Physiological Responses of Phytoflagellates to Dissolved Organic Substrate Additions. 1. Dominant Role of Heterotrophic Nutrition in Poterioochromonas malhamensis (Chrysophyceae)

Algal heterotrophy is a potentially important considerationin the flow of carbon through aquatic food webs. The physiologicalresponses to organic compound additions under various lightintensities were examined with Poterioochromonas malhamensis,a freshwater chrysophyte with an exceptionally high heterotrophiccapability. P. malhamensis demonstrated a much greater potentialfor heterotrophic growth than for photoautotrophic growth. Whenorganic substrates (glucose, glycerol, or ethanol) were addedto the culture medium, the growth rate of P. malhamensis significantlyincreased while the chlorophyll content cell ^–1 decreased,even at light intensities saturating for photoautotrophic growth.After an initial decline in chlorophyll production caused byorganic substrate uptake, chlorophyll cell¹ increased and theuptake rate of organic substrates decreased, despite the persistenceof a relatively high substrate concentration in the medium.The results are consistent with the production of substance(s)by P. malhamensis that conditioned the culture medium, leadingto a relief of the inhibitory effect of organic substrates onchlorophyll production. (Received October 29, 1990; Accepted May 8, 1991)     

Effect of light treatment and endogenous growth hormones on {alpha}-and {beta}-amylase activities in cotyledons of Phaseolus vulgaris L.

The influence of light and endogenous plant-growth regulatorson the evolution of - and ß-amylases in cotyledonsof Phaseolus vulgaris L. was investigated. Both enzymes, whichare not present in ungerminated seeds, appear during germinationof intact seedlings or incubation of excised cotyledons. -Amylaseactivity decreases upon exposure to light. This inhibition iscorrelated with a drastic increase in chlorophyll content anda change in the endogenous gibberellin-inhibitor balance. ß-Amylaseactivity was not affected by light treatment but was by thepresence of endogenous cytokinins. (Received February 3, 1977; )     

Phytoplankton photosynthesis parameters in central Canadian lakes

Values for two biological parameters — (i) P^B_m, the lightsaturated rate of photosynthesis per unit of chlorophyll and(ii) , the slope of the light limited part of the photosynthesisversus light curve per unit of chlorophyll — must be knownin order to be able to estimate rates of phytoplankton primaryproduction from chlorophyll data. These parameters were measuredfor periods of up to 9 years in central Canadian lakes locatedin temperate, subarctic and arctic climatic zones. Regardlessof their geographic locations, lakes where integral photosynthesiswas nutrient limited had lower values of these parameters thandid lakes where integral photosynthesis was light limited. Temperatureset an upper limit to the variability of P^B_m but was not a goodpredictor of its actual value. Year-to-year variability of photosynthesisparameters in the most intensively studied group of lakes waslarge: annual means varied by a factor of three over a 9-yearperiod. Until the sources and extent of this variability areknown, accurate production estimates can be obtained from chlorophylldata only if P^B_m and are measured in each water body and inevery year. Implications for estimating primary production fromremotely-sensed chlorophyll data are discussed.     

Circadian photosynthetic activity of natural marine phytoplankton isolated in a tank

A large (1200 1) seawater sample from the Gulf of Maine waskept under constant temperature and light conditions for a periodof 72 h. Circadian variations were observed in the photosyntheticcapacity (P^B_max) and efficiency (^B); these occurred in phase,and were not related to changes in chlorophyll concentrations.Such observations are consistent with the hypothesis that oscillationsin photosynthetic characteristics of natural phytoplankton aremediated by an endogenous circadian rhythm. ¹ Bigelow Laboratory contribution 87023 and a contribution tothe programs of GIROQ (Groupe interuniversitaire de recherchesoceanographiques du Québec) and of the Maurice-LamontagneInstitute     

The Gamete Pigments of Hormosira banksii (Phaeophyta)

The gamete pigments of Hormosira banksii have been separatedby thin-layer chromatography and identified by absorption spectrophotometry.Male gametes contain predominantly ß-carotene, andfemale gametes predominantly chlorophyll and fucoxanthin. Severalminor pigments have also been tentatively identified.     

