Prothoracicotropic Hormone Bioassay: Pupal-Adult Bombyx Assay

Blockage of adult development by brain removal and its resumption by application of the prothoracicotropic hormone (PTTH) were studied using pupae of a racial hybrid J-122 × C-115 of Bombyx mori . A log-linear dose-response relationship was obrained after injection of a PTTH solution. The Bombyx -unit of PTTH has been defined from this dose-response curve.     

Studies on the change of the respiratory system in roots in relation to the growth of plants V. Effect of root pruning terminal oxidase activity in roots of cereal crops

This experiment studies the comparative activities of terminaloxidases (Fe-containing oxidases, Cu-containing oxidases andCN-resistant oxidases) in roots of rice and wheat seedlingsand the effect of root pruning on these terminal oxidases inthe roots of those crops. Data indicate that (i) excepting achange in the activity of Cu-containing oxidases no appreciabledifference in respiration or in Fe-containing and CN-resistantoxidases was noticed in the two types of seedling roots and(ii) root pruning did not produce any noticeable change in terminaloxidase activities in roots of wheat seedlings. A drastic changein terminal oxidase activities in roots of rice seedlings wasobserved, suggesting that a close relationship exists betweenCN-resistant oxidase and transplanting time. (Received November 21, 1968; )     

Some properties of shikimate: NADP oxidoreductase produced in sweet potato root tissue after slicing

The activity of shikimate: NADP oxidoreductase [EC 1. 1. 1.25] in sweet potato root tissue increased soon after slicing.Enzyme preparations obtained from both sliced tissue and fromfresh tissue probably contained a single enzyme component, andthey showed identical chromatographical behaviour. K_m values of the enzyme for NADP and shikimate were 1.0x10^–4Mand 1.3 x 10^–3M, respectively. Enzyme activity was potentlyinhibited by SH-inhibitors such as p-chloromercuribenzoate andoxidized glutathione. Enzyme activity was affected neither by mononucleotides suchas ATP, ADP and AMP, divalent cations, Mg⁺⁺, Ca⁺⁺ and Mn⁺⁺,nor by metabolites such as tryptophan, phenylalanine, tyrosineand t-cinnamic acid which are involved in aromatic compoundsyntheses. The enzyme rapidly lost its activity. This inactivation reactionshowed a time course consisting of two steps of the first-orderreaction. The inactivated enzyme preparation was not reactivatedby thiol compounds such as cysteine, 2-mercaptoethanol and glutathione,although these reagents, to a certain extent, protected theenzyme from inactivation. The results suggest that denaturationof the enzyme protein was involved in inactivation of the enzyme. ¹Part 74 of the phytopathological chemistry of sweet potatowith black rot and injury. ²Present address: Department of Biology, Faculty of Science,Tokyo Metropolitan University, Setagaya-ku, Tokyo. (Received August 5, 1968; )     

The Hill reaction and oxygen uptake in isolated pine chloroplasts

1. The Hill reaction and oxygen uptake in chloroplasts preparedfrom pine leaves were studied. Pine chloroplasts active forthe Hill reaction were isolated from mature leaves in the presenceof 25% PEG in the isolation buffer. Time courses of the Hillreaction in chloroplasts isolated from leaves at different seasonsdiffered. In chloroplasts isolated in the autumn, Hill activitydecreased rapidly with illumination time. This rapid decreaseof Hill activity was inferred to result from concomitant productionof some inhibitory substance(s) during the Hill reaction.
2. Theprotective effect of PEG on inactivation by aging of pinechloroplastswas found. In the presence of PEG, chloroplastswere stabilizedand Hill activity was maintained even afterstorage for 26 hr;whereas, in the absence of PEG inactivationby aging proceededrapidly and oxygen uptake occurred after20 hr.
3. Chloroplastsisolated without PEG had no ability of the Hillreaction; but,inversely showed pronounced oxygen uptake. Oxygenuptake wasalso observed in aged or DCMU-inhibited chloroplasts.The presenceof benzoquinone strongly suppressed oxygen uptake.

¹ Present address: Laboratory of Biology and Chemistry, Universityof Naval Architecture of Nagasaki, Nagasaki, Japan. (Received January 27, 1971; )     

Electron flow from hydrogen peroxide in photosystem II-catalyzed oxidation-reduction reactions of spinach chloroplast fragments

Oxidation-reduction reactions of photosystem II were investigatedin spinach chloroplast fragments. Chloroplast fragments treatedwith 8-hydroxyquinoline sulfate showed only a low activity forthe 2,6-dichlorophenolindophenol (DPIP) Hill reaction, as wasobserved in chloroplast fragments treated with a high-concentrationof Tris buffer. Hydrogen peroxide could donate electrons tophotoreaction center II in chloroplast fragments treated with8-hydroxyquinoline, high-concentration Tris, or ethylene glycol,but water could not serve as an electron donor in these preparations.Electrons from hydrogen peroxide were transferred to DPIP, ferricyanide,and p-benzoquinone viaphotosystem II. (Received May 12, 1971; )     

Reactions of photosystems I and II in spinach chloroplast fragments treated with ethylene glycol

