PETITE INDUCTION IN YEAST, Saccharomyces cerevisiae, BY PHOTOACTIVATION OF 3-A-ZIDO-6-A-MINO-10-M-ETHYLACRIDINIUM CHLORIDE

Abstract— The photoinduction of petite colonies and cell toxicity in non-growing yeast, Saccharomyces cerevisiae , by 3-a-zido-6-a-mino-10-m-ethylacridinium chloride (AAMAC) has been examined. The results presented here indicate that mitochondrial DNA damage in resting yeast which occurs following irradiation of AAMAC-treated cells for short time periods is probably mediated through a covalent adduct between AAMAC and DNA. Furthermore, the photoreaction which contributes to biological activity is dependent on the presence of oxygen. Pre-irradiated AAMAC, which no longer exhibited the short-term photo-induction of biological effects showed a second biological activity. In this case longer irradiation times, e.g. 30 min, were required to induce petites for resting yeast. Again there was a strong dependence on the presence of oxygen. These results suggest that both processes may be effected through oxygen intermediates (photodynamic processes).     

Synthesis and characterization of ethidium analogs: Emphasis on amino and azido substituents

A series of eight ethidium derivatives has been synthesized in which the substituents at R₃ and R₈ have been varied with hydrogen, azido and amino functions. Three of these compounds are new and their synthesis and characterization by uv-visible, ir and ¹H nmr spectroscopy are presented. The synthesis and characterization of the other compounds are also given for comparison, because the compounds served as precursors, or the synthetic route undertaken for these compounds differed from that reported previously.     

Computing inertia sets using atoms

We consider the problem of computing inertia sets for graphs. By using tools for combining the inertia sets of smaller graphs we can reduce this problem to understanding the inertia sets for three-connected graphs that are not joins. We term such graphs atoms and give the inertia sets for all atoms on at most seven vertices. This can be used to compute the inertia sets for all graphs on at most seven vertices.     

UNSCHEDULED DNA SYNTHESIS IN FROG LENS AT 5°C

Abstract. Unscheduled DNA labeling occurs in the frog even at low temperatures. We conclude, tentatively, that the UV-induced labeling observed in these cold incubated lenses represents repair synthesis of DNA.     

Crystal structure of ethidium monoazide,a photoactive compound that reacts with nucleic acids

The crystal and molecular structure of ethidium monoazide, a photoactive derivative of ethidium, was determined from X-ray diffraction data. Crystals of 8-azidoethidium chloride monohydrate are monoclinic, space groupP2₁/c, witha = 8.336(5),b = 14.778(6),c = 16.873(5) Å, and = 99.90(3) ° The crystals are poorly ordered, weakly diffracting, and unstable in air. Intensity data were collected with an automated diffractometer from a crystal that was sealed in a thin-walled glass capillary. The structure was solved by direct methods and was refined by least squares toR = 0.20 for the complete set of 2126 unique reflections, and toR = 0.11 for 724 reflections with1 > 2(I). The results demonstrate that the photoactive azide moiety is attached to the 8-position of the phenanthridinium moiety. This azide group is almost linear, is nearly in the plane of the phenanthridinium ring, and is oriented away from the phenyl substituent (i.e.,trans to the C(7)-C(8) bond). Possible models for intercalation complexes of 8-azidoethidium with double-helical nucleic acids were examined by modifying published coordinates from crystal structures of ethidium-dinucleotide complexes. When in the conformation observed in this crystal structure, 8-azidoethidium can be substituted for ethidium in the published complexes without suffering any unacceptable steric interactions.     

REPAIR SYNTHESIS IN HUMAN LYMPHOCYTES PROVOKED BY PHOTOLYSIS OF ETHIDIUM AZIDE

Abstract— The azide analog of ethidium was mixed with human lymphocytes and when photolyzed with visible light provoked repair synthesis as shown by incorporation of tritiated thymidine in the presence of hydroxyurea. The use of photolyzed drug, or incubation of drug-cell mixtures in the dark was without effect. These experiments should prove useful in targeting drug action sites and in studying the details of DNA repair.     

DNA EXCISION REPAIR IN LYMPHOCYTES FOLLOWING PHOTOAFFINITY LABELING WITH ETHIDIUM MONOAZIDE

Irradiation of ethidium monoazide by fluorescent light promotes a chemical decomposition of the azide into a highly reactive nitrene intermediate. Covalent bonding of this electrophile to the DNA in the cell provokes repair of damage which can be monitored by incorporation of [³H]-thymidine. Human lymphocytes were labeled with [¹⁴C]-ethidium azide and then allowed to undergo DNA repair. Repair incorporation of [³H]-thymidine showed saturation at 5 µM ethidium azide, but excision of the labeled drug failed to saturate at 20 µM, suggesting that excision and resynthesis are two separate events. Cells were also labeled with the photosensitive drug and/or exposed to UV radiation, and then allowed to undergo a period of DNA repair. The tritium incorporation for the combined insults was less than the sum of the two insults. Quinacrine, progesterone and chloroquine inhibited repair incorporation of [³H]-thymidine, but had no effect on the excision of the drug from the DNA. After damage by ethidium azide, chromatin was isolated from lymphocytes which had been allowed to repair label with [³H]-thymidine. Partial digestion of the chromatin with micrococcal nuclease released 80% of the tritium when approximately 40% of the DNA had been hydrolyzed by the enzyme.