8-METHOXYPSORALEN-PHOTOINDUCED DNA CROSSLINKS AS DETERMINED IN YEAST BY ALKALINE STEP ELUTION UNDER DIFFERENT REIRRADIATION CONDITIONS. RELATION WITH GENETIC EFFECTS

Abstract— DNA damage induced by 8-methoxypsoralen (8-MOP) plus near UV light (UVA) was analyzed in diploid yeast using the alkaline step elution technique. The presence of 8-MOP and UVA induced DNA interstrand cross-links was revealed by the increase of DNA retained on elution filters as compared to untreated controls. The fraction of DNA retained on filters increased linearly with UVA dose. The amount of cross-links was estimated from the fraction of DNA retained on filters using a dose of -radiation leading to a number of DNA strand breaks at least equivalent to the number of 8-MOP induced photoadducts.
When 8-MOP treated cells were exposed to monochromatic light, 365 nm light induced monoadducts and cross-links whereas 405 nm light induced only monoadducts. When submitting 8-MOP plus 405 nm light treated cells to 365 nm irradiation, after removal of unbound 8-MOP by washing, a portion of 8-MOP plus 405 nm light induced monoadducts was converted into cross-links. The amount of monoadducts transformed into cross-links was dependent on the dose of 365 nm irradiation up to a maximum likely to correspond to the number of suitably positioned furan-side monoadducts that could be converted into biadducts. When 8-MOP plus 365 nm light treated cells were reirradiated with 365 nm light, following the same protocol, the maximum level of cross-linking obtainable in yeast was lower than that obtained with 8-MOP in a 405 nm plus 365 nm reirradiation protocol.
In the presence of 8-MOP single exposures to 405 nm light were found to be only slightly genotoxic. However, when followed by second exposures to 365 nm light, a dose-dependent increase in genetic effects, i.e. mutation and gene conversion, was observed in parallel to the induction of DNA crosslinks. These results stress again the prominent role of DNA cross-links in the genotoxicity of 8-MOP.     

INHIBITION OF DNA SYNTHESIS BY PSORALEN-INDUCED LESIONS IN XERODERMA PIGMENTOSUM AND FANCONI''S ANEMIA FIBROBLASTS

Abstract— Exposure of human cells to psoralens and near-UV light produces a mixture of monoadducts and crosslinks in DNA, which inhibit DNA synthesis by blocking replicon initiation and chain elongation. 8-Methoxypsoralen (8-MOP) has a greater effect than angelicin in normal, xeroderma pigmentosum, and Fanconi's anemia cells. Recovery of DNA synthesis is not detectable up to 8 h after exposure. The average distance between lesions that block replication in individual replicons was measured by means of bromodeoxyuridine photolysis. After exposure to 10 μg/mℓ of 8-MOP and 7500 J/m² of near-UV light, blocks were formed every 20 μm. Replicon initiation was inhibited by exposure to near-UV light alone in normal and xeroderma pigmentosum. Exposure to low concentrations of angelicin or 8-MOP plus near-UV light inhibited replicon initiation in normal and Fanconi's anemia cells, but not in xeroderma pigmentosum cells. Inhibition of initiation was not obvious after treatment with high concentrations of 8-MOP or angelicin because of the dominant effect of crosslinks in blocking chain elongation.     

MUTAGENIC AND RECOMBINOGENIC ACTION OF DNA MONOADDUCTS PHOTOINDUCED BY THE BIFUNCTIONAL FUROCOUMARIN 8-METHOXYPSORALEN IN YEAST (Saccharomyces cerevisiae)

