency in complementary mouse models of DNA polymerase ζ activity.

To test the hypothesis that DNA polymerase ζ participates in Ig hypermutation, we generated two mouse models of Pol ζ function: a B cell-specific conditional knockout and a knock-in strain with a Pol ζ mutagenesis-enhancing mutation. Pol ζ-deficient B cells had a reduction in mutation frequency at Ig loci in the spleen and in Peyer's patches, whereas knock-in mice with a mutagenic Pol ζ displayed a marked increase in mutation frequency in Peyer's patches, revealing a pattern that was similar to mutations in yeast strains with a homologous mutation in the gene encoding the catalytic subunit of Pol ζ. Combined, these data are best explained by a direct role for DNA polymerase ζ in Ig hypermutation.

RNase H and postreplication repair protect cells from ribonucleotides incorporated in DNA.

The chemical identity and integrity of the genome is challenged by the incorporation of ribonucleoside triphosphates (rNTPs) in place of deoxyribonucleoside triphosphates (dNTPs) during replication. Misincorporation is limited by the selectivity of DNA replicases. We show that accumulation of ribonucleoside monophosphates (rNMPs) in the genome causes replication stress and has toxic consequences, particularly in the absence of RNase H1 and RNase H2, which remove rNMPs.

Replication of ribonucleotide-containing DNA templates by yeast replicative polymerases.

The major replicative DNA polymerases of S. cerevisiae (Pols α, δ, and ɛ) incorporate substantial numbers of ribonucleotides into DNA during DNA synthesis. When these ribonucleotides are not removed in vivo, they reside in the template strand used for the next round of replication and could potentially reduce replication efficiency and fidelity.

Genome instability due to ribonucleotide incorporation into DNA.

Maintaining the chemical identity of DNA depends on ribonucleotide exclusion by DNA polymerases. However, ribonucleotide exclusion during DNA synthesis in vitro is imperfect. To determine whether ribonucleotides are incorporated during DNA replication in vivo, we substituted leucine or glycine for an active-site methionine in yeast DNA polymerase ϵ (Pol ϵ).

Abundant ribonucleotide incorporation into DNA by yeast replicative polymerases.

Measurements of nucleoside triphosphate levels in Saccharomyces cerevisiae reveal that the four rNTPs are in 36- to 190-fold molar excess over their corresponding dNTPs. During DNA synthesis in vitro using the physiological nucleoside triphosphate concentrations, yeast DNA polymerase epsilon, which is implicated in leading strand replication, incorporates one rNMP for every 1,250 dNMPs. Pol delta and Pol alpha, which conduct lagging strand replication, incorporate one rNMP for every 5,000 or 625 dNMPs, respectively.

Mutagenicity of the 1-nitropyrene-DNA adduct N-(deoxyguanosin-8-yl)-1-aminopyrene in mammalian cells.

The mutagenesis of the major DNA adduct N-(deoxyguanosin-8-yl)-1-aminopyrene (C8-AP-dG) formed by 1-nitropyrene was compared with the analogous C8-dG adducts of 2-aminofluorene (AF) and N-acetyl-2-aminofluorene (AAF) in simian kidney (COS-7) cells. The DNA sequence chosen for this comparison contained 5'-CCATC GCTACC-3' that has been used for solution NMR investigations. The structural and conformational differences among these lesions are well-established [Patel, D.

Polycyclic aromatic hydrocarbon-DNA adducts determined by semiquantitative immunohistochemistry in human esophageal biopsies taken in 1985.

Esophageal endoscopic biopsy samples were obtained in 1985 in Linxian, China, a region with very high esophageal cancer incidence rates, and where ingested food is known to contain substantial amounts of polycyclic aromatic hydrocarbons (PAHs). In this study, the automated cellular imaging system (ACIS) was used for localization and semi-quantitation of PAH-DNA adducts. Fresh tissue sections were cut from archived paraffin blocks and incubated with an antiserum elicited against DNA modified with 7beta,8alpha-dihydroxy-9alpha,10alpha-epoxy-7,8,9,10-tetrahydro-benzo[a]pyrene (BPDE).