Photosynthetic characteristics of detached barley leaves during greening in the presence of SAN 9785

The effects of the pyridazinone compound SAN 9785 on the photosynthetic competence of leaves, on the photochemical activity of isolated thylakoids and on the formation and spectral properties of chlorophyll-protein complexes were studied during a 72-h greening period of detached etiolated leaves of barley (Hordeum vulgare L. cv. Horpácsi kétsoros). It was established that i) the photosynthetic capacity of the leaves decreased considerably (by 80 and 90%, as determined by¹⁴CO₂ fixation and fast fluorescence induction measurements, respectively); ii) the photochemical activity of isolated thylakoids from water to potassium ferricyanide and from dichlorophenol indophenol/ascorbate to methylviologen exhibited only slight reductions when expressed on a chlorophyll basis compared with the control; iii) the slow fluorescence induction curves of the treated leaves demonstrated the presence of a peculiar fluorescence component interrupting the quenching of fluorescence at around 1 min illumination; iv) a shortage of the chlorophyll-protein complex of photosystem I (CPI) occurred with a higher content of the monomer of the light harvesting complex in the thylakoids of treated leaves; and v) the fluorescence spectrum of the CPI band present in treated leaves indicates the destruction of the structural integrity of this complex during isolation from the membrane.Abbreviations Chl chlorophyll - CPI, CPII chlorophyll-protein complexes of the reaction centres of PSI and PSII - DCMU 3-(3,4-dichlorophenyl)-1,1-dimethylurea - DPIP 2,6-dichlorophenol indophenol - DPIPH₂ chemically reduced form of DPIP - F _o fluorescence of constant yield - F _v fluorescence of variable yield - F _i ,F _m mitial and maximum yield of fluorescence - LHCP³ monomer of the light-harvesting complex - LHCP² and LHCP¹ oligomers of the light-harvesting complex LHCP³ - PSI, PSII photosystems I, II - SAN 9785 4-chloro-5-(dimethylamino)-2-phenyl-3(2H)-pyridazinone, also known as BASF 13-338 - SDS-PAGE sodium dodecyl sulphate-polyacrylamide gel electrophoresis     

On the role of ß-carotene in the reaction center chlorophyll a antennae of photosystem I

Absorption and low temperature fluorescence emission spectra were measured on chloroplast thylakoids and on purified reaction center chlorophyll a-protein complexes of photosystem I, CP-a₁. A clear association between the presence of ß-carotene and the occurrence of far red absorbing and emitting chlorophyll a components of the reaction center antennae of photosystem I was demonstrated. For this study chloroplasts and CP-a₁ were obtained from normal and carotenoid deficient plant material of various sources. The experimental material included 1) lyophilized pea chloroplasts extracted with petroleum ether, 2) the carotenoid deficient mutant C-6E of Scenedesmus obliquus and 3) wheat chloroplasts derived from normal and SAN-9789 treated plants. Removal of carotenoids, most likely principally ß-carotene, caused a loss of long wavelength absorbing chlorophylls in chloroplasts and purified CP-a₁, and the loss or diminution of the long wavelength peak seen in the low temperature fluorescence emission spectrum. This association between ß-carotene and special chlorophyll a forms may explain both the photoprotective and antenna functions ascribed to ß-carotene. In the absence of carotenoids in wheat and in the Scenedesmus mutant, the chlorophyll a antenna of photosystem I was extremely photosensitive. A triplet-triplet resonance energy transfer from chlorophyll a to ß-carotene and a singlet-singlet energy transfer from excited ß-carotene to chlorophyll would explain the photoprotective and antenna functions, respectively. The role of this association in determining some of the fluorescence properties of photosystem I is also discussed.     

Flowering of Cultivated Green and SAN 9789-Treated Chenopodium rubrum Plants Exposed to White,Blue, and Red Light

Green plants and plants devoid of photosynthetic pigments were compared with regard to their ability to flower under various growth conditions. Green plants of Chenopodium rubrum L. and plants treated with norflurazon SANDOZ-9789 (SAN) were grown on sucrose-containing media with or without hormones (GA₃, BA, IAA, ABA) under short-day photoperiodic or continuous illumination with white, blue, or red light. Green and SAN-treated albino plants produced flowers only under short-day conditions. The flowering of green plants was independent of the presence of sucrose and hormones in the medium as well as of the light quality. The albino plants produced flowers under white and blue light but did not flower in red light. The addition of GA₃ or BA to the medium induced flowering of albino plants exposed to red light. The functional interaction of photoreceptors in the flowering control is discussed.     

Soret absorption of intermediate(s) in protochlorophyllide to chlorophyllide photoreduction trapped at low temperature

Absorption spectra at ca 100 K from 400 to 750 nm and fluorescence emission spectra at 77 K from 600 to 750 nm were obtained from: 1) etiolated leaves of the H-ordeum vulgare L. (barley) mutant albozonata 2 and SAN 9789-treated Avena sativa L. (oat) with low levels of carotenoids, and 2) preparations of protochlorophyllide holo-chrome from Phaseolus vulgaris L. cv. Commodore (bean).
This allowed clear resolution for the first time of the Soret bands of the green pigments before and after light-induced accumulation of intermediate(s) in protochlorophyllide to chlorophyllide photoreduction and after conversion of the intermediate(s) to chlorophyllide by warming the samples to 233 K in darkness. Although the intermediate(s) differ(s) in absorption and fluorescence in the red wavelength region from both protochlorophyllide and chlorophyllide, the extinction in the Soret band is not distinguishable from that of chlorophyllide. These observations indicate that the C7-C8 double bond in ring IV of protochlorophyllide has been altered in intermediate(s) accumulated at low temperature in intense light, such that the transition state exhibits the character of a π complex.     