CHANGES IN EMISSION SPECTRA OF PHOTOSYNTHETIC PIGMENTS IN VIVO

1. The effects of 3-(4'-chlorophenyl)-1, 1-dimethylurea (CMU)onthe fluorescence of photosynthetic pigments in vivo wereinvestigatedin blue-green, red and brown algae and in isolatedspinach chloroplasts.CMU caused an increase in steady statelevel of fluorescenceof chlorophyll a, but did not influencethe fluorescence ofphycobilins. The spectrum of the fluorescenceincrement hada peak at 685 m/µ and a shoulder at 730–740mµ.These two bands probably arise from chlorophyll a(C_f684) belongingto pigment system II.
2. On excitation of chlorophylla in a red alga, Porphyra yezoensis,a fluorescence band witha peak at 720 mµ was observedbesides a shoulder at 685mµ. The 720 m band is inferredto arise from chlorophylla (probably, C_f-1) in pigment systemI.
3. On addition of CMUto the algal cells, the induction of fluorescencewas modifiedto take a simple time course. The induction wasobserved onlywith respect to the fluorescence of chlorophylla, but not inthe fluorescence of phycobilins. The spectrumof the "transient"fluorescence showed two emission bands ofchlorophyll a at 685mµ and 740 mµ, and was quitesimilar in form tothe spectrum of the CMU-caused increase insteady state fluorescence.
4. These facts were interpreted in terms of the correlation offluorescence of chlorophyll a and the photochemical reactionsof photosynthesis

(Received July 20, 1967; )     

Effects of Metal Chelators on Leaf Senescence in Maize and Hydrangea

The senescence of maize and hydrangea leaves after being detachedand kept in the dark was studied in terms of the loss of chlorophyll.Chlorophyll was more rapidly degraded in maize than hydrangeaduring the incubation period in the dark. The loss of chlorophyllin the dark was effectively inhibited in both plants by ,'-dipyridyland o-phenanthroline at concentrations between 0.01 and 0.1mM. Three other chelators of iron produced lesser inhibitionand only at higher concentrations. EDTA prevented the loss ofchlorophyll in maize leaves at concentrations above 10 mM, butdid not do so in hydrangea leaves. Detached leaves floated on EDTA, ,'-dipyridyl or o-phenanthrolinesolutions and exposed to light exhibited a marked bleaching.The bleaching was partially inhibited by applying ascorbic acid. (Received December 26, 1981; Accepted October 18, 1982)     

Novel role for alphavbeta5-integrin in retinal adhesion and its diurnal peak

_v5-Integrin is the sole integrin receptor at the retinal pigment epithelium (RPE)-photoreceptor interface and promotes RPE phagocytic signaling to the tyrosine kinase Mer tyrosine kinase (MerTK) once a day in response to circadian photoreceptor shedding. Herein we identify a novel role for _v5-integrin in permanent RPE-photoreceptor adhesion that is independent of _v5's function in retinal phagocytosis. To compare retinal adhesion of wild-type and ₅-integrin^–/– mice, we mechanically separated RPE and neural retina and quantified RPE protein and pigment retention with the neural retina. Lack of _v5-integrin with normal expression of other RPE integrins greatly weakened retinal adhesion in young mice and accelerated its age-dependent decline. Unexpectedly, the strength of wild-type retinal adhesion varied with a diurnal rhythm that peaked 3.5 h after light onset, after the completion of phagocytosis, when integrin signaling to MerTK is minimal. Permanent _v5 receptor deficiency attenuated the diurnal peak of retinal adhesion in ₅-integrin^–/– mice. These results identify _v5-integrin as the first RPE receptor that contributes to retinal adhesion, a vital mechanism for long-term photoreceptor function and viability. Furthermore, they indicate that _v5 receptors at the same apical plasma membrane domain of RPE cells fulfill two separate functions that are synchronized by different diurnal rhythms. circadian rhythm; knockout; photoreceptors; retinal pigment epithelium     

Chlorophyll-Carotenoid Protein Complexes from the Diatom, Phaeodactylum Tricornutum: Spectrophotometric, Pigment and Polypeptide Analyses

