Untersuchungen zur Kompartimentierung der freien Aminosäure Alanin in den Farnvorkeimen von Dryopteris filix-mas (L.) Schott im Rotlicht und im Blaulicht

Summary In fern gametophytes (= sporelings) there is a strong correlation between the degree of blue light mediated photomorphogenesis and the protein content of the organism (cf. Mohr, 1963). In a previous paper (Payer et al., 1969) we have shown that blue light specifically increases the rate of protein synthesis in the fern sporelings over the rate which is maintained under red light. — In the present paper blue light mediated protein synthesis has been dealt with further using one representative amino acid, alanine, which was labelled with ¹⁴C from ¹⁴CO₂ under steady state conditions of photosynthetic ¹⁴C incorporation under blue or red light.Synthesis of free alanine is proportional to the rate of photosynthesis (Table 1). For a number of reasons we conclude that alanine is derived directly from primary photosynthetic products. Since the pool size of the thoroughly ¹⁴C-labelled pool of free alanine is much less than the actual, pool size of this amino acid, (Table 1), and since the specific activity of the isolated ¹⁴C-alanine is much below the value we can expect on the basis of the specific activity of the ¹⁴CO₂ applied we conclude that there are separate pools of free alanine; active (with respect to protein synthesis) and inactive pools which do not mingle. Taking into account this possibility of compartmentation of pools of free amino acids we have calculated in the case of ¹⁴C-alanine the rate of protein synthesis for two extreme instances (Table 2). A comparison of the theoretical values with the actual data indicates that indeed protein synthesis is fed from active pools of amino acids while the inactive pools are possibly located in the vacuoles. The total pool of alanine is much larger in red grown than in blue grown sporelings while the active pools seem to have the same size under both conditions. The cells of the red grown sporelings have much larger vacuoles than the cells of the blue grown sporelings.The rate of protein synthesis is under our conditions 1.8 times higher in blue light than in red light. The rate of turnover of the total protein is 0.29% per hour in the blue and 0.23% in the red light. The absolute turnover of protein is 1.5 times higher in blue light than in red light. It is concluded that the blue light mediated increase of protein synthesis is very real. Blue light must act specifically at the level of polypeptide synthesis.     

Ein spezifischer Einfluß von Blaulicht auf den Einhau von photosynthetisch assimiliertem 14C in das Protein von Farnvorkeimen [Dryopteris filix-mas (L.) Schott]

Summary Morphogenesis and metabolism of the early gametophytes (=sporelings) of the common male fern are controlled by light. The normal two-dimensional development of the gametophytes takes place only in white or blue light; in red light alone, on the other hand, the sporelings remain filamentous even under conditions of equal photosynthetic rate.The problem has been whether blue light exerts its morphogenic influence through differential gene activation. In other words: does blue light mediate the synthesis of morphogenic enzymes which are required for normal morphogenesis. In an earlier paper (Drumm and Mohr, 1967) we have shown that blue light increases the rate of RNA synthesis within an hour whereas the first indication of a morphogenic change due to blue light is only discernible about 3 hours after the onset of blue light (Figs. 1,2). Furthermore we have shown (Mohr, 1965) that Actinomycin D specifically inhibits the blue light mediated morphogenic alterations, and Bergfeld (1967) has shown that blue light will rapidly lead to changes in nuclei and nucleoli in the fern sporelings. In the present paper it has been shown that blue light does increase the rate of protein synthesis about an hour after the transfer of the sporelings from the red into the blue light of equal quantum flux density (350 pE·cm^-2·s^-1).The rate of protein synthesis was measured in shortterm experiments (40min) using ¹⁴CO₂. The photosynthetic rate was the same in red and blue; it was not influenced by the transfer(Fig. 3). Likewise the rate of ¹⁴C incorporation into the pool of free amino acids was not significantly different in red and blue light (Fig.4). On the other hand, the rate of incorporation of ¹⁴C into the protein increased rapidly after the transfer of the sporelings from the red into the blue light (Fig. 5). The same phenomenon (no influence of blue light on the specific activity of the free amino acid; a strong promotive influence on the specific activity of the protein-bound amino acid) was observed in the case of alanine which was investigated in detail (Figs. 6, 7). Since the increase of the protein content of the sporelings is not significant during the first six hours after transfer to blue light (Fig. 8) the protein induced by blue light and directly related to morphogenesis can only be a very small fraction of the total protein of the sporeling.The data strongly support the hypothesis (Ohlenroth and Mohr, 1964), that the morphogenic effect of blue light on the fern sporelings is due to the induction of morphogenic enzymes by blue light.     

