Deneb Karentz

The Induction and Repair of DNA Photodamage in the Environment

Since its discovery in bacteria 25 years ago, DNA repair, particularly in response to damage induced by UV radiation, has been a major focus of molecular biology. The roles played by DNA damage and repair in the aging and carcinogenic processes have been of particular interest. More recently, degradation of stratospheric ozone and the resultant increase in UV-B radiation at the earth’s surface have focused our attention on the environmental effects of solar DNA damage. Although we have learned much about DNA damage tolerance mechanisms in prokaryotes and some lower eukaryotes, our understanding of the molecular events that determine biological effects of UVR in complex higher eukaryotes is far from complete.

Influence of Ozone-Related Increases in Ultraviolet Radiation on Antarctic Marine Organisms1

SYNOPSIS. Every spring for the past two decades, depletion of stratospheric ozone has caused increases in ultraviolet B radiation (UVB, 280–320 nm) reaching Antarctic terrestrial and aquatic habitats. Research efforts to evaluate the impact of this phenomenon have focused on phytoplankton under the assumption that ecosystem effects will most likely originate through reductions in primary productivity; however, phytoplankton do not represent the only significant component in ecosystem response to elevated UVB. Antarctic bacterioplankton are adversely affected by UVB exposure; and invertebrates and fish, particularly early developmental stages that reside in the plankton, are sensitive to UVB. There is little information available on UV responses of larger Antarctic marine animals (e.g., birds, seals and whales). Understanding the balance between direct biological damage and species-specific potentials for UV tolerance (protection and recovery) relative to trophic dynamics and biogeochemical cycling is a crucial factor in evaluating the overall impact of ozone depletion. After more than a decade of research, much information has been gathered about UV-photobiology in Antarctica; however, a definitive quantitative assessment of the effect of ozone depletion on the Antarctic ecosystem still eludes us. It is only obvious that ozone depletion has not had a catastrophic effect in the Antarctic region. The long-term consequences of possible subtle shifts in species composition and trophic interactions are still uncertain.

Sublethal effects of ultraviolet b radiation on miracidia and sporocysts of Schistosoma mansoni: intramolluscan development, infectivity, and photoreactivation.

Schistosoma mansoni occurs in tropical regions where levels of ultraviolet B (UVB; 290-320 nm) light are elevated. However, the effects of UVB on parasite transmission are unknown. This study examines effects of UVB on the miracidia and sporocysts of S. mansoni, focusing specifically on intramolluscan development, infectivity, and the ability to photoreactivate (repair DNA damage using visible light). Histology revealed that miracidia irradiated with 861 J·m−2 underwent abnormal development after penetrating Biomphalaria glabrata snails. Total number of sporocysts in snail tissues decreased as a function of time postinfection (PI), among both nonirradiated and irradiated parasites; however, this decrease was greater in the latter. Moreover, whereas the proportion alive of nonirradiated sporocysts increased PI, that of irradiated sporocysts, i.e., derived from irradiated miracidia, decreased. Irradiation of miracidia with UVB resulted in decreased prevalence of patent infection (defined by presence of daughter sporocysts) in a dose-dependent manner, and no infections occurred at a dose of 861 J·m−2. Like many aquatic organisms, including the snail host, parasites were able to photoreactivate if exposed to visible light following UVB irradiation, even subsequent to penetrating snails. These photoreactivation results suggest cyclobutane–pyrimidine dimers in DNA as the primary mechanism of UVB damage, and implicate photoreactivation, rather than nucleotide excision, as the main repair process in S. mansoni.

Repair-Deficient Xeroderma Pigmentosum Cells Made UV Light Resistant by Fusion with X-Ray-Inactivated Chinese Hamster Cells

Xeroderma pigmentosum (XP) is an autosomal recessive human disease, characterized by an extreme sensitivity to sunlight, caused by the inability of cells to repair UV light-induced damage to DNA. Cell fusion was used to transfer fragments of Chinese hamster ovary (CHO) chromosomes into XP cells. The hybrid cells exhibited UV resistance and DNA repair characteristics comparable to those expressed by CHO cells, and their DNA had greater homology with CHO DNA than did the DNA from XP cells. Control experiments consisted of fusion of irradiated and unirradiated XP cells and repeated exposure of unfused XP cells to UV doses used for hybrid selection. These treatments did not result in an increase in UV resistance, repair capability, or homology with CHO DNA. The hybrid cell lines do not, therefore, appear to be XP revertants. The establishment of these stable hybrid cell lines is an initial step toward identifying and cloning CHO DNA repair genes that complement the XP defect in human cells. The method should also be applicable to cloning genes for other diseases, such as ataxia-telangiectasia and Fanconis anemia.

Mutation and expression of the XPA gene in revertants and hybrids of a xeroderma pigmentosum cell line

A series of ultraviolet (UV)-resistant cell lines have been generated from a UV-sensitive XP group A cell line homozygous for a stop codon (TGA) in the chromosome 9XPA gene. Three lines generated by chemical mutagenesis acquired the ability to excise (6–4) photoproducts but not cyclobutane dimers from the whole genome; two lines generated by a fusion procedure with hamster cells acquired the ability to excise both (6–4) photoproducts and cyclobutane dimers from the whole genome. A central region of the hamsterXPA gene was cloned and sequenced. With the use of species-specific primers in the polymerase chain reaction, we found that the hybrid cell lines do not contain a hamsterXPA gene. Sequence analysis showed that all of the UV-resistant cell lines contain reversions of the human stop codon, resulting in missense mutations (glycine or leucine for arginine) or wild-type sequences. The concentration of XPA protein in revertant cell lines was about one-half that in normal cells, which would be expected from heterozygous cells; there was no evidence that the mutant proteins were less stable than the wild-type proteins. These results are consistent with the idea that the XPA protein initiates repair by binding to damaged sites with various affinities, depending on the photoproduct and the transcriptional state of the region. A concentration of XPA protein near 50% is needed before repair can proceed into nontranscribed regions of the genome. The revertant cell lines represent a class of missense mutations in theXPA gene that may have altered specificity and that can be used to understand some of the regulatory differences in repair of photoproducts in various regions of the genome.

