Pill-Soon Song

PHOTOCHEMISTRY AND PHOTOBIOLOGY OF PSORALENS

Abstract— The photochemistry and photobiology of psoralens have been rcvicwed on the basis of selected literature. In addition to the well establishcd photoaddition of psoralens to DNA. photoreac‐tions involving RNA. tRNA and proteins warrant further studies. Although thymine seems to be one of the predominant bases for the psoralen photocycloaddition in DNA. photoreactions of other bases such as adenine and the mechanisms other than cycloaddition should also be investigated in order to fully assess the specificity of psoralen photoreactions and their application BS a probe for biopolymer structure and as a tool for photobiological studies of microorganisms. The relativc photobiological importance of monoaddition vs crosslinking of psoralens to nucleic acids also remains to be fully elucidated.

Hypericin and its photodynamic action

Hypericin, 7,14-dione-1,3,4,6,8,13-hexahydroxy10,ll-dimethyl-phenanthrol[ 1 , l O ,9,8opgralperylene, derives its name from Hypericum, a genus of plants which biosynthesize the pigment and its related derivatives and stored in minute glands located in different organs. Hypericin constitutes up to 0.05% of the plant substances in some species of the genus (Netien and Lebreton, 1964). St. John’s wort, H. perforatum, is probably the richest source of hypericin (Giese, 1980a; Georgiev et al., 1983; Vanhaelen and Vanhaelen-Fastre, 1983). Hypericum hirsutum also contains a large amount of hypericin (Kitanov et al., 1978; Know and Dodge, 1985). Hypericin and closely related pigments have also been found in some Australian insects (Banks et al., 1976). Hypericin-like pigments are present in the protozoa Blepharisma and Stentor (Giese, 1980a,b; 1981) and in Fagopyrum esculentum (Wender et al., 1943; Wender, 1946). Hypericism, a state of skin sensitivity to the visible light in animals, is apparently caused by the ingestion of hypericin-containing plants and feed. The photosensitizing effects of skin by hypericin result in severe skin irritation, high body temperature and sometimes death of the animal. Hypericin subcutaneously injected in rats or mice induces the typical photodynamic effect on body temperature when incandescent light is applied (Brockmann et a/., 1939; Chick and Ellinger, 1941; Brockmann et al., 1942; Pace 1942; Brockmann, 1947,1952; Metzner, 1958; Bwangamol, 1967; Giese, 1972).

Functional Characterization of Phytochrome Interacting Factor 3 in Phytochrome-Mediated Light Signal Transduction

Phytochromes regulate various light responses through their interactions with different signaling proteins, such as phytochrome interacting factor 3 (PIF3). However, the physiological functions of PIF3 in light signaling are not yet fully understood. To increase our understanding of these roles, we characterized a T-DNA insertional pif3 mutant and transgenic plants overexpressing the full-length PIF3. Transgenic overexpressing lines displayed longer hypocotyls and smaller cotyledons under red light and reduced cotyledon opening under both red and far-red light, whereas the pif3 mutant showed the opposite phenotypes. The accumulation of anthocyanin and chlorophyll further indicated complicated features of PIF3 function. The accumulation of anthocyanin was increased and the content of chlorophyll was decreased in the overexpression lines. Our data indicate that PIF3 plays complex roles depending on the type of light response and the light conditions.

Spectroscopic characterization of β-lactoglobulin-retinol complex

1. 1.|The absorption spectrum of retinol when bound to β-lactoglobulin is vibrationally resolved. The circular dichroism spectrum exhibits the same structure, as does the fluorescence excitation spectrum. 2. 2.|Two molecules of retinol are bound per protein dimer, with a binding constant (Kd) of 2 · 10−8 M. Also, by fluorescence titration it was found that the monomer binds one molecule of retinol with essentially the same Kd. 3. 3.|Energy transfer occurs from tryptophan (donor) to retinol (acceptor) with a rate constant, k, of 4.4 · 108 s−1. The distance between the centers of mass of the transition is 34 A, corresponding to the energy transfer efficiency of 44%. 4. 4.|The fluoresence lifetime of retinol increases dramatically on binding to β-lactoglobulin, from approx. 2 to approx. 10 ns, as does the fluorescence quantum yield. 5. 5.|The retinol binding to β-lactoglobulin does not show a pH dependence and the binding site is hydrophobic. 6. 6.|On the Sephadex G-100 column, retinol is chemically modified to a retro derivative which binds even more strongly to β-lactoglobulin than does retinol. 7. 7.|The β-lactoglobulin-retinol complex rotates anisotropically in solution with a fast (3 ns) and a slower (12 ns) component. This may be attributed to retinol being bound at a flexible region of the protein, where only segmental flexibility is observed, weighted by its proximity to one of the major axis rotational times.

Light and Brassinosteroid Signals Are Integrated via a Dark-Induced Small G Protein in Etiolated Seedling Growth

Plant growth and development are regulated through coordinated interactions between light and phytohormones. Here, we demonstrate that a dark-induced small G protein, pea Pra2, regulates a variant cytochrome P450 that catalyzes C-2 hydroxylation in brassinosteroid biosynthesis. The cytochrome P450 is dark-induced and predominantly expressed in the rapidly elongating zone of etiolated pea epicotyls, where Pra2 is also most abundant. Transgenic plants with reduced Pra2 exhibit a dark-specific dwarfism, which is completely rescued by exogenous brassinolide. Overexpression of the cytochrome P450 results in enhanced hypocotyl growth even in the light, which phenocopies the etiolated hypocotyls. We therefore propose that Pra2 and its orthologs are molecular mediators for the cross-talk between light and brassinosteroids in the etiolation process in plants.