Die Wirkung von d-Aminosäuren auf die Struktur und Biosynthese des Peptidoglycans

Zusammenfassung In einem Konzentrationsbereich von 0,02–0,2 M hemmt d-Serin das Wachstum aller untersuchten Bakterien. Gleichzeitig traten morphologische Veränderungen der Bakterienzellen auf. In den nucleotidaktivierten Vorstufen von gehemmten Zellen wurden die d-Alaninreste des Peptidanteils ganz oder teilweise durch d-Serin ersetzt. Auch das Peptidoglycan enthielt d-Serin anstelle von d-Alanin, jedoch weiniger als in den Vorstufen. Zusätzlich war das modifizierte Peptidoglycan zu einem geringeren Anteil quervernetzt als das normale. Vier weitere d-Aminosäuren (Threonin, Valin, Leucin, Methionin) verursachten bei einer Konzentration von 0.2 M ähnliche Wirkungen wie d-Serin. Die Wirkungsweise von d-Aminosäuren auf die Peptidoglycansynthese kann daher allgemein wie folgt beschreiben werden: In Gegenwart von wachstumshemmenden Konzentrationen an d-Aminosäuren werden modifizierte nucleotidaktivierte Peptidoglycanvorstufen synthetisiert, die zu einem geringeren Ausmaß in das Peptidoglycan eingebaut und im Peptidoglycan schlechter quervernetzt werden. Der Ersatz von d-Alanin in Position 4 der Peptiduntereinheit ist dabei in der Regel am wirkungsvollsten. Nur bei Corynebacterium insidiosum und Staphylococcus aureus erwies sich der Ersatz in Position 5 als stärker hemmend. Diese Wirkungsweise entspricht weitgehend derjenigen von Glycin. Im Unterschied zur Wirkung von Glycin kann l-Alanin in Position 1 der Peptiduntereinheit nicht durch d-Aminosäuren ersetzt werden.

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Mode of action of d-amino acids on the biosynthesis of peptidoglycan

The mechanism of growth inhibition by d-amino acids was studied. d-Serine at concentrations from 0.02–0.2 M was sufficient to cause partial growth inhibition in seven species of bacteria representing the four most common types of peptidoglycan. The inhibited cells displayed morphological alterations. In the nucleotide-activated peptidoglycan precursors of these cells, d-alanine residues in position 4 and/or 5 of the peptide moiety were partially or even completely replaced by d-serine. The peptidoglycan also contained d-serine instead of d-alanine, but the percentual content of d-serine was significantly lower than that in the precursors. In addition, the modified peptidoglycan was less cross-linked than the normal one. Four other d-amino acids (d-threonine, d-valine, d-leucine, d-methionine) at concentrations of about 0.2 M caused similar effects as did d-serine when applied to Corynebacterium callunae and Bacillus subtilis. Thus the mode of action of d-amino acids on peptidoglycan synthesis can be generally described as follows: in their presence, at growth inhibiting concentrations modified nucleotide-activated peptidoglycan precursors are formed in which d-alanine residues are replaced by the d-amino acids. They are less efficiently incorporated into peptidoglycan. A high percentage of the modified muropeptides remains non-cross-linked, since they are poor substrates for the transpeptidation reaction. In the majority of the organisms, cross-linking was decreased when d-alanine in position 4 of the peptide subunit was replaced, in two organisms (Corynebacterium insidiosum and Staphylococcus aureus) replacement in position 5 was most effective, however. The low extent of crosslinkage is consistent with the morphological aberrations of inhibited cells. In previous studies with glycine, results were described that were in close analogy to those obtained with d-amino acids. However, glycine can replace not only d-alanine residues in position 4 and 5 but also l-alanine in position 1 of the peptide subunit.

