Oxidative Stress Is Not a Major Contributor to Somatic Mitochondrial DNA Mutations

The accumulation of somatic mitochondrial DNA (mtDNA) mutations is implicated in aging and common diseases of the elderly, including cancer and neurodegenerative disease. However, the mechanisms that influence the frequency of somatic mtDNA mutations are poorly understood. To develop a simple invertebrate model system to address this matter, we used the Random Mutation Capture (RMC) assay to characterize the age-dependent frequency and distribution of mtDNA mutations in the fruit fly Drosophila melanogaster. Because oxidative stress is a major suspect in the age-dependent accumulation of somatic mtDNA mutations, we also used the RMC assay to explore the influence of oxidative stress on the somatic mtDNA mutation frequency. We found that many of the features associated with mtDNA mutations in vertebrates are conserved in Drosophila, including a comparable somatic mtDNA mutation frequency (∼10⁻⁵), an increased frequency of mtDNA mutations with age, and a prevalence of transition mutations. Only a small fraction of the mtDNA mutations detected in young or old animals were G∶C to T∶A transversions, a signature of oxidative damage, and loss-of-function mutations in the mitochondrial superoxide dismutase, Sod2, had no detectable influence on the somatic mtDNA mutation frequency. Moreover, a loss-of-function mutation in Ogg1, which encodes a DNA repair enzyme that removes oxidatively damaged deoxyguanosine residues (8-hydroxy-2′-deoxyguanosine), did not significantly influence the somatic mtDNA mutation frequency of Sod2 mutants. Together, these findings indicate that oxidative stress is not a major cause of somatic mtDNA mutations. Our data instead suggests that somatic mtDNA mutations arise primarily from errors that occur during mtDNA replication. Further studies using Drosophila should aid in the identification of factors that influence the frequency of somatic mtDNA mutations.     

Evidence of multiple paternity in spiny dogfish (Squalus acanthias) broods based on microsatellite analysis

An understanding of alternative mating systems is a fundamental requirement for the effective management of vulnerable species. Microsatellite analysis of spiny dogfish Squalus acanthias broods from the north‐west Atlantic provides novel evidence of polyandry in this species.     

Bandign isolated metaphase chromosomes by a sequential flurescent G/Q technique.

G/Q-banding is a new, rapid, fluorescent technique for banding isolated chromosomes that incorporates characteristics of both G- and Q- banding. G-bands, while easily characterized, are often inconsistent when using isolated chromosomes, and Q-bands, while reliable, fade rapidly under UV exposure, making prolonged observation and photography difficult. G/Q-banding combines these techniques to sequentially utilize quinacrine staining over Giemsa banding to produce slow-fading fluorescent G/Q-bands. The background fluorescence in G/Q preparations fades quickly under continued UV exposure, while the chromosomes remain brightly banded and can be observed and photographed for at least five minutes. G/Q-banding was extended to whole cell chromosome spreads and produced results identical to those obtained with isolated chromosomes. Whole cell karyotypes indicate that G/Q-bands generally correspond to Q-bands. Advantages of G/Q-banding as a unique and universal technique over current double-staining procedures are discussed.     

The serological responses of calves to live Salmonella dublin vaccine--a comparison of different serological tests

The serological responses to live Salmonella dublin vaccine was assessed in three groups of calves; three-day-old colostrum-deprived (3DO C-), three-day-old colostrum-fed (3DO C+) and three-month-old (3MO), by the following tests; serum agglutination test (SAT), indirect haemagglutination test (IHA), complement-fixation test (CFT) and antiglobulin test (AGT). Serological activity was detected by all the tests in the 3MO calves. In the 3DO C+ calves no serological activity was detected by either the somatic SAT or IHA but low levels of CF and somatic AGT antibodies were produced. In 3DO C- calves serological activity, often at low levels, was detected by all the tests except the somatic SAT. High levels of flagellar agglutinins were detected in both groups of 3DO calves. It was concluded that with the exception of the flagellar SAT the tests were affected by the age of the calf and in 3DO calves also by the presence of colostral antibodies. However, the use of the SAT in 3MO calves would provide an indication as to the potency of salmonella vaccines.     

Neurodevelopmental toxicity assessment of flame retardants using a human DNT in vitro testing battery

Due to their neurodevelopmental toxicity, flame retardants (FRs) like polybrominated diphenyl ethers are banned from the market and replaced by alternative FRs, like organophosphorus FRs, that have mostly unknown toxicological profiles. To study their neurodevelopmental toxicity, we evaluated the hazard of several FRs including phased-out polybrominated FRs and organophosphorus FRs: 2,2′,4,4′-tetrabromodiphenylether (BDE-47), 2,2′,4,4′,5-pentabromodiphenylether (BDE-99), tetrabromobisphenol A, triphenyl phosphate, tris(2-butoxyethyl) phosphate and its metabolite bis-(2-butoxyethyl) phosphate, isodecyl diphenyl phosphate, triphenyl isopropylated phosphate, tricresyl phosphate, tris(1,3-dichloro-2-propyl) phosphate, tert-butylphenyl diphenyl phosphate, 2-ethylhexyl diphenyl phosphate, tris(1-chloroisopropyl) phosphate, and tris(2-chloroethyl) phosphate. Therefore, we used a human cell–based developmental neurotoxicity (DNT) in vitro battery covering a large variety of neurodevelopmental endpoints. Potency according to the respective most sensitive benchmark concentration (BMC) across the battery ranked from <1 μM (5 FRs), 1<10 μM (7 FRs) to the >10 μM range (3 FRs). Evaluation of the data with the ToxPi tool revealed a distinct ranking (a) than with the BMC and (b) compared to the ToxCast data, suggesting that DNT hazard of these FRs is not well predicted by ToxCast assays. Extrapolating the DNT in vitro battery BMCs to human FR exposure via breast milk suggests low risk for individual compounds. However, it raises a potential concern for real-life mixture exposure, especially when different compounds converge through diverse modes-of-action on common endpoints, like oligodendrocyte differentiation in this study. This case study using FRs suggests that human cell–based DNT in vitro battery is a promising approach for neurodevelopmental hazard assessment and compound prioritization in risk assessment.

Graphical abstract

     

Biparental contributions of the H2A.B histone variant control embryonic development in mice

Histone variants expand chromatin functions in eukaryote genomes. H2A.B genes are testis-expressed short histone H2A variants that arose in placental mammals. Their biological functions remain largely unknown. To investigate their function, we generated a knockout (KO) model that disrupts all 3 H2A.B genes in mice. We show that H2A.B KO males have globally altered chromatin structure in postmeiotic germ cells. Yet, they do not show impaired spermatogenesis or testis function. Instead, we find that H2A.B plays a crucial role postfertilization. Crosses between H2A.B KO males and females yield embryos with lower viability and reduced size. Using a series of genetic crosses that separate parental and zygotic contributions, we show that the H2A.B status of both the father and mother, but not of the zygote, affects embryonic viability and growth during gestation. We conclude that H2A.B is a novel parental-effect gene, establishing a role for short H2A histone variants in mammalian development. We posit that parental antagonism over embryonic growth drove the origin and ongoing diversification of short histone H2A variants in placental mammals.

The unusual short histone variant H2A.B is a novel parental-effect gene that plays an important role in early mammalian development. Parental antagonism over embryonic growth resource allocation may have driven the origin and ongoing diversification of short histone H2A variants in placental mammals.     

Sarmentosin epoxide,a new cyanogenic compound from Sedum cepaea

A new cyanogenic compound, sarmentosin epoxide, was isolated from the aerial parts of Sedum cepaea. Its structure was established mainly by ¹H