Chloroplast membranes of the diatom Phaeodactylum tricornutumwere dissociated by means of mild detergents, digitonin andlauryl-dimethylamide oxide. After electrophoresis on polyacrylamidegel, all the pigments were still attached to proteins, and sixpigment-protein complexes were obtained. Three Chl -ß-carotenecomplexes had the characteristics of Photosystem I and PhotosystemII antennae. The other three contained Chl , Chl c and fucoxanthinand were related to light-harvesting complexes. Chlorophyllide, a product of the hydrolysis of Chl by chlorophyllase, hasbeen found only in light-harvesting complexes. The spectralproperties, pigment and polypeptide compositions of the complexesare discussed in relation to those of other plants and to theparticular organization of the chloroplasts of diatoms. (Received December 20, 1986; Accepted April 14, 1987)     

Properties of light-harvesting chlorophyll a/b-protein, and photosystem I chlorophyll a-protein, purified from digitonin extracts of spinach chloroplasts by isoelectrofocusing

A procedure for purifying both light-harvesting chlorophylla/b-protein and photosystem I chlorophyll -protein from digitoninextracts of spinach chloroplasts is described. This procedureuses isoelectrofocusing on Ampholine at the last step and permitsisolating all of the chlorophyll-proteins from the same extractin a better yield and a highly pure state. The purified light-harvesting chlorophyll a/b-protein whichhas an isoelectric point (pi) of 4.35 (?0.1) and a single polypeptideof 24 kilodaltons (kD), shows slightly higher chlorophyll a/Aratio of 1.35 than the values reported for the preparationsobtained by anionic detergents. This chlorophyll-protein exhibitsa markedly high and sharp fluorescence band at 681 nm at 77?Kwhich is not found on the chloroplast emission spectrum. Photosystem I chlorophyll a-protein focuses on Ampholine intotwo bands with pi values of 4.75 (?0.1) and 4.80 (?0.1). Thesetwo fractions show the same absorption spectra (maximum at 678nm at room temperature) and emission spectra (maximum at 734nm at 77?K) and have the same constituent polypeptides: onelarge band at 55–64 kD and six minor bands (21.5, 20,19, 18, 16 and 15 kD). The polypeptide composition and the P-700to chlorophyll a ratio (1 to ca. 80) of this preparation arevery similar to those of the photosystem I reaction center preparationobtained from Swiss chard chloroplasts by Bengis and Nelson(8). (Received October 31, 1978; )     

Enzymatic Degradation of Chlorophyll in Chenopodium album

The breakdown of chlorophyll (Chi) in crude extracts of Chenopodiumalbum (white goose foot) in the dark was examined. Derivativesof pheophorbide were formed when Chi or chlorophyllide wasincubated with depigmented crude extracts. The formation ofpheophorbide was completely prevented by heat treatment of extracts,indicating that the reaction was enzymatic, and the presenceof a Mg-releasing enzyme, the so called Mg-dechelatase, waspostulated. This hypothesis was strongly supported by the observationthat the formation of pheophorbide was inhibited by 51% by 10mM MgCl₂. Analysis by high-performance thin-layer chromatography(HPTLC) and liquid chromatography (HPLC) showed that the appearanceof chlorophyllide , pheophorbide 13²-hydroxychlorophyllide and pyropheophorbide was accompanied by a concomitant decreasein levels of Chi The formation of 13²-hydroxychloro-phyllide was not clearly an enzymatic reaction and requires furtherexamination. It appears that Chl is degraded in a crude extractof C. album via the following enzymatically catalyzed reactions (Received September 10, 1990; Accepted November 15, 1990)     

An NTP-Binding Protein That Cross with a Ras-Specific Antibody in the Myceia of Neurospora crassa