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Verwendete Abkürzungen Dab Diaminobuttersäure - m-Dmp meso-Diaminopimelinsäure - GlcNAc oder G N-Acetylglucosamin - MurNAc oder M N-Acetylmuraminsäure     

Der Einfluß von Rot- und Blaulicht auf die Photosynthese vonAcetabularia mediterranea und auf die Verteilung des assimilierten Kohlenstoffs

Zusammenfassung Bei Zellen der marinen Grünalge Acetabularia mediterranea liegen nach 2stündiger Photosynthese im Weißlicht (8000 Lux) etwa 80% des fixierten¹⁴C in äthanollöslicher Form vor, etwa 12% entfallen auf Stärke, 2–3% auf Protein und 6% auf die Zellwand.Werden die Zellen mit Rotlicht (Dauerlicht, 3800 erg · cm^–2 · sec^–1) bestrahlt, so fällt die Einbaurate in allen 4 Fraktionen stark ab (Abb. 1). Dabei nimmt der¹⁴C-Anteil in der äthanollöslichen Fraktion innerhalb von 3 Wochen zu Lasten der Stärke und Zellwand von 80% auf ca. 90% zu. Im Gegensatz dazu wird im Blaulicht (Dauerlicht, 5600 erg · cm^–2 · sec^–1) mit der Bestrahlungsdauer der Einbau in Stärke, Zellwand und Protein gefördert (Abb. 1).Trotz sinkender Einbauraten von¹⁴C in Stärke nimmt im Rotlicht der Stärkegehalt pro Zelle zu, liegt dagegen im Blaulicht trotz höherer¹⁴C-Einbauraten deutlich unter demjenigen der Rotlichtzellen (Tabelle 1 und 2). Die Akkumulation von Stärke im Rotlicht dürfte demnach auf einer Hemmung des Stärkeabbaus beruhen.Der Gehalt an löslichen Kohlenhydraten (Fructose, Glucose, Saccharose, Fructosane) stagniert in Rotlichtzellen und steigt in Blaulichtzellen um ein Mehrfaches an (Tabelle 1).Bestrahlung mit Blaulicht nach Rotlichtvorbehandlung führt zu einem Ansteigen der Photosyntheseintensität. Nach 8stündiger Bestrahlung nimmt die Fixierungsrate zu und erreicht nach 48- bis 72stündiger Bestrahlung etwa das 5- bis 6fache des am Ende der Rotlichtbestrahlung gemessenen Wertes (Abb. 2).Diesem Anstieg der Fixierungsrate muß offenbar eine Synthese von Proteinen vorausgehen (Abb. 3). Auch der¹⁴C-Einbau in Stärke und die Zellwand steigt bereits vor der Gesamtfixierung an, und außerdem wird der Abbau der während der Rotlichtvorbehandlung akkumulierten Stärke eingeleitet (Tabelle 2).Der Hauptanteil des¹⁴C in der löslichen Fraktion entfällt auf die löslichen Kohlenhydrate. Bestrahlung mit Blaulicht nach Rotlichtvorbehandlung führt zunächst zu einer Abnahme des¹⁴C-Einbaus in die löslichen Kohlenhydrate, gefolgt von einem starken Anstieg bis zur 72. Stunde und einem erneuten Abfall (Abb. 4). Während der¹⁴C-Einbau in Fructose, Saccharose und Glucose diesem Kurvenverlauf folgt, steigt der Einbau in Inulin bis zur 72. Stunde kontinuierlich an (Abb. 5).Demgegenüber ist der auf die basische (Aminosäuren) und die saure Fraktion entfallende Anteil gering. Der¹⁴C-Einbau in beide nimmt im Blaulicht kontinuierlich zu (Abb. 4). Aminosäuren werden in den Zellen auch nach 3wöchiger Bestrahlung mit Rotlicht gebildet. Ferner ist der Gehalt an Aminosäuren am Ende der Rotlichtvorbehandlung am höchsten (Tabelle 3). Die Syntheserate von Protein in Rotlicht dürfte demnach nicht durch die Aminosäurekonzentration begrenzt werden.Die Ursache für den Abfall der Photosyntheseintensität bei Rotlichtbestrahlung ist den vorliegenden Daten nicht zu entnehmen. Die Möglichkeiten, die dabei eine Rolle spielen könnten, werden diskutiert.