Prevention of Ultraviolet Radiation Damage in Antarctic Marine Invertebrates

One aspect of investigating the ecological effects of ozone depletion on marine communities is to identify species-specific characteristics relative to strategies for dealing with prevention and repair of ultraviolet B (UV-B, 280–320 ran) damage. In adult marine invertebrates protection from UV-B exposure can be provided by either external coverings (e.g., teste, shell, body wall) or by the presence of UV-absorbing molecules (e.g., mycosporine-like amino acid compounds) or by both in cells and tissues. External coverings are not opaque to UV-B. Using biological dosimetry, penetration of UV-B wavelengths through the exteriors of adult invertebrate has been observed. In addition, selective partitioning of UV-absorbing compounds occurs in adult invertebrate tissues. The highest concentrations of these substances are found in ovaries and eggs. Planktonic larval stages receive higher UV exposures than benthic adults and have considerably less optical shielding. Since egg cells tend to have higher concentrations of UV-absorbing compounds than other body tissues, this may provide for increased protection of embryos and larvae during planktonic development.

Ozone depletion and UV-B radiation in the Antarctic—Limitations to ecological assessment

Abstract Ozone depletion has been detected in the upper atmosphere over the Antarctic since the late 1970s. This depletion results in increased fluences of incident ultraviolet-B radiation in Antarctic environments. There is considerable information available about the chemical and physical characteristics of this atmospheric phenomenon, but the ecological impact of this annual pollution cycle has not been determined. Only a few studies on the UV-photobiology of Antarctic organisms have been initiated, and neither the extent of ecosystem modification nor the time scale for ecosystem response are known.

Distribution of mycosporine-like amino acids in the sea hare Aplysia dactylomela: effect of diet on amounts and types sequestered over time in tissues and spawn

We investigated the interaction of diet and accumulation of UV-absorbing mycosporine-like amino acids (MAAs) in body tissues and spawn of the sea hare Aplysia dactylomela to determine if MAA accumulation reflects type and level of dietary intake. Food sources were the red algae Acanthophora spicifera, Centroceras clavulatum, and Laurencia sp., and the green alga, Ulva lactuca. Adults were maintained on these foods for 40 days, after which feces were collected and tissues separated by dissection. Field animals were similarly sampled at this time. All spawn from experimental and field animals was collected over the study period. Samples, including seaweed foods, were analysed for six MAAs. Overnight consumption experiments using a variety of common seaweeds and one seagrass from A. dactylomelas habitat showed that the four seaweeds selected as foods were among those best-eaten by Aplysia. After 40 days levels of specific MAAs in the tissues of experimental animals showed excellent correlation with those in their diets, suggesting that the MAAs were dietarily-derived. Relative MAA contents in spawn from all diet groups correlated well with those in spawn from field animals. Commonest MAAs in spawn were porphyra-334, shinorine, and palythine, in this order. Concentrations of these MAAs were maintained at constant levels over time in spawn from all diet groups eating red algae and from field animals. Spawn from the Ulva dietary group showed an initial significant decline in MAA concentrations, but levels stabilized after the first 2 weeks. Skin was rich in porphyra-334 and shinorine, and levels of these in experimental animals correlated well with comparable levels in the skin of field animals. Digestive glands contained high levels of asterina-330, particularly those of the Centroceras dietary group, where concentrations reached a maximum of 21 mg dry g(-1).

Beyond Xeroderma Pigmentosum: DNA Damage and Repair in an Ecological Context. A Tribute to James E. Cleaver†

The ability to repair DNA is a ubiquitous characteristic of life on Earth and all organisms possess similar mechanisms for dealing with DNA damage, an indication of a very early evolutionary origin for repair processes. James E. Cleavers career (initiated in the early 1960s) has been devoted to the study of mammalian ultraviolet radiation (UVR) photobiology, specifically the molecular genetics of xeroderma pigmentosum and other human diseases caused by defects in DNA damage recognition and repair. This work by Jim and others has influenced the study of DNA damage and repair in a variety of taxa. Today, the field of DNA repair is enhancing our understanding of not only how to treat and prevent human disease, but is providing insights on the evolutionary history of life on Earth and how natural populations are coping with UVR‐induced DNA damage from anthropogenic changes in the environment such as ozone depletion.

Effects of UVB on interactions between Schistosoma mansoni and Biomphalaria glabrata.

Ultraviolet B (UVB, 280-315nm) radiation is detrimental to both of larvae of the digenetic trematode Schistosoma mansoni and its snail intermediate host, Biomphalaria glabrata. We explored effects of UVB on three aspects of the interaction between host and parasite: survival of infected snails, innate susceptibility and resistance of snails to infection, and acquired resistance induced by irradiated miracidia. Snails infected for 1 week showed significantly lower survival than uninfected snails following irradiation with a range of UVB intensities. In contrast to known immunomodulatory effects in vertebrates, an effect of UVB on susceptibility or resistance of snails to infection could not be conclusively demonstrated. Finally, exposure of susceptible snails to UVB-irradiated miracidia failed to induce resistance to a subsequent challenge with nonirradiated miracidia, a result similar to that reported previously with ionizing radiation.