Mismatch repair proteins are activators of toxic responses to chromium-DNA damage

Chromium(VI) is a toxic and carcinogenic metal that causes the formation of DNA phosphate-based adducts. Cr-DNA adducts are genotoxic in human cells, although they do not block replication in vitro. Here, we report that induction of cytotoxicity in Cr(VI)-treated human colon cells and mouse embryonic fibroblasts requires the presence of all major mismatch repair (MMR) proteins. Cr-DNA adducts lost their ability to block replication of Cr-modified plasmids in human colon cells lacking MLH1 protein. The presence of functional mismatch repair caused induction of p53-independent apoptosis associated with activation of caspases 2 and 7. Processing of Cr-DNA damage by mismatch repair resulted in the extensive formation of gamma-H2AX foci in G(2) phase, indicating generation of double-stranded breaks as secondary toxic lesions. Induction of gamma-H2AX foci was observed at 6 to 12 h postexposure, which was followed by activation of apoptosis in the absence of significant G(2) arrest. Our results demonstrate that mismatch repair system triggers toxic responses to Cr-DNA backbone modifications through stress mechanisms that are significantly different from those for other forms of DNA damage. Selection for Cr(VI) resistant, MMR-deficient cells may explain the very high frequency of lung cancers with microsatellite instability among chromate workers.     

The cellular responses to DNA damage

The ability to survive spontaneous and induced DNA damage, and to minimize the number of heritable mutations that this causes, is essential to the maintenance of genome integrity for all organisms. Early studies on model eukaryotes focused on genes acting in defined DNA repair pathways. More recent work with the budding and fission yeasts and mammalian cells has started to integrate the DNA damage response with cell physiology and the cell cycle.     

The responses of some Lepidoptera to labiate herb and white clover extracts

Herbs have been traditionally used as intercrops with crop plants on the assumption that their odour repels pest species. Alcohol extracts and essential oils of labiate herbs were tested in the laboratory for deterrent/repellent responses to ovipositing Plutella xylostella (L.) and feeding larvae of P. xylostella and Pieris brassicae L.Alcohol extracts of hyssop, rosemary, sage, thyme and white clover reduce oviposition by P. xylostella on pieces of brassica leaf in dual-choice tests. Essential oils of sage and thyme reduce oviposition on pieces of brassica leaf. Feeding, in dual-choice tests, by final instar P. xylostella and P. brassicae larvae is reduced by application of alcohol extracts of herbs.

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Résumé Des plantes médicinales ont traditionnellement été utilisées en association avec des plantes cultivées dans l'espoir que leur odeur repousserait les insectes nuisibles. Des extraits à l'alcool et des essences volatiles de labiées médicinales ont été essayés au laboratoire pour examiner leur effets répulsif et dissuadant pour la ponte de Plutella xylostella et l'alimentation des chenilles de P. xylostella et Pieris brassicae. Les extraits alcooliques d'hysope, de romarin, de sauge, de thym et de trèfle blanc réduisent la ponte de P. xylostella sur des morceaux de feuilles de chou traitées. Les essences volatiles de sauge et de thym réduisent la ponte de P. xylostella sur des morceaux de feuilles de chou dans des expériences de choix. L'alimentation des chenilles de dernier stade de P. xylostella et P. brassicae dans des expériences de choix est réduite par l'application d'extraits alcooliques des labiées médicinales.