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The effect of red and blue light on photosynthesis ofAcetabularia mediterranea and on the distribution of assimilated carbon

Summary After photosynthesis for two hours in white light (8000 lux), cells of the marine chlorophycean algaAcetabularia mediterranea contain about 80% of the¹⁴C incorporated in ethanol soluble form, about 12% in starch, 2–3% in protein, and 6% in the cell wall.When cells are irradiated with red light (continuous light, 3800 erg · cm^–2 · sec^–1), the incorporation rate for all four fractions is sharply reduced (Fig. 1). Concomitantly, the¹⁴C content in the ethanol soluble fraction rises in three weeks from 80% to about 90%, to the debit of starch and cell wall. In contrast to these findings, incorporation into starch, cell wall, and protein under blue light (continuous light, 5600 erg · cm^–2 · sec^–1) rises with the irradiation time (Fig. 1).Starch content per cell rises under red light in spite of declining incorporation rates of¹⁴C into starch, whereas it is clearly reduced in blue light below the values for red light cells, notwithstanding the increased¹⁴C incorporation rates (Tables 1 and 2). Accumulation of starch under red light seems to be due, therefore, to an inhibition of starch degradation.Soluble carbohydrate content (fructose, glucose, sucrose, fructosans) stagnates in red light cells and is multiplied in blue light cells (Table 1).Blue light irradiation after red light pretreatment increases the intensity of photosynthesis. The assimilation rate rises after an irradiation period of eight hours, reaching, after 48 to 72 hours of irradiation, about five to six times the level at the end of the red light period.Obviously, this rise in the assimilation rate must be preceded by protein synthesis (Fig. 3).¹⁴C incorporation into starch and cell wall rises even before the increase in total fixation, too, and, in addition, degradation of starch accumulated during the red light pretreatment is initiated (Table 2). The main amount of¹⁴C in the soluble fraction falls to soluble carbohydrates. Irradiation with blue light after red light pretreatment results at first in a reduction of¹⁴C incorporation into soluble carbohydrates, followed by a sharp increase till the 72nd hour and another decline (Fig. 4).¹⁴C incorporation into fructose, sucrose, and glucose follows this pattern, whereas incorporation into inulin increases continuously till the 72nd hour (Fig. 5).The amount falling to the basic and the acid fractions is small, in contrast.¹⁴C incorporation into both fractions rises continuously in blue light (Fig. 4).Amino acids are formed in the cells even after a three-week period of red light irradiation. Furthermore, the amino acid content is highest at the end of the red light pretreatment (Table 3). Thus, the rate of protein synthesis in red light seems not to be limited by amino acid concentration.The cause for the reduction of photosynthesis under irradition with red light does not become obvious from the data obtained. Factors possibly playing a role in this process are discussed.

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Die Untersuchungen wurden durch Sachmittel der Deutschen Forschungsgesellschaft unterstützt. Frau I. MAASS danke ich für die sorgfältige Mithilfe bei den Versuchen.     

Ein Vergleich der Wirkung von ionisierender Strahlung auf peptidgebundene und freie Aminosäuren in festem Zustand

Zusammenfassung Untersuchungen an thermischen Polymeren von-Aminosäuren in festem Zustand zeigen, daß in diesen insbesondere Tryptophan, Histidin, Cystin, Lysin und Methionin eine höhere Strahlenempfindlichkeit als in den bisher untersuchten Proteinen aufweisen. Diese Ergebnisse werden verglichen mit ähnlichen Untersuchungen an Filmen von Aminosäuremischungen, die in noch stärkerem Umfang auf einen beträchtlichen Energietransfer oder Chargetransfer in Richtung auf die vier genannten Aminosäuren schließen lassen. Die Ergebnisse werden auch in Hinsicht auf die Strahlenempfindlichkeit von Aminosäuren in Proteinen und auf die Inaktivierung von Enzymen diskutiert.