A Ras-related NTP-binding protein was partially purified froma membrane fraction derived from the mycelia of Neurospora crassa.[-³²P]ATP and [-³²P]GTP were incubated with mem brane and solublefractions which were then irradiated with UV light to inducecrosslinking of tightly bound nucleotides. After SDS-polyacrylamidegel electrophoresis, blotting onto a nitrocellulose filter andautoradiography it was apparent that most of the proteins thatbound [-³²P]-GTP also bound [-³²P]ATP. Pretreatment of the membranefraction with Ras-specific antibody effectively blocked thebinding of [-³²P]ATP and [-³²P]GTP to several ATP-GTP-bindingproteins. The band of a protein with a molecular weight of 26kDa on the SDS-polyacrylamide gel cross-reacted strongly withthe Ras-specific antibody. The protein was extracted from thegel and further purified by repeated gel electrophoresis. Thepurified protein bound [-³²P]ATP, [-³²P]-GTP, [-³²P]CTP and[-³²P]UTP at 1.6x10^– M and was autophosphorylated in thepresence of [-³²P]ATP and [-³²P]GTP at 1.7x10 M. Pretreatmentof the protein with Ras-specific antibody partially blockedthe autophosphorylation in the presence of these nucleotides.The binding of [-³²P]ATP to the NTP-binding protein was blockedby addition of ATP at 10^–4–10^–3 M. ATP ata concentration of 10^–4 M prevented the binding of [-³²P]to a greater extent than did GTP at the same concentration.Binding of [-³²P]CTP and [-³²P]UTP to the protein was also observed. (Received October 7, 1991; Accepted July 14, 1992)     

A Transient Stimulation of Net Photosynthesis of Rye Leaves by {alpha}-Hydroxy-2-pyridinemethanesulphonic Acid ({alpha}-HPMS) due to Inhibition of Photorespiratory CO2 Release

The effects of -hydroxy-2-pyridinemethanesulphonic acid (-HPMS)upon net photosynthesis (P_n, the CO₂ compensation point (),post-lower illumination burst of CO₂ (PLIB) and post-lower temperatureburst of CO₂ (PLTB) in detached rye (Secale cereale L.) leaveswere investigated. At low concentrations ( 0.5 mol m^–3),-HPMS initially stimulated P_n and decreased the magnitude ofboth PLIB and PLTB. The decreased at all concentrations of-HPMS (0.05–5.0 mol m^–3. The effects of -HPMS onP_n and were time-dependent and, after a few minutes, the P_nwas inhibited while values increased considerably. At a higherconcentration (5.0 mol m ^–3), the transient effects of-HPMS were shorter () or not observed at all (P_n. Both PLIBand PLTB, when expressed in relation to P_n, increased at higherlevels of this compound. Similar data with respect to the effectsof -HPMS on PLIB and PLTB were found for leaves of dandelion(Taraxacum officinale L.). The results suggest that -HPMS may stimulate P_n by inhibitingphotorespiration, as originally suggested by Zelitch (1966),but only at low concentrations and over a short time span. Thedecrease of PLIB and PLTB values at low -HPMS levels is consistentwith these processes being a residual activity of the glycolatepathway. Key words: CO₂ compensation point, -hydroxy-2-pyridinemethanesulphonic acid, photorespiration, photosynthesis     

srb: a Bacillus subtilis Gene Encoding a Homologue of the {alpha}-Subunit of the Mammalian Signal Recognition Particle Receptor

We cloned a Bacillus subtilis gene (srb) encoding a homologueof the mammalian signal recognition particle receptor -subunit(SR). The gene is 987 bp in length and encodes a 329-amino acidprotein. The deduced amino acid sequence of the protein shared26.6, 36.2 and 49.7% identity with those of mammalian SR, archaebacterialDP and Escherichia coli FtsY, respectively. The protein containsthree conserved GTP-binding elements like the other three SRPreceptor proteins, though the N-terminal portion of the putativeB. subtilis protein was shorter than the others. Secondary structureprediction showed that an amphipathic -helix is positioned inthe N-terminal region. A defect in srb inhibited cell growthand protein translocation.     