     

Insect herbivory in an intact forest understory under experimental CO2 enrichment

Human-induced increases in atmospheric CO₂ concentration have the potential to alter the chemical composition of plant tissue, and thereby affect the amount of tissue consumed by herbivorous arthropods. At the Duke Forest free-air concentration enrichment (FACE) facility in North Carolina (FACTS–1 research facility), we measured the amount of leaf tissue damaged by insects and other herbivorous arthropods during two growing seasons in a deciduous forest understory continuously exposed to ambient (360 l l^–1) and elevated (~560 µl l^–1) CO₂ conditions. In 1999, there was a significant interaction between CO₂ and species such that winged elm (Ulmus alata) showed lower herbivory in elevated CO₂ plots, whereas red maple (Acer rubra) and sweetgum (Liquidambar styraciflua) did not. In 2000, our results did not achieve statistical significance but the magnitude of the result was consistent with the 1999 results. In 1999 and 2000, we found a decline (10–46%) in community-level herbivory in elevated CO₂ plots driven primarily by reductions in herbivory on elm. The major contribution to total leaf damage was from missing tissue (66% of the damaged tissue), with galls, skeletonized, and discolored tissue making smaller contributions. It is unclear whether the decline in leaf damage is a result of altered insect populations, altered feeding, or a combination. We were not able to quantify insect populations, and our measurements did not resolve an effect of elevated CO₂ on leaf chemical composition (total nitrogen, carbon, C/N, sugars, phenolics, starch). Despite predictions from a large number of single-species studies that herbivory may increase under elevated CO₂, we have found a decrease in herbivory in a naturally established forest understory exposed to a full suite of insect herbivores and their predators.     

A slow opportunist: physiological and growth responses of an obligate understory plant to patch cut harvesting

Understory light environments change rapidly following timber harvest, and while many understory species utilize and benefit from the additional light, this response is not ubiquitous in shade-obligate species. I examined the effects of patch cut timber harvest on the physiology and growth of an obligate forest understory species to determine if disturbances via timber harvest are physiological stressors or whether such disturbances provide physiological benefits and growth increases in understory species. Forest canopy structure, along with photosynthesis, respiration, water use efficiency, stomatal conductance, and growth rates of American ginseng were quantified one summer before and two summers after patch cut timber harvest. Survival following timber harvest was lower than that observed at undisturbed populations; however, growth of survivors increased post-harvesting, with growth increasing as a function of canopy openness. Light response curves as well as photosynthesis and respiration rates indicated that plants were not well acclimated to higher light levels in the growing season after timber harvest, but rather to two growing seasons after harvest. Relative growth rate formed a positive linear relationship with maximum photosynthesis following timber harvest. My study suggests that ginseng is a “slow opportunist”, because while it benefits from sudden light increases, acclimation lags at least one growing season behind canopy changes. American ginseng is surprisingly resilient in the face of a discrete environmental shift and may benefit from forest management strategies that mimic the natural disturbance regimes common in mature forests throughout its range.     

Influences of invasive herb Coreopsis lanceolata on riparian endemic herbs in relation to the understory light availability

In Japan, invasions of alien herb Coreopsis lanceolata are often observed in riparian endemic vegetation, but the invasion influences on light environments and endemic species are insufficiently evaluated. In this study, we investigated the influences of C. lanceolata and native dominant turf Zoysia japonica on the densities of riparian plants (two endemics Artemisia capillaris and Potentilla chinensis and one alien Lespedeza inschanica) in relation to the understory light availability using 594 0.2 m × 0.2 m plots. Available understory light was influenced negatively by C. lanceolata, not Z. japonica, due probably to the tall stature and dense foliar canopy of C. lanceolata. Understory light availability positively affected densities of riparian endemics, although the relation with L. inschanica was unclear. A. capillaris and P. chinensis densities were negatively related with C. lanceolata, not Z. japonica, implying that C. lanceolata reduces endemic populations. Shading by C. lanceolata would be one of prime factors reducing the endemics. L. inschanica density was not correlated with C. lanceolata cover. C. lanceolata could not shade L. inschanica effectively due to the taller stature of L. inschanica. We considered that C. lanceolata reduces endemic riparian species but coexists with L. inschanica. Preventing invasion and dominance of C. lanceolata is desirable to conserve endemic riparian vegetation.     