Abstract— For the same furocoumarin 8-MOP and the same total number of photoadditions, the genetic activity of DNA monoadducts and a mixture of mono- and biadducts photoinduced by the bifunctional furocoumarin 8-methoxypsoralen (8-MOP) is compared in the yeast Saccharomyces cerevisiae. In the presence of 8-MOP, 405 nm irradiation induces only monoadducts, whereas 365 nm irradiation induces mono- and biadducts (interstrand cross-links) in DNA. This is shown by heat denaturation-renaturation experiments on calf thymus DNA treated in vitro and by alkaline step elution analysis of DNA from treated yeast cells. For the same photobinding of tritiated 8-MOP to DNA in diploid yeast, about 20 times higher doses are needed with 405 nm than with 365 nm irradiation. Re-irradiation experiments reveal that part of the monoadducts induced by 8-MOP and 405 nm irradiation can be effectively converted into DNA interstrand cross-links by exposures to 365 nm radiation after washing-out of unbound 8-MOP molecules. 8-MOP and 405 nm irradiation induce per lethal hit cytoplasmic "petite" mutations in yeast as efficiently as the monofunctional furocoumarin 3-carbethoxypsoralen (3-CPs) and 365 nm irradiation, both treatments being much more efficient than 8-MOP and 365 nm irradiation. At equal survival, treatments with 8-MOP and 405 nm radiation are clearly less efficient than treatments with 8-MOP and 365 nm radiation for the induction of forward ( CAN *) and reverse ( HIS +) mutations in haploïd yeast and for the induction of mutations ( ILV +) and genetically aberrant colonies including mitotic crossing-over in diploid yeast. The two treatments are equally efficient for the induction of mitotic gene conversion. At equal photobinding of 8-MOP, the monoadducts induced by 405 nm irradiation are found less effective than the mixture of mono-and biadducts induced by 365 nm irradiation for the induction of cell killing, mutations and mitotic recombination.     

New aspects of the repair and genotoxicity of psoralen photoinduced lesions in DNA.

Several approaches are described aiming at a better understanding of the genotoxicity of psoralen photoinduced lesions in DNA. Psoralens can photoinduce different types of photolesions including 3,4- and 4',5'-monoadducts and interstrand cross-links, oxidative damage (in the case of 3-carbethoxypsoralen (3-CPs)) and even pyrimidine dimers (in the case of 7-methylpyrido(3,4-c)psoralen (MePyPs)). The characterization and detection of different types of lesions has been essential for the analysis of their possible contributions to genotoxicity. For example, oxidative damage photoinduced by 3-CPs can be detected by the formamidopyrimidine glycosylase (FPG) protein. Furthermore, it is shown how the presence of MePyPs induced monoadducts may interfere with the photoreactivation of concomitantly induced pyrimidine dimers, how the ratio of monoadducts and interstrand cross-links (CL) affects the occurrence of double-strand breaks during the repair of photolesions and genotoxicity. In vitro treatment of yeast plasmids, followed by transformation, also indicates that the repair of photoadducts on exogenous DNA differs for 8-methoxy-psoralen (8-MOP) induced mono- and diadducts and for monoadducts alone. The recombinational rad52 dependent pathway is not needed for the repair of 8-MOP induced monoadducts. The results obtained suggest that the genotoxic effects of psoralens are conditioned by the nature, number, ratio and sequence distribution of the photolesions induced in DNA.     

THE EFFECT OF DARK COMPLEXING ON THE PHOTOSENSITIZED FORMATION OF 8-METHOXYPSORALEN CROSS-LINKS WITH DNA*

The near-UV dose at 365 nm required for the formation of 8-methoxypsoralen (8-MOP) cross-links with calf thymus DNA was found to be insensitive to the concentration ratio of nucleotides to 8-MOP over the range 100:1 to 5:1. The proposed explanation is that only the dark-complexed 8-MOP contributes to the photosensitized formation of monoadducts and cross-links. A kinetics analysis indicates that the fluence required for cross-linking should be inversely proportional to the square root of the fraction of dark-complexed 8-MOP per nucleotide (the binding ratio), a quantity that is insensitive to the ratio of nucleotides to 8-MOP. These findings predict a small dependence of photosensitivity on the total 8-MOP concentration in cellular systems in which cross-links are the major lethal lesion.     

