Chromatic Regulation of Photosystem Composition in the Cyanobacterial Photosynthetic System: Kinetic Relationship between Change of Photosystem Composition and Cell Proliferation

Kinetics of the change of photosystem (PS) composition in cyanobacteriainduced by chromatic light were studied in relation to cellproliferation. The study was made for two unicellular strains,Synechococcus NIBB 1059 and Synechocystis (Aphanocapsa) PCC6714. We found that (1) the change to a higher or lower PS I/IIratio was due to acceleration or suppression of apparent PSI formation, and (2) it progressed on a similar time scale tothat of the cell proliferation. The apparent rate constant ofthe change in the PS I/II ratio was proportional to that ofcell proliferation, µ, when this was low, but at highvalues of µ the increase in the rate constant of the changein the PS I/II ratio became smaller, causing a deviation fromthe linear relationship. Results indicate that under autotrophicconditions, the photoregulated composition change occurs asa result of thylakoid development, which accompanies cell proliferation. (Received June 23, 1986; Accepted December 5, 1986)     

Transformation of mouse BALB/c 3T3 cells with human basic fibroblast growth factor cDNA.

The expression of human basic fibroblast growth factor (bFGF) cDNA in mouse BALB/c 3T3 clone A31 cells induced morphological transformation. These transformed cells grew well and reached more than a sixfold-higher saturation density than parental A31 cells even in serum-free medium. They were able to form colonies in soft agar. The phenotypic alteration in the transformed cells was reversed by the addition of anti-human bFGF antibodies to the medium. These results suggest that the cellular transformation mediated by bFGF is caused by autocrine stimulation with secreted bFGF molecules.     

Nudeotide Pools and Purine Metabolism in Tissue Cultures of Wild-Type Datura innoxia and Adenine-Requiring Auxotroph

Cells of the auxotrophic mutant, Ad1, of Datura innoxia requiredadenine, adenosine, or inosine for their growth on solid agarmedium which contained Murashige-Skoog salts, 2,4-dichloro-phenoxyaceticacid, and sucrose. Thirteen purine and pyrimidine nucleotidesin extracts of wild-type and Ad1 cells were separated and quantifiedby HPLC. Levels of ADP-glucose and UMP were significantly higherin Ad1 than in wild-type cells, but those of other nucleotideswas found when Ad1 cells were transferred to fresh medium withoutadenine. The rate of the biosynthesis de novo of purines, asestimated from the rate of incorporation of ¹⁴C from [2-¹⁴C]-glycine and [¹⁴C]formate into adenine nucleotides, was reducedin Ad1 cells to 21 and 13% of the wild-type rate, respectively.The activities involved in the salvage of adenine and adenosinein Ad1 cells were similar to those in wild-type cells. Ad1 cellshad the capability to convert adenine to guanine nucleotidesand guanine to adenine nucleotides. ¹ Part 27 of the series, "Metabolic Regulation in Plant CellCulture". (Received March 7, 1988; Accepted August 3, 1988)     

THE EFFECT OF IRON NUTRITION ON PHOTOSYNTHESIS AND NITROGEN FIXATION IN CULTURES OF TRICHODESMIUM (CYANOPHYCEAE)1

Cultures of Trichodesmium NIBB 1067 were grown in the synthetic medium AQUIL with a range of iron added from none to 5 × 10^?7 M Fe for 15 days. Chlorophyll-a, cell counts, and total cell volume were two or three times higher in medium with 10^?7 M Fe than with no added Fe. Oxygen production rate per chlorophyll-a was over 60% higher with higher iron. Increased iron stimulated photosynthesis at all irradiances from about 12–250 μE · m^?2· s^?1. Nitrogen fixation rate, estimated from acetylene reduction, for 10^?7 and 10^?8 M Fe cultures was approximately twice that of the cultures with no added Fe. The range of rates of O₂ production and N₂ fixation in cultures at the iron concentrations we used were similar to the rates from natural samples of Trichodesmium from both the Atlantic, and the Pacific oceans. This similarity may allow this clone to be used, with some caution, for future physiological ecology studies. This study demonstrates the importance of iron to photosynthesis and nitrogen fixation and suggests that Trichodesmium plays a central role in the biogeochemical cycles of iron, carbon and nitrogen.     

1-Acylglycerophosphocholine O-acyltransferase in maturing safflower seeds

The activity of 1-acylglycerophosphocholine (1-acyl-GPC) O-acyltransferase (EC 2.3.1.23) varied during maturation of safflower (Carthamus tinctorius L.) seeds, and activity per seed was highest in the middle period of seed development when triacylglycerol (TAG) is most rapidly synthesized. The specific activity of acyl transfer in a 20000·g particulate preparation exceeded 500nmol·min^-1·(mg protein)^-1 and was higher than those of any other enzymes involved in TAG synthesis (K. Ichihara et al., 1993, Plant Cell Physiol. 34, 557–566). This suggested the presence of a large flux of acyl-CoA to phosphatidylcholine in the cell. The reaction was specific to C₁₆ and C₁₈ acyl-CoAs with a double bond at position 9. Lauroyl- and erucoyl-CoA were completely ineffective, while ricinoleoyl- and elaidoyl-CoA were utilized efficiently. The relative order of specificity for native acyl-CoA species was linoleoyl > oleoyl stearoyl = palmitoyl. When acyl-CoA mixtures were presented, preference for the unsaturated species rather than the saturated species was even more apparent. The enzyme preferentially utilized 1-C₁₆-acyl- and 1-C₁₈-acyl-GPC molecular species, and 1-palmitoyl-, 1-stearoyl-, 1-oleoyl-and 1-linoleoyl-GPC equally served as acyl acceptor. No activity was detected with 1-octanoyl-GPC, and 1-erucoyl-GPC produced little effect. The effectiveness of 1-alkyl-GPC was comparable to that of 1-acyl-GPC. It was thus concluded that the enzyme recognizes the chain lengths of the acyl donor and acceptor, and the double bond at position 9 of the acyl donor.Abbreviations DAG diacylglycerol - DTNB 5,5-dithiobis(2-nitrobenzoic acid) - GP sn-glycerol 3-phosphate - GPC sn-glycero-3-phosphocholine - GPE sn-glycero-3-phosphoethanolamine - GPI sn-glycero-3-phosphoinositol - PC phosphatidylcholine - TAG triacylglycerol     