Relationship between Photosynthesis and Chlorophyll Content during Leaf Senescence of Rice Seedlings

Photosynthetic oxygen evolution, chlorophyll contents and chlorophylla /b ratios of 3rd to 6th leaves of rice seedlings were measuredto examine whether or not inactivation of photosynthesis duringsenescence is related to loss of chlorophyll. Photosyntheticactivity decreased more rapidly than chlorophyll content duringleaf senescence; as a result, the lower the leaf position, thelower was the rate of oxygen evolution determined on the basisof chlorophyll. Chlorophyll ab ratio also decreased with advancingsenescence. Electrophoretic analysis revealed that the declinein chlorophyll ab ratio is due to more rapid degradation ofthe reaction center complexes than light-harvesting chlorophyllab proteins of photosystem II and that the photosystem I reactioncenter disappears in parallel with the inactivation of photosynthesis.A simple method was developed to estimate the amounts of chlorophylla associated with the reaction center complexes of the two photosystemsfrom the total chlorophyll contents and chlorophyll ab ratiosof leaves. Rates of oxygen evolution, determined on the basisof chlorophyll a bound to the reaction center complexes, remainedconstant throughout the course of senescence. Thus, inactivationof photosynthesis is closely related with loss of the reactioncenter complexes during leaf senescence of rice seedlings. Theresults suggest that loss of photosynthesis is mainly causedby loss of a functional unit of photosynthesis or by a decreasein the number of whole chloroplasts. (Received April 22, 1987; Accepted August 13, 1987)     

Pigment Composition of a Novel Oxygenic Photosynthetic Prokaryote Containing Chlorophyll d as the Major Chlorophyll

The principal pigment found in the majority of oxygenic photosyntheticorganisms is known to be chlorophyll a. However, we isolateda new oxygenic photosynthetic prokaryote that contained chlorophylld as a predominant pigment with chlorophyll a being a minorpigment. Chlorophyll d had previously been noted but its naturaloccurrence and function remained unclear. Cells of the new prokaryotehad an absorption maximum at red region of 714–718 nmdue to chlorophyll d absorption, but no characteristic absorptionpeak of chlorophyll a around 680 nm was observed. Chlorophylld of the new organism was identified spectrophotometricallyin several solvents and its chemical structure was confirmedby NMR and FABMS analysis. The cell also contained a chlorophyllc-like pigment, zeaxanthin and a-carotene but not chlorophyllb and ß-carotene. The content of chlorophyll d accountedfor more than 2% of the cell dry weight, while the content ofchlorophyll a was less than 0.1%. The chlorophyll a/d ratioremained between 0.03 and 0.09 under different culture conditions.The light absorption characteristics and the high content ofchlorophyll d along with the small content of chlorophyll aindicated the existence of a new light utilization mechanisminvolving chlorophyll d. (Received October 7, 1996; Accepted December 16, 1996)     

Identification and characterization of an {alpha}-mannosidase from Trypanosoma cruzi

In this report we describe the first purification and characterizationof the acid -mannosidase from the human parasite Trypanosomacruzi. The purified enzyme exhibited a native mol. wt of 240000 Da and is apparently composed of four identical subunitsof mol. wt 58 000 Da. Each of the four subunits contains oneN-linked high-mannose-type oligosaccharide. The -mannosidaseexhibited a pH optimum of 3.5 and a pI of 5.9. This low pH optimumand the ability of swainsonine to inhibit its activity suggestthat the -mannosidase is a lysosomal enzyme. Antibodies againstthe T.cruzi enzyme did not react with mammalian lysosomal -mannosidaseand, conversely, antibody against a rat lysosomal -mannosidasedid not react with the T.cruzi enzyme. Thus, the T.cruzi enzymeappears to be distinct from its mammalian counterpart. -mannosidase lysosomal enzyme Trypanosoma cruzi     

Mycobacterial arabinan biosynthesis: the use of synthetic arabinoside acceptors in the development of an arabinosyl transfer assay