Near-UV induced interstrand cross-links in anthraquinone-DNA duplexes

Anthraquinone (AQ) has been extensively used as a photosensitizer to study charge transfer in DNA. Near-UV photolysis of AQ induces electron abstraction in oligonucleotides leading to AQ radical anions and base radical cations. In general, this reaction is followed by the transport of base radical cations to sites of low oxidation potential, that is, GG, and conversion of G radical cations to DNA breaks. Here, we show that AQ also produces interstrand cross-links in DNA duplexes. About half of the cross-links collapse to single strands in hot piperidine treatment. The structure of stable interstrand cross-links was deduced by MS, NMR, and sequence substitution. The cross-links consist of a covalent link between the methyl group of T on one strand with either C6 or C7 of AQ on the other strand. The formation of interstrand cross-links decreased in O2 compared to deoxygenated solutions. In the presence of O2, the yield of breaks at GG doublets was 10-fold greater than that of cross-links for end tethered AQ, while cross-links exceeded breaks for centrally located AQ. The formation of stable cross-links can be explained by initial charge transfer from T to excited AQ, deprotonation of T radical cations, and condensation of the latter species with AQ radicals. These studies reveal a novel pathway of damage in the photolysis of AQ-DNA duplexes.     

The Lack of Efficacy of 4,6,4'-Trimethylangelicin to Induce Immune Suppression in an Animal Model for Photopheresis: a Comparison with 8-MOP

Photopheresis is an extracorporeal form of photochemo-therapy with 8-methoxypsoralen (8-MOP) and UVA (PUVA). Patients ingest 8-MOP and then a psoralen-rich buffy coat is obtained by centrifugation and mixed with saline. This mixture is recirculated through a UVA radiation field and then reinfused. Photopheresis appears to be effective for several T cell-mediated disorders, because the treatment results in a specific immune response against the pathogenic clone of T cells involved. With PUVA therapy, the whole body of the patient is exposed to UVA, after ingestion of 8-MOP. Upon UVA exposure 8-MOP binds to, amongst others, DNA and induces DNA monoadducts and interstrand cross-links. As a result of these photoadducts photocarcinogenicity is a risk in PUVA. In PUVA for psoriasis, it proved that angular furocoumarins, although almost incapable of inducing DNA cross-links (less carcinogenic), are still effective. In order to determine if monoadducts induced by photopheresis could also be effective we used, specifically, 4,6,4'-trimethylangelicin (TMA). In this report, we compare the photodegradation of both TMA and 8-MOP under conditions relevant to the in vivo situation, as well as the effect both compounds have on the viability of rat lymphocytes as measured with the 3–(4,5-dimethylthia-zol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay. We show that TMA did not induce immunosuppression in vivo , even after extensive irradiation. In addition a dose dependency of 8-MOPNVA versus the induced immune suppression was carried out. It was shown that there is a log doselresponse correlation of r = 0.9205.     

REPAIR OF 8-METHOXYPSORALEN INDUCED DNA INTERSTRAND CROSS-LINKS IN Tetrahymena thermophila

Abstract— The protozoan Tetrahymena thermophila was treated with 8-methoxypsoralen in combination with long wavelength ultraviolet irradiation and the DNA-repair response was studied. Following the treatment a lag-period in cell proliferation was observed, the duration of which was proportional to the amount of psoralen used, and both swelling and deformation of the cells were observed. The treatment with 3 μg 8-methoxypsoralen/mℓ and a light dose of 8 kJ m^-2 resulted in a 10-fold decrease in DNA and RNA synthesis activity, while the protein synthesis was only moderately affected. Using the same conditions the lag-period was 30 h, and during this time the psoralen induced DNA interstrand cross-links were removed. Alkaline elution experiments showed that the repair process involves DNA single strand scissions, whereas no double strand DNA scissions were detected.     