Photochemical reactions of chloroplast fragments isolated fromspinach leaves were measured in the presence of ethylene glycolor were measured after washing with an ethylene glycol-containingmedium. 2,6-Dichlorophenolindophenol (DPIP) photoreduction,oxygen evolution and oxygen uptake (a photosystem I reaction)were investigated in ethylene glycol-treated chloroplast fragments.By washing with ethylene glycol, oxygen evolution was stronglyinhibited, but oxygen uptake was not much affected by ethyleneglycol washing. Chloroplast fragments in 50% ethylene glycolmaintained a high rate of DPIP photoreduction (85% of the controlactivity in an ethylene glycol-less medium). In 67% ethyleneglycol, DPIP photoreduction mediated by photosystem II was eliminatedand only a small rapid reduction mediated by photosystem I wasobserved. Chloroplast fragments inhibited by ethylene glycolphotoreduced DPIP in the presence of p-aminophenol added asan artificial electron donor to photosystem II. The restoredactivity of DPIP photoreduction was inhibited by 3-(3',4'- dichlorophenyl)-1,1-dimethylurea. (Received September 8, 1970; )     

INHIBITORY EFFECT OF GROWTH RETARDANTS ON THE INDUCTION OF FLOWERING IN WINTER WHEAT

1. Growth retardants, CCC, Amo-1618, Phosfon-D and B-995, appliedduring seed vernalization inhibited the ear development of winterwheat. CCC applied during green plant vernalization inhibitedflowering,but it had no appreciable effect on the final stemlength. Theinhibition by CCC was reversed by foliar applicationof gibberellin.On the other hand, CCC applied after the vernalizationperiodaffected the final stem length but not flowering.
2. Theamount of endogenous gibberellin-like substance(s) was greaterin the vernalized plant than in the non-vernalized plant. Nogibberellin-like substance was detected in the CCC-treated plant.
3. Endogenous gibberellin-like substance(s), whose biosynthesisis inhibited by some growth retardants, may play a part in thevernalization process in winter wheat.

¹Present address: National Institute of Agricultural Sciences,Nishigahara, Kitaku, Tokyo     

Micromere Differentiation in the Sea Urchin Embryo: Two-Dimensional Gel Electrophoretic Analysis of Newly Synthesized Proteins

A method for large-scale culture of isolated blastomeres of sea urchin embryos in spinner flasks was developed. Micromeres and meso-, macromeres isolated from sea urchin embryos at the 16-cell stage were cultured by this method and the patterns of protein synthesis by their descendants were examined by two-dimensional gel electrophoresis of [³⁵S] methionine-labeled proteins. Six distinct proteins with molecular weights of 140–kDa, 105–kDa, 43–kDa, 32–kDa, and 28–kDa (two components) were specifically synthesized by differentiating micromeres. Quantitative analysis of the two-dimensional gel patterns demonstrated that all these proteins, except the 32–kDa protein, appeared at the time of ingression of primary mesenchyme cells (PMC's) in vivo , several hours earlier than the onset of spicule formation. The synthesis of 32–kDa protein was paralleled to active spicule formation and the uptake of Ca²⁺. Cell-free translation products directed by poly (A)⁺ RNAs isolated from descendant cells of micromeres and meso-, macromeres were compared by two-dimensional gel electrophoresis. Several spots specific to the micromere lineage were detected. However, none of them comigrated with the proteins synthesized specifically by the cultured micromeres. The results suggest that the expression of these proteins specific to differentiating micromeres may involve post-translational modification.     

GENERAL PATTERN, 35SO4-INCORPORATION AND INTRACELLULAR LOCALIZATION OF GLYCANS IN DEVELOPING SEA URCHIN (ANTHOCIDARIS) EMBRYOS

Glycosaminoglycan (GAG) prepared from sea urchin embryos ( Anthocidaris crassispina ) at various stages with or without pulse ³⁵SO₄-labelling was separated into various fractions by chromatography on DEAE-cellulose with a linear NaCl concentration gradient: fraction "P" (nonacidic) and fractions "A" through "F" (of increasing acidities). The ³⁵SO₄-radioactivity was negligible in "P" and "A", largest in "B" and "C", and decreased in the other fractions three alphabetical order. During development (hatched blastulae to gastrulae) the glycans in fractions "P" and "A" decreased in amount, whereas those in "E" and "F" increased. "E" contained heparin-like (AMPS-1) and dermatanpolysulfate-like (AMPS-2) GAG in addition to a sulfated fucogalactan-like (E₁) glycan. Another sulfated fucogalactan-like (F₁) glycan was found in "F". A sulfated polysialic acid-like (S₁) glycan was found in "C". An EDTA-extract of gastrulae gave AMPS-2, E₁ and F₁. The mitochondria-rich fraction gave AMPS-1, whereas the yolk granule-rich fraction gave S₁. Most of the other still unidentified components in "B", "C", and "D" appeared to be derived from glycoproteins and were mainly located in the crude yolk-mitochondrial and cytosol fractions.     

SYNCHRONOUS MASS-CULTURE OF CHLORELLA

A new culture apparatus was constructed to obtain large quantitiesof synchronized cells of Chlorella ellipsoidea. Two flat culturechambers made of lucid acrylate resin (each 20 liters in capacity)were placed in a water thermostat together with a bank of 17daylight fluorescent lamps, and the culture was run by the methodstarting from a homogeneous population of D_s-cells accordingto TAMIYA et al. It was demonstrated that with this apparatusone can obtain as much as 200–400 mg (dry weight) eachof algal material at 8 or 9 different stages in one cell cycle.Completely synchronized cell cycles could be repeated as manyas 10 times in one series of experiment, indicating that theapparatus can produce 2–4g (dry weight) each of algalcells of different developmental stages. (Received April 2, 1964; )