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A comparison of the effect of ionizing radiation on peptide- bound and free amino acids in the solid state

Summary Irradiation of thermal polymers of-amino acids with X-rays in the solid state produces a significantly increased destruction of tryptophan, histidine, cystine, lysine and methionine as compared with the response of constituent amino acids in proteins. These results are discussed with respect to related results obtained by irradiation of dry films of amino acid mixtures which indicate an even stronger energy or charge transfer towards the four amino acids mentioned. The results are also discussed with respect to the radiation sensitivity of constituent amino acids in proteins and the inactivation of enzymes.

     

Die Wirkung von Actinomycin D auf die phytochromabhängigen Veränderungen des RNS-Gehaltes im Senfkeimling (Sinapis alba L.)

Summary Actinomycin D (10 g/ml) cancels completely the phytochrome-mediated RNA net synthesis in the cotyledons of the mustard seedling whereas RNA net synthesis in the cotyledons of the dark-grown seedling is only partially inhibited (Fig. 1). — In the hypocotyl Actinomycin D of the same concentration lowers the RNA contents in the light (i.e. far-red)-grown seedling as well as in the dark-grown seedling down to the same level (Fig. 2). In the presence of Actinomycin D phytochrome has no significant influence on the RNA contents neither in the cotyledons nor in the hypocotyl (Fig. 1,2).The data support the view that P₇₃₀, the active phytochrome, acts through differential gene activation in the cotyledons and predominantly through differential gene repression in the hypocotyl (cf. Mohr, 1966; Schopfer, 1967a, b). —The data further support the conception that active genes (as defined by Mohr, 1966 and Schopfer, 1967a, b) are much less sensitive towards Actinomycin D than potentially active and repressible genes (cf. Schopfer, 1967a; Mohr and Bienger, 1967).     

Transport und Stoffwechsel von 2-[14C]Abscisinsäure in Wurzelsegmenten von Phaseolus coccineus L.

Summary Movement of 2-[¹⁴C]ABA through 1.5 cm and 5 cm long root segments of P. coccineus L. was acropetally polarised. The velocity of acropetal transport of [¹⁴C]ABA in 1.5 cm long segments was 4–5 mm·h^-1. Up to 11 h after the start of incubation [¹⁴C]ABA could be extracted unchanged. Beyond this time radioactivity became associated to an unidentified compound, which shows chromatographic qualities similar to those of Milborrow's Metabolite C (Biochemistry & Physiol. Plant Growth Substances, 1531, Runge Press Ottawa, 1968; and Chem. Comm. 966, 1969).

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Abkürzungen ABA Abscisinsäure - PPO 2,5-Diphenyloxazol     

Die Zeitwirkung von Temperaturänderungen auf die CO2-Abgabe von Kartoffelparenchym

Ohne ZusammenfassungMit 1 Textabbildung.     

Die Wirkung von Kinetin, 2,4-DNP und Antimetaboliten auf die Veränderungen im Aminosäurengehalt welkender Pflanzenblätter

Summary It has been established that in intact plants suffering from scarcity of water as well as in isolated withering leaves the amount of proline increases considerably. The main factor in the isolated leaves is thus the scarcity of water and not the injury due to the defoliation. The proline content of the leaves withering in the dark increases only for a few days and then decreases with the diminution of the carbohydrates. Of the active substances tested only 2,4-DNP inhibited the proline synthesis during the withering. It is very probable that in the course of withering the great amount of proline forms during the oxidation of carbohydrates via -ketoglutarate. The oxydative phosphorylation is uncoupled by 2,4-DNP. Kinetin, 2,4-D and antimetabolites applied do not inhibit the abnormal increase of proline.     

Über die Wirkung kurzfristiger Kinetingaben auf Protein,Aminosäuren und RNS bei Lemna minor L.