Responses of root length/leaf area ratio and specific root length of an understory herb, Pteridophyllum racemosum, to increases in irradiance

The understory evergreen perennial Pteridophyllum racemosum Sieb. et Zucc. (Papaveraceae) has the ability to increase root mass per unit transpiring leaf area (RMA) if irradiance increases gradually over several years. In this study, we examined how P. racemosum changes its root length/leaf area ratio and specific root length when the species encounters abrupt increases in irradiance, such as sudden and unexpected canopy openings. Plants were transplanted from a low light condition in a subalpine wave-regenerating forest (photon flux density on the forest floor relative to the full sun (RPFD) was 2.7%) to a high light condition in a glasshouse (30% RPFD) (LH treatment). Transplantation from the low light condition in the forest to a low light condition in the glasshouse (LL) and transplantation from a high light condition in the forest (33% RPFD) to a high light condition in the glasshouse (HH) were also conducted as controls. Compared to the LL plants, the LH plants exhibited significant increases in RMA and root length/leaf area ratio from 30 to 70 days after transplantation. On the other hand, the effect of increased irradiance on specific root length (SRL) was weak, and both the LL and LH plants showed increased SRL from 30 to 70 days after transplantation. Increased SRL results from longer root length per unit construction cost. We concluded that increased root length/leaf area ratio of P. racemosum in response to abrupt increases in irradiance was caused by a combination of enhanced carbon allocation to roots with increased SRL.     

Heavy metal accumulation and distribution in forest understory herb species of Carpathian beech ecosystems

Abstract

This study addresses the issue of heavy metal (HM) accumulation and distribution for three different plant species, Carex pilosa, Dentaria bulbifera, Galium odoratum, in Carpathian beech ecosystems. Data are presented on HM concentrations in forest understory vegetation and a preliminary insight into different HM allocation patterns is provided. Bioaccumulation factors (BCFs) and shoot/root ratios differed considerably among the species and between polluted and unpolluted regions. HMs were accumulated in forest plants as follows: Cu > Zn > Cd >Pb in unpolluted areas and Zn> Cd > Cu >Pb in polluted areas. Zn was preferentially distributed to roots and Cu to shoots. The distribution of Cd and Pb in different plant parts was specific in terms of the species-dependence. Cd and Pb levels in Carex pilosaand Galium odoratumwere more strictly controlled in the transfer zone of root-shoot, compared to Dentaria bulbifera.The highest BCFs were found in Carex pilosafor Cu (5.9) and in Dentaria bulbiferawas found the highest shoot/root ratio for Cd (3.1).     

Telomeres are double-strand DNA breaks hidden from DNA damage responses

A network of ATM/ATR-mediated events regulates cell cycle checkpoints and genomic integrity and contributes to the processing of DNA double-strand breaks in both genomic DNA and at telomeres. In yeast and in human cells, investigators, including, and Herbig et al., published in this issue of Molecular Cell, are beginning to decipher the signaling pathways involved at the telomeres.     

Competition as an amplifier of short-term vegetation responses to climate: an experimental test

1. Responses to a spring warming treatment were measured on five common herbaceous species grown in outdoor microcosms in Northern England. Although elevated temperature had a beneficial effect on canopy height and plant cover in monospecific cultures of all species, strongly divergent responses to warming occurred in mixtures.
2. We show that the effect of interspecific competition was to modify and amplify the vegetation response to the warming treatment through effects on phenology and morphological development.
3. The observed responses between the species to warming are consistent with predicted differential responses linked to genome size.
4. We conclude that the mechanism of competitive interaction proposed by Boysen-Jensen (1929 ) coupled with the insights related to genome size provide a basis for explaining and predicting the role of interannual variation in temperature in determining year to year fluctuations in the relative abundance of species in productive perennial herbaceous vegetation.     