DNA damage induced by 4,6,8,9-tetramethyl-2H-furo[2,3-h]quinolin-2-one, a new furocoumarin analog: biological consequences

4,6,8,9-Tetramethyl-2H-furo[2,3-h]quinolin-2-one (HFQ) and its isomer FQ (1,4,6,8-tetramethyl-2H-furo[2,3-h]quinolin-2-one) showed very strong antiproliferative activity in mammalian cells, about two times greater than 8-methoxypsoralen (8-MOP). Both compounds induced DNA-protein cross-links (DPC) but not interstrand cross-links. The FQ generated DPC in a biphotonic process, yielding a new kind of diadduct, whereas HFQ induced DPC by a monophotonic one, probably without its physical participation in the covalent bridge. These lesions gave different toxic responses. Sensitization of FQ led to extensive DNA fragmentation and to a number of chromosomal aberrations. Conversely, HFQ seemed to be completely inactive and 8-MOP gave intermediate results. A strict relationship between DPC formation and induction of chromosomal aberrations was observed. The HFQ did not induce light skin erythemas, whereas FQ was more phototoxic than 8-MOP, thus suggesting that FQ lesions, DPC in particular, may be implicated in skin phototoxicity. Ehrlich ascites cells, a transplantable mouse tumor, inactivated by furoquinolinone sensitization and injected into healthy mice, protected them from a successive challenge by viable tumor cells. This response appeared to be based on an immune mechanism. Comparable amounts of base substitution revertants were scored when testing furoquinolinones and 8-MOP in bacteria but no DPC were detected. This suggests that classic mutagenesis tests on bacteria are insufficient to give adequate information on furocoumarin genotoxicity. Given its features, HFQ can be regarded as an interesting new agent for psoralen plus UVA photochemotherapy and photopheresis.     

REPAIR OF 8-METHOXYP SORALEN PHOTOINDUCEDCROSS-LINKS and MUTAGENESIS: ROLE OF THE DIFFERENT REPAIR PATHWAYS IN YEAST

Abstract— In Saccharomyces cerevisiae, a re-irradiation with a saturing dose of UVA after pretreatment with 8-methoxypsoralen (8-MOP) plus low doses of UVA and removal of unbound 8-MOP lead to an increase in frequency of forward mutants in strains defective in the excision (radl-3, radl-Δ, rad2-6) or in the recombinational (rad52-l) repair pathways. Such an enhancement attributable to DNA interstrand cross-links was not observed in mutants blocked in a mutagenic repair pathway (rad6-Δ and pso2-l). These results are interpreted as revealing the existence of an alternative pathway to excision of DNA cross-links. This pathway appears to be error-prone and independent from the recombinational pathway. The RAD6 or the PSO2 gene products are likely to interfere with this process.     

Photochemistry of nucleic acids in cells.

A survey of the recent aspects of the main photoreactions induced by far-UV radiation in cellular DNA is reported. This mostly includes the formation of cyclobutadipyrimidines, pyrimidine(6-4)pyrimidone photoadducts and related Dewar valence isomers in various eukaryotic and prokaryotic cells, as monitored by using either specific or more general assays. Information is also provided on mechanistic aspects regarding the formation of 5,6-dihydro-5-(alpha-thyminyl) thymine, the so-called "spore photoproduct" within far-UV-irradiated bacterial spores. The second major topic of the review deals with the effects of near-UV radiation and visible light on cellular DNA which are mostly mediated by photosensitizers. The main photoreactions of furocoumarins with DNA, one major class of photosensitizers used in the phototherapy of skin diseases, involve a [2 + 2] cycloaddition to the thymine bases according to an oxygen-independent mechanism. In contrast a second type of photosensitized reaction which appears to play a major role in the genotoxic effects of both near-UV and visible light requires the presence of oxygen. The photodynamic effects which are mediated by either still unidentified endogenous photosensitizers or defined exogenous photosensitizers lead to the formation of a wide spectrum of DNA modifications including base damage, oligonucleotide strand breaks and DNA-protein cross-links.     

