DNA slip-outs cause RNA polymerase II arrest in vitro: potential implications for genetic instability

The abnormal number of repeats found in triplet repeat diseases arises from ‘repeat instability’, in which the repetitive section of DNA is subject to a change in copy number. Recent studies implicate transcription in a mechanism for repeat instability proposed to involve RNA polymerase II (RNAPII) arrest caused by a CTG slip-out, triggering transcription-coupled repair (TCR), futile cycles of which may lead to repeat expansion or contraction. In the present study, we use defined DNA constructs to directly test whether the structures formed by CAG and CTG repeat slip-outs can cause transcription arrest in vitro. We found that a slip-out of (CAG)20 or (CTG)20 repeats on either strand causes RNAPII arrest in HeLa cell nuclear extracts. Perfect hairpins and loops on either strand also cause RNAPII arrest. These findings are consistent with a transcription-induced repeat instability model in which transcription arrest in mammalian cells may initiate a ‘gratuitous’ TCR event leading to a change in repeat copy number. An understanding of the underlying mechanism of repeat instability could lead to intervention to slow down expansion and delay the onset of many neurodegenerative diseases in which triplet repeat expansion is implicated

Inhibitory effect of a short Z-DNA forming sequence on transcription elongation by T7 RNA polymerase

DNA sequences capable of forming unusual secondary structures can be a source of genomic instability. In some cases that instability might be affected by transcription, as recently shown for the Z-DNA forming sequence (CG)14, which causes genomic instability both in mammalian cells and in bacteria, and this effect increases with its transcription. We have investigated the effect of this (CG)14 sequence on transcription with T7 RNA polymerase in vitro. We detected partial transcription blockage within the sequence; the blockage increased with negative supercoiling of the template DNA. This effect was not observed in a control self-complementary sequence of identical length and base composition as the (CG)14 sequence, when the purine–pyrimidine alternation required for Z-DNA formation was disrupted. These findings suggest that the inhibitory effect on T7 transcription results from Z-DNA formation in the (CG)14 sequence rather than from an effect of the sequence composition or from hairpin formation in either the DNA or the RNA product

Building on the past, shaping the future: The environmental mutagenesis and genomics society

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/97167/1/em21765.pd

Medieval menarche: changes in pubertal timing in the aftermath of the Black Death

OBJECTIVES: Bioarchaeological evidence suggests stature increased in males but decreased in females after the Black Death (1348-1350 CE). Because tradeoffs between growth and reproduction can result in earlier ages at menarche and lower limb length, we assess menarcheal age between 1120 and 1540 CE to better understand the health of medieval adolescent females before and after the plague. MATERIALS AND METHODS: Our sample comprises 74 adolescent females from St Mary Spital, London (1120-1540 CE) within the age range during which menarche occurs (10-25 years). They were assessed as being pre- or post-menarcheal and divided into three groups: Early Pre-Black Death (n=13), Late Pre-Black Death (n=38), and Post-Black Death (n=23). Changes in the ages of pre- and post-menarcheal females were assessed using Mann-Whitney tests. RESULTS: The average age of post-menarcheal females increased from the Early- to Late Pre-Black Death periods and declined after the Black Death. CONCLUSIONS: Short stature can reflect unfavorable growth environments, while younger menarcheal age indicates improved living conditions. The paradoxical pattern of female, but not male, stature reduction after the Black Death might reflect the association of early menarche with lower limb length and signal that adolescent females experienced improved health conditions after the epidemic. Our focus on pre- and post-menarche within a limited age span provides a novel approach for inferring average ages of menarche over time. Pathways to skeletal development and reproductive investment are part of an integrated system, providing a bridge between life history research in bioarchaeology and human biology

Nicotine Overrides DNA Damage-Induced G1/S Restriction in Lung Cells

As an addictive substance, nicotine has been suggested to facilitate pro-survival activities (such as anchorage-independent growth or angiogenesis) and the establishment of drug resistance to anticancer therapy. Tobacco smoking consists of a variety of carcinogens [such as benzopyrene (BP) and nitrosamine derivatives] that are able to cause DNA double strand breaks. However, the effect of nicotine on DNA damage-induced checkpoint response induced by genotoxins remains unknown. In this study, we investigated the events occurred during G1 arrest induced by γ-radiation or BP in nicotine-treated murine or human lung epithelial cells. DNA synthesis was rapidly inhibited after exposure to γ-radiation or BP treatment, accompanied with the activation of DNA damage checkpoint. When these cells were co-treated with nicotine, the growth restriction was compromised, manifested by upregulation of cyclin D and A, and attenuation of Chk2 phosphorylation. Knockdown of cyclin D or Chk2 by the siRNAs blocked nicotine-mediated effect on DNA damage checkpoint activation. However, nicotine treatment appeared to play no role in nocodazole-induced mitotic checkpoint activation. Overall, our study presented a novel observation, in which nicotine is able to override DNA damage checkpoint activated by tobacco-related carcinogen BP or γ-irradiation. The results not only indicates the potentially important role of nicotine in facilitating the establishment of genetic instability to promote lung tumorigenesis, but also warrants a dismal prognosis for cancer patients who are smokers, heavily exposed second-hand smokers or nicotine users

Bone histomorphometric measures of physical activity in children from Medieval England

Objectives: Histomorphometric studies show consistent links between physical activity patterns and the microstructure underlying the size and shape of bone. Here we adopt a combined bone approach to explore variation in microstructure of ribs and humeri related to physical activity and historical records of manual labor in skeletal samples of children (n=175) from medieval England. The humerus reflects greater biomechanically induced microstructural variation than the rib which is used here as a control. Variation in microstructure is sought between regions in England (Canterbury, York, Newcastle), and between high- and low-status children from Canterbury. Materials and Methods: Thin-sections were prepared from the humerus or rib and features of bone remodeling were recorded using high-resolution microscopy and image analysis software. Results: The density and size of secondary osteons in the humerus differed significantly in children from Canterbury when compared to those from York and Newcastle. Amongst the older children, secondary osteon circularity and diameter differed significantly between higher and lower status children. Discussion: By applying bone remodeling principles to the histomorphometric data we infer that medieval children in Canterbury engaged in less physically demanding activities than children from York or Newcastle. Within Canterbury, high-status and low-status children experienced similar biomechanical loading until around seven years of age. After this age low-status children performed activities that resulted in more habitual loading on their arm bones than the high-status children. This inferred change in physical activity is consistent with historical textual evidence that describes children entering the work force at this age