Summary Lemna minor L. was cultivated on nutrient medium with kinetin (10^-5 m) for 3 hours at 28°C and ca. 5,000 lux. Samples were taken at different times and their amino acid, protein and RNA content was determined.The amino acid content in the cultures with kinetin corresponds to that in the cultures without kinetin. The amino acid concentration seems to be regulated by mechanisms independent of kinetin. On the other hand, kinetin increases the quantity of protein immediately i.e., within the first two minutes. This increase stops after ca. 30 minutes, but the excess of protein remains for a long time. In the first 30 minutes the RNA concentration also rises. This increase is not as strong as that of the protein nor does it remain. After 30 minutes, when the rise of protein content stops, the RNA concentration begins to drops within 1 hour to that of the control cultures without kinetin.The extent of the protein and RNA increase induced by kinetin depends strongly upon the physiological state of the plants. This may be because the plants produce different quantities of endogenous cytokinins under different conditions.The protein induced by kinetin was then examined after centrifugation at 100,000 x g. In samples taken during the first 5 minutes, the quantity of protein in the supernatant and in the sediment is increased. In samples taken after this period, the increase of protein in the supernatant disappears and only that in the sediment remains. After some time, therefore, all the excess protein consists of structural protein, which is found in the sediment. The findings are compared with those of other authors and an unspecific stimulation of RNA is suggested.     

Zur Ätiologie der Anreicherung von Aminosäuren und Amiden im Wurzelraum vonHelianthus Annuus L

Ohne ZusammenfassungMit 5 Textabbildungen     

Die Häufigkeit und die Verteilung der Perlschnurleisten auf den Händen von geistig normalen Menschen und Mongoloiden

Ohne ZusammenfassungDirektor: Prof. Dr. Dr. Vogel Mit 2 Textabbildungen     

Die Wirkung von niedriger Temperatur,Stickstoffatmosphäre und Entkopplern auf die longitudinale Radioaktivitäts-verteilung nach akropetaler und basipetaler Applikation von [2-14C]ABA in langen Wurzelsegmenten von Phaseolus coccineus L.

Summary Acropetal movement of [2-¹⁴C] abscisic acid (ABA) in long root segments of P. coccineus is drastically reduced by destruction of the tissue, low temperature, anaerobic conditions and pretreatment with CCCP (carbonyl-cyanide-m-chloro-phenylhydrazone) at uncoupling concentrations. Basipetal transport is not affected by destruction of the tissue but markedly inhibited by the other treatments. Consequently polarity of ABA-transport disappears only after the roots have been killed. The importance of using long segments for hormone transport studies is discussed.

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Abkürzungen ABA Abscisinsäure - CCCP Carbonyl-cyanid-m-chlor-phenylhydrazon - DNP Dinitrophenol - IAA Indolessigsäure - PPO Diphenyloxazol Vorübergchende Anschrift bis zum 31. 3. 1976: Botanisches Institut der Hochschule für Bodenkultur, A-1180 Wien 18, Gregor-Mendel-Straße 33, Austria.     

Die Wirkung der äusseren Bedingungen auf die Veränderung des Geschlechts und auf die Entwicklung von Daphnia pulex de Geer

Ohne Zusammenfassung     

Der Einfluss von Phenäthylpyridinen und Pyridylphenylazetylenen auf die Wurzelatmung vonSinapis alba L.

Aktivita sukcinát-dehydrogenázy SDH a sukcinát-oxydázového systému SO byla mě?ena sestý den po vyklí?ení v homogenátu z ko?en? klí?ních rostlin ho??ice, které byly pěstovány v ?ivném roztoku, obsahujícím některý ze t?í polohových isomer? fenethylpyridinu (IIIA, B, C) nebo pyridylfenylacetylenu (IVA, B, C) v koncentraci 5×10^?4 m. Slou?eniny fenethylpyridinového typu prakticky neovlivňují aktivitu dehydrogenázy kyseliny jantarové SDH aniin vivo, aniin vitro, av?ak pyridylfenylacetyleny aktivitu tohoto enzymuin vivo výrazně stimulují. Naproti tomu oba typy slou?enin v pokusném uspo?ádání jakin vivo, takin vitro pr?kazně inhibují aktivitu sukcinát-oxydázového systému SO. Ú?inek fenethylpyridin? i pyridylfenylacetylen? je tedy pravým opakem ú?innosti trans-styrylpyridin? (typ I).