DNA damage responses to oxidative stress

The DNA damage response is a hierarchical process. DNA damage is detected by sensor proteins such as the MRN complex that transmit the information to transducer proteins such as ATM and ATR, which control the damage response through the phosphorylation of effector proteins. The extent of the DNA damage determines cell fate: cell cycle arrest and DNA repair or the activation of apoptotic pathways. In aerobic cells, reactive oxygen species (ROS) are generated as a by-product of normal mitochondrial activity. If not properly controlled, ROS can cause severe damage to cellular macromolecules, especially the DNA. We describe here some of the cellular responses to alterations in the cellular redox state during hypoxia or oxidative stress. Oxidative damage in DNA is repaired primarily via the base excision repair (BER) pathway which appears to be the simplest of the three excision repair pathways. To allow time for DNA repair, the cells activate their cell cycle checkpoints, leading to cell cycle arrest and preventing the replication of damage and defective DNA.     

Ipsilateral vagotomy to unilaterally ovariectomized pre-pubertal rats modifies compensatory ovarian responses

The present study evaluates the participation of the vagus nerve in pre-pubertal rats with unilateral ovariectomy on puberty onset, and on progesterone, testosterone and estradiol serum levels, and the compensatory responses of the ovary. Unilateral vagotomy did not modify the onset of puberty in unilaterally ovariectomized rats. Ovulation rates of animals with the left vagus nerve sectioned and the left ovary in-situ was lower than in rats with only unilateral ovariectomy. Sectioning the left vagus to 32-day old rats with the left ovary in-situ resulted in lower compensatory ovarian hypertrophy than in rats with right unilateral ovariectomy. Twenty-eight or 32-day old animals with sectioning of the right vagus nerve and the right ovary in situ showed higher compensatory ovulation. Twenty-eight -day old rats with the right ovary in situ had higher progesterone and testosterone levels than animals of the same age with the left ovary in-situ. Compared to animals with the right ovary in situ, animals treated at 32-days of age, sectioning the ipsi-lateral vagus nerve resulted in higher progesterone levels. Higher progesterone levels were observed in 28- and 32 days old rats with the left ovary in situ and left vagus nerve sectioned. Thirty-two day old animals with the right ovary in situ and right vagus nerve sectioned had higher progesterone levels than rats of the same age with the left ovary in situ and left vagus nerve sectioned. Left vagotomy to 28-day old rats with the left ovary in situ resulted in higher testosterone levels, a reverse response to that observed in animals with sectioning of the right vagus and the right ovary in situ. Thirty-two day old rats with the left ovary in situ and left vagus nerve sectioned showed lower testosterone levels than animals without vagotomy and with the left ovary in situ.     

Cyclophosphamide teratogenesis: evidence for compensatory responses to induced cellular toxicity

Cyclophosphamide (CP) administered ip to pregnant mice on day 10 of gestation (day of plug = day 0) is teratogenic (exencephaly, cleft palate, and limb malformations) at 20 mg/kg and embryolethal at higher doses. In the present study, CP was administered at 1, 5, 10, or 20 mg/kg on day 10 of gestation. Embryos were removed at 8 and 28 hr postdosing, and two embryos from each litter were immediately stained with Nile blue sulfate (NBS) to identify areas of cell death. The remaining embryos were frozen and forelimb buds subsequently removed for flow cytometric (FCM) analysis of the cellular DNA synthetic cycle. Additional litters were examined near term (day 17) for morphological abnormalities; these data were correlated with embryonic toxicity as detected by NBS staining and FCM analysis. Only the highest dose produced malformations. In marked contrast, a dose-related increase in the percentage of limb bud cells in the S (DNA synthetic) phase of the cell cycle was detectable at all doses. Inhibition of DNA synthesis was detected at all doses 8 hr post exposure and persisted through 28 hr for doses greater than or equal to 10 mg/kg. NBS staining indicated increased cell death in the alar plate of the neural tube 28 hr after exposure to 10 mg/kg CP and generally increased cell death in areas of rapid cell proliferation throughout the embryo at 20 mg/kg. The absence of an overt teratogenic response at dose levels that produced significant perturbation of the cell cycle indicates that a measure of embryonic damage can be compensated for or repaired. The implications of these findings for the existence of thresholds in developmental toxicity are discussed.     