4, 4'', 5''-TRIMETHYL-8-AZAPSORALEN, A NEW-PHOTOREACTIVE AND NON-SKIN-PHOTOTOXIC BIFUNCTIONAL BIOISOSTER OF PSORALEN

Photochemical and photobiological properties of a new isoster of psoralen, 4,4',5'-trimethyl-8-azapsoralen (4,4',5'-TMAP), have been studied. This compound shows a high DNA-photobinding rate, higher than that of 8-methoxypsoralen (8-MOP), forming both monoadducts and inter-strand cross-links. The yield of cross-links, however, is markedly lower than that of 8-MOP. Antiproliferative activity of 4,4',5'-TMAP, in terms of DNA synthesis inhibition in Ehrlich ascites tumor cells, is higher than that of 8-MOP. Mutagenic activity on E. coli WP2 R46+ cells appeared similar to or even lower than that of 8-MOP. This new compound applied on depilated guinea pig skin and irradiated with UVA did not show any skin-phototoxicity. On the basis of these properties 4,4',5'-TMAP appears to be a potential photochemotherapeutic agent.     

SENSITIVITY OF DNA REPAIR-DEFICIENT STRAINS OF ESCHERICHIA COLI K-12 TO VARIOUS FUROCOUMARINS AND NEAR-ULTRAVIOLET RADIATION

Abstract— Survival curves were obtained for DNA repair-deficient strains of Escherichia coli K-12 ( polA1, uvrB5 , and recA56 ) exposed to near-ultraviolet radiation [black light (BL)] in the presence of the DNA cross-linking agent 8-methoxypsoralen (8-MOP) or in the presence of photosensitizers forming primarily monoadducts with DNA [angelicin; 3-carbethoxypsoralen (3-CPs); 5,7-dimethoxycoumarin (DMC)], and after exposure to blue light (BluL) in the presence of 8-MOP or 3-CPs. An interpretation of these data suggests that DNA polymerase I is required for the major pathway of monoadduct repair, but appears to play little or no role in the repair of 8-MOP cross-links. The uvrB and recA strains were very sensitive, both to the cross-linking agent and to the monoadduct formers. The markedly different results for BL plus DMC or 3-CPs compared to angelicin suggests that the DMC and 3-CPs monoadducts are repaired by a different mechanism than are the angelicin monoadducts, or else DMC and 3-CPs undergo photochemical side reactions that produce DNA lesions other than the expected monoadducts. From photochemical evidence, we predicted that fewer 8-MOP monoadducts should be converted to cross-links by BluL vs BL; this appears to be the case. 3-CPs showed dramatically different biological results when irradiated with BL vs BluL, suggesting that 3-CPs may form more types of photoproducts than the expected monoadducts; BluL, however, appears to favor monoadduct formation.     

Molecular dosimetry of 8-MOP + UVA-induced DNA photoadducts in Saccharomyces cerevisiae: correlation of lesions number with genotoxic potential

Acid hydrolysis of purified DNA extracted from cells of a haploid repair-proficient (RAD) yeast strain that had been treated with 8-MOP + UVA revealed the existence of two major and one minor thymine photoproduct. At survival levels of the RAD strain between 100% and 1% furanside monoadducts constituted the major DNA lesion, followed by diadducts that, at the lowest survival level, nearly reached 50% of the thymine photoproducts; pyrone-side monoadducts were only detectable at the highest UVA exposure dose applied and clearly constitute a minority photoproduct. The number of induced diadducts was verified by determination of interstrand cross-links via denaturation and renaturation of 8-MOP + UVA-treated DNA from RAD and rad2 yeast strain. 8-MOP + UVA was shown to induce two types of locus-specific mutations: reversion of the lys1-1 ochre allele was between 20- to 50-fold higher than that of the his4-38 frameshift allele. Mutant yield for the lys 1-1 reversion was the same in RAD and excision repair-deficient rad2-20 strains whereas frameshift mutagenesis was about eightfold higher in the rad2-20 background.     