Information on the biosynthesis of the D-arabinans of the cellwall of Mycobacterium tuberculosis is rapidly emerging, withthe promise of new targets for drug development against tuberculosis.Accordingly, arabinosyl transferase assays were developed utilizingsynthesized [1–¹⁴C]-β-D-arabinofuranosyl-1-monophosphoryldecaprenolas donor and a variety of O- and S-alkyl arabinosides as acceptors.These were: -D-Araf-(15)--D-Araf-O- and -S-alkyl di-arabinosidesand -D-Araf-(15)--D-Araf-(15)--D-Araf-O- and -S-alkyl triarabinosides.Whereas the O- and S-alkyl monosaccharide acceptors were inactive,the O- and S-alkyl disaccharide and the O- and S-alkyl trisaccharideacceptors (<C12) possessed considerable acceptor activity,and the trisaccharide acceptors were more potent than the correspondingdisaccharides. The O-alkyl disaccharide acceptors with a C8alkyl chain were more active than those containing the C6 orC10 analogs. Chemical analysis of the enzymatically synthesizedproducts of the reactions demonstrated that β-D-arabinofuranosyl-1-monophosphoryldecaprenolwas an effective donor for two of the three potential arabinosyltransferases: β-D-arabinofuranosyl-1-monophosphoryldecaprenol:arabinan (15) arabinosyl transferase and β-D-arabinofuranosyl-1-monophosphoryl-decaprenol:arabinan β(12) arabinosyl transferase. The β(12) arabinosyltransferase activity was more in evidence in the presence ofthe O-alkyl disaccharide acceptor, whereas both transferaseswere about equivalent in the presence of the S-alkyl trisaccharideacceptor. The tuberculosis drug, ethambutol, a known mycobacterialarabinosyl transferase inhibitor, was inactive within thesearabinosyl transferase/acceptor based assay systems, supportingother evidence that a third activity, responsible for the formationof 13 linkage, is the drug target. acceptor arabinan biosynthesis glycosyltrans-ferase assay mycobacteria     

A PHENOLIC PIGMENT REACTIVE TO L-LACTATE CYTOCHROME C REDUCTASE OF YEAST

1. A phenolic pigment was extracted from baker's yeast. The pigmentis slowly autooxidizable, and rapidly oxidized with Rhus-laccaseor polyphenol oxidase and reduced by dithionite.
2. The pigmentdissolved in ethylether had an absorption peak at258 mµ,shoulders at 289 and 382 mµ and a plateauat 450–500mµ. The difference spectrum between oxidizedand reducedforms of the pigment showed a wide plateau around500 mµ.
3. The pigment supported the oxygen uptake by reconstructed enzymesystem: L-lactate, L-lactate cytochhrome c reductase and Rhuslaccaseor polyphenol oxidase. In its absence, no oxygen uptake tookplace. The pigment was replaced successfully with p-quinone,catechol and menadione, but not with ubiquinone. The sequenceof hydrogen transport can be represented: L-lactate L-lactatecytochrome c reductase "phenolic pigment" oxidase oxygen.

(Received August 11, 1967; )     

Temperature dependence of chlorophyll {alpha} fluorescence in lettuce and spinach chloroplasts at sub-zero temperatures

The temperature dependence of chlorophyll fluorescence wasmeasured in spinach and lettuce chloroplasts at sub-zero temperaturesin the presence of 50% ethylene glycol. In the presence of 5mM Mg²⁺, a fluorescence maximum appeared at –31?C in boththe spinach and lettuce chloroplasts, while in the presenceof only 5 mM Na⁺ as cations the maximum shifted to –20?Cin the spinach chloroplasts and to –11?C in the lettucechloroplasts. Since the occurrence of a maximum in the temperatureversus fluorescence curve is an indication for the transitionof the physical phase of thylakoid membrane lipids between theliquid crystalline and the phase-separation state (16, 18),these findings suggest that the (major) phase transition ofmembrane lipids occurs at these low temperatures in chloroplastsof higher plants and also that the phase transition temperatureis markedly lowered by the presence of divalent cations. Ethylene glycol at a concentration of 50% had almost no effecton the temperature dependence of chlorophyll fluorescence ina lamellar membrane preparation of Anabaena variabilis. In awater suspension of dimyristoylphosphatidylcholine, the additionof ethylene glycol to 50% did not alter the characteristic featureof the temperature dependence of fluorescence of 1-anilinonaphthalene-8-sulfonate.These findings suggest that 50% ethylene glycol does not affectthe temperature of the transition of the physical phase of membranelipids. ¹ C.I.W.-D.P.B. Publication No. 592. ² Present Address: Department of Biophysics and Biochemistry,Faculty of Science, University of Tokyo, Hongo 113, Tokyo, Japan. (Received June 22, 1977; )