Effects of light and nutrient availability on chasmogamy and cleistogamy in an understory tropical herb,Calathea micans (Marantaceae)

The effects of light and nutrient availability on chasmogamous and cleistogamous flower and fruit production were investigated in an understory tropical herb, Calathea micans (Marantaceae). I censused chasmogamous and cleistogamous flower and fruit production at five permanently marked demographic study sites in Costa Rica, characterized by different successional stages. Using a transplant experiment, I examined whether an increase in light and/or nutrients would increase chasmogamous and/or cleistogamous flower production. In natural populations as well as in the experiment, chasmogamous reproduction increased with light; nutrient availability significantly increased chasmogamous reproduction in the transplant experiment. Field observations indicated that very few plants reproduced and low investment in reproduction might have been due to poor environmental conditions. Larger plants had a higher probability of reproducing, but plant size did not have an effect on the mode of reproduction, chasmogamy vs. cleistogamy. However, the production of new vegetative shoots increased the probability of producing chasmogamous inflorescences. Cleistogamy may allow the plant to reproduce even in conditions of poor habitat quality and assures some seed set even in conditions unfavorable to plant growth.     

Global metabolic responses of NMRI mice to an experimental Plasmodium berghei infection

We present a metabolism-driven top-down systems biology approach to characterize metabolic changes in the mouse resulting from an infection with Plasmodium berghei, using high-resolution (1)H NMR spectroscopy and multivariate data analysis techniques. Twelve female NMRI mice were infected intravenously with approximately 20 million P. berghei-parasitized erythrocytes. Urine and plasma samples were collected 4-6 h before infection, and at days 1, 2, 3, and 4 postinfection. Multivariate analysis of spectral data showed differentiation between samples collected before and after infection, with growing metabolic distinction as the time postinfection progressed. Our analysis of plasma from P. berghei-infected mice showed marked increases in lactate and pyruvate levels, and decreased glucose, creatine, and glycerophosphoryl choline compared with preinfection, indicating glycolytic upregulation, and increased energy demand due to P. berghei infection. The dominant changes in the urinary metabolite profiles included increased levels of pipecolic acid, phenylacetylglycine, and dimethylamine, and decreased concentrations of taurine and trimethylamine- N-oxide, which may, among other factors, indicate a disturbance of the gut microbial community caused by the parasite. Although several of the observed metabolic changes are also associated with other parasitic infections, the combination of metabolic changes and, in particular, the occurrence of pipecolic acid in mouse urine postinfection are unique to a P. berghei infection. Hence, metabolic profiling may provide a sensitive diagnostic tool of Plasmodium infection and the control of malaria more generally.     

Distribution of genetic diversity among disjunct populations of the rare forest understory herb, Trillium reliquum

We assessed genetic diversity and its distribution in the rare southeastern US forest understory species, Trillium reliquum. In all, 21 loci were polymorphic (PS=95.5%) and the mean number of alleles per polymorphic locus was 3.05. However, genetic diversity was relatively low (Hes=0.120) considering the level of polymorphism observed for this outcrossing species. A relatively high portion of the genetic diversity (29.7%) was distributed among populations. There was no relationship between population size and genetic diversity, and we did not detect significant inbreeding. These results are best explained by the apparent self-incompatibility of this species, its longevity and clonal reproduction. To address questions regarding the history of T. reliquum's rarity, we compared results for T. reliquum with that of its more common and partially sympatric congener, T. cuneatum. Despite shared life history traits and history of land use, we observed significant genetic differences between the two species. Although T. cuneatum contains slightly lower polymorphism (Ps=85%), we detected significantly higher genetic diversity (Hes=0.217); most of its genetic diversity is contained within its populations (GST=0.092). Our results suggest that not only is there little gene flow among extant T. reliquum populations, but that rarity and population isolation in this species is of ancient origins, rather than due to more recent anthropogenic fragmentation following European colonization. The Chattahoochee River was identified as a major barrier to gene exchange.