DNA damage induced by 4,6,8,9-tetramethyl-2H-furo[2,3-h]quinolin-2-one, a new furocoumarin analog: photochemical mechanisms

Some photochemical and photobiological properties of 4,6,8,9-tetramethyl-2H-furo[2,3-h]quinolin-2-one (HFQ) were studied in comparison with its isomer 1,4,6,8-tetramethyl-2H-furo[2,3-h]quinolin-2-one (FQ) and 8-methoxypsoralen (8-MOP). The HFQ photobinds to DNA forming furan-side monoadducts (MAHFQ) that have molecular structure very similar to those of FQ (MAFQ). Unlike MA8-MOP and MAFQ, MAHFQ no longer photoreact. The HFQ, like FQ, produces moderate amounts of singlet oxygen but no superoxide anions. The HFQ and FQ induce numbers of DNA-protein cross-links (DPC), much more plentiful than those of 8-MOP (about two and seven times, respectively) but no interstrand cross-links. The mechanism of DPC formation was studied in vivo in mammalian cells by alkaline elution and in vitro using a new test mixing histones and DNA from calf thymus. The latter is a very useful technique for the double irradiation protocol. The DNA (or histones) are separately exposed to a first UVA dose in the presence of the sensitizer; then, after its unbound molecules have been removed, histones (or DNA) are added to assemble the chromatin-like complex that is irradiated again. According to in vitro and in vivo methods, DPC appear to be formed by FQ and 8-MOP by a biphotonic process that starts with monoadduct induction in DNA, followed by their conversion into DPC. In the resulting lesions, the sensitizer molecule forms a covalent bridge between the two macromolecules (DPC at length greater than zero). Instead, HFQ induces DPC by a monophotonic process; thus, HFQ is probably not a physical part of the bridge between DNA and proteins, which may be linked together directly, like DPC at zero length induced by UVC.     

A NEW MONOFUNCTIONAL PYRIDOPSORALEN: PHOTOREACTIVITY and REPAIR IN YEAST

Abstract— The newly synthesized derivative of psoralen, the pyrido (3,4-c) 7 methylpsoralen (MePyPs), acts in combination with 365 nm ultraviolet as a monofunctional agent on yeast DNA. In vivo, its photoaffinity for DNA is much higher than that of the bifunctional agent, 8-methoxypsoralen (8-MOP). The MePyPs photo-induced monoadducts are almost completely removed from wild type cells DNA as efficiently as 8-MOP photo-induced adducts during post-treatment incubation. This process is blocked in excision-repair defective mutants (6 to 10% residual excision in radl- Δ or rad2- Δ ). For an equal number of photoinduced lesions, the DNA single strand breaks which are produced concomitantly to MePyPs or 8-MOP photoadducts excision are rapidly rejoined in the case of 8-MOP whereas they are only partly resealed for the MePyPs treatment. The high photo-toxicity of MePyPs, a promising agent for photo-chemotherapeutic use, is explained in terms of the high photoaffinity for DNA.     

Mechanism of action of 4-hydroxymethyl-1,6,8-trimethylfuro[2,3-h]quinolin-2(1H)-one, a very active angular furocoumarin-like sensitizer