Chlorophyll b accumulation and grana formation in low intensities of red light

Abstract When dark grown leaves of wheat (Triticum aesivum L.) were given a brief irradiation, there was an immediate onset of chlorophyll (Chl) b synthesis in the dark. This synthesis led to a rather slow accumulation of Chl b, which ceased when the Chl b/Chl a ratio had reached a value of about 0.1. The Chl b synthesis occurred also when the seedlings were treated with the herbicide SAN 9789. Leaves grown under different intensities of red light accumulated Chl b and Chl a, resulting in a ratio Chl b/Chl a which depended on the light intensity. If the light intensity was low, Chl a accumulated to a level about ten times the level of PChlide of the dark grown leaves. This occurred without any increase in the Chl b/Chl a ratio. There was no difference between SAN 9789-treated seedlings and water controls in this respect. Above a certain threshold of irradiance, the Chl b/Chl a ratio in the control leaves increased rapidly with the irradiation intensity. The increase in Chl b/Chl a ratio coincided with formation of grana in the plastids. This increase was not found and grana formation was completely absent in the seedlings treated with SAN 9789. The possibility of two different stages in the Chl b synthesis is discussed.     

Chelerythrine and sanguinarine dock at distinct sites on BclXL that are not the classic BH3 binding cleft

The ratio of the levels of pro-survival and pro-apoptotic members of the Bcl-2 protein family is thought to be an important regulatory factor for determining the sensitivity of the mammalian cells to apoptotic stimuli. High levels of expression of pro-survival members such as Bcl(XL) in human cancers were frequently found to be a good prognostic indicator predicting poor response to chemotherapy. The pro-survival members of the Bcl-2 family mediate their effects through heterodimerization with the BH3 region of the pro-apoptotic members. Structural analyses of the binding complex of the BH3 peptide and Bcl(XL) showed that a hydrophobic groove termed the BH3 binding cleft is the docking site for the BH3 region. Chemical mimetics of the BH3 region such as BH3I-1 that target the BH3 binding cleft indeed exhibit pro-apoptotic activities. Chelerythrine (CHE) and sanguinarine (SAN) are natural benzophenanthridine alkaloids that are structurally homologous to each other. CHE was previously identified as an inhibitor of Bcl(XL) function from a high-throughput screen of natural products, but its mode of interaction with Bcl(XL) is not known. By determining the effect of site-directed mutagenesis on ligand binding and using saturation transfer difference (STD) NMR experiments, we have verified locations of these docked ligands. Surprisingly, CHE and SAN bind separately at the BH groove and BH1 region of Bcl(XL) respectively, different from the BH3 binding cleft where other known inhibitors of Bcl(XL) target. Interestingly, certain residues on the flexible loop between helices alpha1 and alpha2 of Bcl(XL) are also perturbed upon CHE, but not SAN or BH3I-1 binding. Although CHE and SAN are similarly effective as BH3I-1 in displacing bound BH3 peptide, they are much more effective in inducing apoptosis, raising the possibility that CHE and SAN might be able to antagonize other pro-survival mechanisms in addition to the one that involves BH3 region binding.     

The Effects of Growth Regulators on Flowering of Chenopodium murale Plants in vitro

In vitro culture of Chenopodium murale L. (ecotype 197) green and herbicide SAN 9789 - treated "white" plants was established and the effects of benzylaminopurine (BAP), indole-3-acetic acid (IAA) and gibberellic acid (GA₃) on growth and flowering were tested. Green plants did not flower on glucose free media, while 17 % of plants flowered on 5 % glucose-containing medium. SAN 9789 (10^–5 M) inhibited growth and flowering. BAP and IAA (0.1 – 5 mg dm^–3) also inhibited growth and flowering of green and "white" plants. GA₃ (10 mg dm^–3) stimulated leaf development in green plants, but had no significant effect on "white" plants, and stimulated flowering of green (41 %) and "white" (33 %) plants.     

Correlation between pigment composition and apoproteins of the light-harvesting complex II (LHC II) in wheat (Triticum aestivum)

Seedlings of wheat (Triticum aestivum L. cv. Walde, Weibull) grown in continuous weak red light (16 mW m⁻²) with or without SAN-9789, contained significantly lower amounts of chlorophylls and carotenoids compared to untreated plants grown in a greenhouse. The Chl alb ratios were 3.6 in the greenhouse-grown plants, 5.1 in untreated and ca 16 in SAN-treated plants grown in weak red light, respectively. The main difference in polypeptide composition of thylakoids isolated from red light-grown plants, compared to those grown in the greenhouse, was a lower amount of proteins of the light-harvesting complex (LHC) II. The amount of apo-LHC and LHC were correlated to the xanthophyll to β-carotene ratios in these plants. The absence of grana and the absence of proteins of the light-harvesting complex 11 in SAN-treated plants, support the general dogma that these proteins are involved in the formation of grana. Since the amount of apo-LHC and LHC could be correlated to the presence of carotenoids as well as the chlorophylls, it is concluded that the carotenoids are necessary for the correct assembly and stabilization of the apoproteins of LHC II in the thylakoid membranes.     

Absence of the Spindle Assembly Checkpoint Restores Mitotic Fidelity upon Loss of Sister Chromatid Cohesion

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