Content of iron and copper in the nuclei and induction of pH 9-labile lesions in L5178Y sublines inversely cross-sensitive to H2O2 and x-rays

Cells from the L5178Y murine lymphoma subline LY-R are twice as resistent to killing by ionizing radiation than the subline LY-S. In contrast, LY-R cells are more sensitive to killing by H₂O₂, the effect being more pronounced at 37 °C than 0 °C. Initial DNA damage after H₂O₂ treatment (both temperatures, 5 min) has been estimated by the comet assay (single-cell gel electrophoresis) and fluorescent halo technique. According to both methods, the initial damage is significantly higher in LY-R cells, particularly that inflicted at O °C. Differences between DNA unwinding and rewinding abilities at pH 9 and 6.9 (estimated by the fluorescent halo technique) point to a considerable difference in pH-9-labile damage between the sublines, as observed previously for x-irradiated cells (Kapiszewska et al. 1992). In contrast to findings with x-irradiated cells, however, after H₂O₂ treatment this damage is more extensive in LY-R cells than in LY-S cells. Thus, the initial pH-9-labile damage corresponds to the pattern of sensitivity to H₂O₂ and x-rays. We suggest that this is caused by different proportions of cuprous and ferric ions found in the nuclei of LY sublines and by the different ability of these ions to react with H₂O₂ and water radiolysis products. The copper/iron ratio in the nucleus is 1.31 in LY-R cells and 4.84 in LY-S cells.     

Photoreversible absorption changes of guanidine-HCltreated phycocyanin and allophycocyanin isolated from the blue-green alga Tolypothrix tenuis

In the search for the photoreceptor in photocontrolled phycoerythrinformation, photoreversible absorption changes of chromoproteinsin vivo and in vitro were studied with the blue-green alga Tolypothrixtenuis. Neither intact cells nor crude extracts of soluble proteinsshowed any significant absorption changes which were reversiblyinduced by green and red light. However, the photoresponse wasobservable when the crude protein extracts were treated withthe chaotropic reagent guanidine-HCl (0.4 M, for 1 hr in thedark). Isolated phycocyanin and allophycocyanin also showedthe same photoresponse after the guanidine-HCl treatment. Thedifference spectrum (green minus red) of guanidine-HCl-treatedphycocyanin was almost identical with that shown by phycochromea of Bj?rn and Bj?rn (3), and the allophycocyanin showed thesame difference spectrum as those of phycochrome c of Bj?rnand Bj?rn and the photoreversible pigment isolated by Scheibe(7). Urea at a concentration higher than 1 M or alkaline incubation(pH 8.5) also showed the same effect. The results were interpretedas indicating that phycocyanin and allophycocyanin obtain theability for photoresponsiveness when their protein conformation,probably around the chromophore site, is modified. (Received October 30, 1978; )     

The Pro to glycine mutation of staphylococcal nuclease simplifies the unfolding—folding kinetics

Kinetics of unfolding and refolding of a staphylococcal nuclease mutant, in which Pro¹¹⁷ is replaced by glycine, have been investigated by stopped-flow circular dichroism, and the results are compared with those for the wild-type protein. In contrast to the biphasic unfolding of the wild-type nuclease, the unfolding of the mutant is represented by a single-phase reaction, indicating that the biphasic unfolding for the wild-type protein is caused by cis-trans isomerization about the prolyl peptide bond in the native state. The proline mutation also simplifies the kinetic refolding. Importance of the results in elucidating the folding mechanism is discussed.     

Comparison of cytochrome P-450 species which catalyze the hydroxylations of the aromatic ring of estradiol and estradiol 17-sulfate

For identification of microsomal cytochrome P-450 (P-450) enzymes which catalyze 2- or 4-hydroxylations of estrogens in the rat liver, estradiol (E₂) and estradiol 17-sulfate (E₂-17-S) were selected as the substrates and incubated with various kinds of purified P-450 enzymes: PB-1, PB-2, PB-4 and PB-5 obtained from phenobarbital-treated male rats (Sprague-Dawley); MC-1 and MC-5 from 3-methylcholanthrene-treated male rats; and UT-1, UT-2, UT-4 and UT-5 from untreated animals. The reactions were carried out under the P-450-reconstructed system, and the resulting products were determined by HPLC using electrochemical detection. All the enzymes tested were shown to have varying degrees of catalytic activities for 2-hydroxylation of the two substrates; UT-1 and UT-2 had the highest activity. Of the induced P-450 enzymes, PB-2 and MC-1 showed fairly high catalytic activity for 4-hydroxylation of E₂. The P-450 enzymes obtained from the untreated male rats, especially UT-4, showed the highest catalytic activity for 4-hydroxylation of the two substrates. From these results and also from kinetic experiments, the P-450 enzymes which catalyze 2- and 4-hydroxylations of estrogen were considered to be different species. A part of E₂ was converted to such metabolites as estrone and those having a hydroxyl group at positions 6β, 15 or 16, each production of which was estimated to be catalyzed by single or multiple P-450s.