The molecular structure of 1,4,6,8-tetramethylfuro[2,3-h]quinolin-2(1H)-one (FQ), a recent furocoumarin-like photosensitizer, has been modified with the aim of reducing its strong genotoxicity, by replacing the methyl group at 4 position with a hydroxymethyl one, and so obtaining 4-hydroxymethyl-1,6,8-trimethylfuro[2,3-h]quinolin-2(1H)-one (HOFQ). This modification gave rise to a strong reduction of lipophilicity and dark interaction with DNA. The formation of monoadducts (MA) was deeply affected, whereas the induction of bifunctional adducts between DNA and proteins (DPC(L>0)) was replaced by an efficient production of DNA-protein cross-links at zero length (DPC(L=0)), probably via guanine damage. Because of its angular molecular structure, HOFQ does not form interstrand cross-links (ISC): therefore, DPC(L=0) and MA represent the main lesions induced by HOFQ in DNA. In comparison with FQ (which induces MA and DPC(L>0)) and 8-methoxypsoralen (8-MOP) (MA, ISC, DPC(L>0)), HOFQ seems to be a more selective agent. In fact, contrary to FQ and 8-MOP, HOFQ, together with a noticeable antiproliferative activity, shows low levels of point mutations in bacteria and of clastogenic effects in mammalian cells. HOFQ is also an efficient apoptosis inducer, especially in comparison with 8-MOP, when tested at equitoxic experimental conditions; this property might be correlated with the complete HOFQ inability of inducing skin erythemas, a well-known side effect of classic furocoumarin photosensitization.     

A PHOTOSYSTEM I-PHENOSAFRANINE SOLAR CELL

Abstract Bacteriophage λ_vir was inactivated when it was irradiated with near-UV light in the presence of chlorpromazine. DNA strand breakage in the treated phage was indicated by alkaline sucrose gradient centrifugation. The number of the breaks was increased with increasing fluence. Although the inactivation rate was enhanced with a decreasing salt concentration in the reaction mixture and under a nitrogen atmosphere, the number of the strand breaks was not altered in either case. Therefore, the DNA strand breakage is not a sole lethal damage in the treated phage. The addition of NaN₃ repressed the inactivation and the reaction in a D₂O medium enhanced the inactivation even if the reaction mixture was irradiated under anaerobic conditions. Under anaerobic conditions, the inactivation occurs presumably via a radical mechanism.     

INDUCTION OF DNA-PROTEIN CROSS-LINKS IN CHINESE HAMSTER CELLS BY THE PHOTODYNAMIC ACTION OF CHLOROALUMINUM PHTHALOCYANINE AND VISIBLE LIGHT

Abstract— Chloroaluminum phthalocyanine (CAPC) is an efficient photosensitizer for the inactivation of Chinese hamster V79 cells. In order to investigate possible molecular mechanisms in the photo-dynamic action of CAPC and visible light, the induction and repair rate of two classes of DNA lesions have been determined, i.e. DNA single-strand breaks and DNA-protein cross-links. In cells pretreated with 1 μ.M CAPC, a fluence of 12 kJ/m² of red light (>600 nm) kills approximately 50% of the cells and induces 3 to 3.5 Gy-equivalents of single-strand breaks. The repair of these breaks was slower than the repair of single-strand breaks induced by -irradiation. The photodynamic action of CAPC also induces a large number of DNA-protein cross-links which, in contrast to -radiation-induced DNA-protein cross-links, do not appear to be repaired during 4 h of post-treatment incubation in fresh medium. These studies suggest that DNA may be an important target for the cytotoxicity of CAPC + red light.     

Decreased survival of UV-irradiated Staphylococcus aureus in the presence of 8-methoxypsoralen in the post-irradiation plating medium

For Staphylococcus aureus, the presence of 8-methoxypsoralen (8-MOP) in the post-irradiation plating medium increased the lethal effect of far-UV light (FUV; approximately 254 nm) and of 8-MOP plus near-UV light (8-MOP+NUV; approximately 365 nm), an effect similar to that caused by acriflavine which inhibits DNA repair. In the repair-proficient strain, the presence of 8-MOP in the plating medium was almost as effective in inhibiting the repair of damage caused by FUV as that caused by 8-MOP photoadditions. Survival data obtained with Rec(-)-like and Uvr(-)-like strains suggest that 8-MOP in the plating medium, although possibly inhibiting recombination repair, was much more effective in inhibiting excision repair of FUV damage. Regarding 8-MOP+NUV treatment, 8-MOP in the plating medium had a lesser effect in the repair-deficient strains, differing from that observed after FUV treatment, which is consistent with the notion that different types of damage are caused by the two treatments.