The photochemistry and photobiology of furocoumarins (Psoralens)

Publisher Summary This chapter describes the photochemistry and photobiology of furocoumarins and the applications of furocoumarins in biology and medicine. Furocoumarins and their congeners are found in many plants. The photoreactivity of psoralens regarding the cycloaddition to DNA is determined by three main factors: kinetic, steric, and electronic. RNA can also be photochemically modified with psoralens and near-UV radiation. When psoralens are intercalated in DNA, the singlet reaction can be a predominant path, because the singlet excited psoralen intercalated need not diffuse too far to encounter its substrate. The probability of the photocycloaddition of psoralen to thymine in solution is enhanced if the excited singlet state of the psoralen encounters thymine and forms an exciplex. In mammalian cells, PUVA-induced mutagenesis is linear with dose, following one-hit kinetics. PUVA treatment is a well-established clastogen. Caffeine enhances the yield of chromosome aberrations when added after PUVA treatment. PUVA treatment inactivates transforming DNA in Bacillus subtilis. DNA cross-links contribute to this effect by preventing entry of DNA into the bacterial cells.

Phytochrome-Specific Type 5 Phosphatase Controls Light Signal Flux by Enhancing Phytochrome Stability and Affinity for a Signal Transducer

Environmental light information such as quality, intensity, and duration in red (approximately 660 nm) and far-red (approximately 730 nm) wavelengths is perceived by phytochrome photoreceptors in plants, critically influencing almost all developmental strategies from germination to flowering. Phytochromes interconvert between red light-absorbing Pr and biologically functional far-red light-absorbing Pfr forms. To ensure optimal photoresponses in plants, the flux of light signal from Pfr-phytochromes should be tightly controlled. Phytochromes are phosphorylated at specific serine residues. We found that a type 5 protein phosphatase (PAPP5) specifically dephosphorylates biologically active Pfr-phytochromes and enhances phytochrome-mediated photoresponses. Depending on the specific serine residues dephosphorylated by PAPP5, phytochrome stability and affinity for a downstream signal transducer, NDPK2, were enhanced. Thus, phytochrome photoreceptors have developed an elaborate biochemical tuning mechanism for modulating the flux of light signal, employing variable phosphorylation states controlled by phosphorylation and PAPP5-mediated dephosphorylation as a mean to control phytochrome stability and affinity for downstream transducers.

REP1, a basic helix-loop-helix protein, is required for a branch pathway of phytochrome A signaling in Arabidopsis.

Phytochromes are primary photoreceptors mediating diverse responses ranging from induction of germination to floral induction in higher plants. We have isolated novel recessive rep1 (reduced phytochrome signaling 1) mutants, which exhibit a long-hypocotyl phenotype only under far-red light but not under red light. Physiological characterization showed that rep1 mutations greatly reduced a subset of phytochrome A–regulated responses, including the inhibition of hypocotyl elongation, cotyledon expansion, modulation of gravitropic growth of hypocotyl, and induction of the CAB (encoding chlorophyll a/b binding protein) gene, without affecting the accumulation of anthocyanin, far-red-preconditioned blocking of greening, induction of germination, and induction of CHS (encoding chalcone synthase) and FNR (encoding ferredoxin-NADP+ oxidoreductase) genes. These results suggest that REP1 is a positive signaling component, functioning in a branch of the phytochrome A signaling pathway. Molecular cloning and characterization of the REP1 gene revealed that it encodes a light-inducible, putative transcription factor containing the basic helix-loop-helix motif.

PHOTO‐CONJUGATION OF 8–METHOXYPSORALEN WITH PROTEINS

Abstract— 8–Methoxypsoralen (8–MOP) is shown to form a covalent conjugate with bovine serum albumin (BSA) by UVA irradiation in the presence of 02. The photoreaction is shown to involve oxidation of 8–MOP itself as a first step, producing an oxidation product which reacts readily with protein. Thus, this photoreaction appears to be completely different from the known 8–MOP photo‐cycloaddition reaction with DNA. Among several proteins studied, human serum albumin, histone type 11, RNAse A and lysozyme also undergo photoinduced addition by 8–MOP. Chemical modifications of various amino acid residues in BSA revealed tyrosine‐OH as one of the reaction sites. Irradiation (12 h) with UVA at 1.78 J/ds intensity resulted in approximately 1.5 mot of 8–MOP bound to 1 mol of BSA.

Luminescence spectra and photocycloaddition of the excited coumarins to DNA bases.

Abstract— The lowest excited singlet and triplet states of coumarin, psoralen, and 4‐hydroxy‐coumarin have been assigned to the (π,π*) type on the basis of the luminescence spectroscopy and MO calculations. The mechanism of photocycloaddition of courmarin and psoralen to thymine has been described in terms of the perturbational MO model and MO reactivity indices. All possible cycloaddition patterns have been examined. Results suggest that the 3,4‐bond of coumarin in the excited state is somewhat more reactive than the same bond of psoralen in the excited state. It is also predicted that the 3,4‐bond of psoralen in the triplet state is more reactive than the 4′, 5′‐bond. The results have been favorably correlated with the electronic characteristics of excited coumarin molecules and with available experimental data on the relative yields of photoadducts.