RNA and protein metabolism in the particulate fractions of the gametophytes of bracken fern during growth in red and blue light

Summary The metabolism of RNA and protein in the gametophytes of bracken fern (Pteridium aquilinum) is affected by the quality of light in which they are grown. When sporelings were grown as two-dimensional gametophytes in blue light, particulate fractions separated from the sporelings exhibited greater incorporation of uridine-³H and leucine-³H into RNA and protein, respectively, than those from sporelings grown as one-dimensional protonema in red light. After various periods of exposure of gametophytes to red or blue light in the presence of uridine-³H, the nuclei-rich fraction showed the highest specific activity in RNA, and irrespective of incubation time, blue light was more effective than red light. The possibility that enhanced synthesis of RNA in the nucleus in response to blue light is significantly related to the morphological growth pattern of the gametophytes, is discussed.     

THE TRANSITION FROM FILAMENTOUS TO TWO-DIMENSIONAL GROWTH IN FERN GAMETOPHYTES. III. INTERACTION OF CELL ELONGATION AND CELL DIVISION

Periodic cell divisions were induced in gametophytes of Pteridium aquilinum by daily irradiation with white light. In white-dark cycles, the rate of cell division was promoted by increased time in white light; cell elongation was not affected. The time of transition to two-dimensional growth (days to 5% 2-D) was closely associated with the mitotic rate. For white-red cycles, the rate of elongation was controlled by the intensity of red light (wavelengths over 550 nm). This increased elongation delayed the initiation of 2-D development. In both cases the rate of transition to 2-D growth was correlated with the amount of elongation per division.     

Chloroplastenausbildung und Morphogenese der Gametophyten von Dryopteris filix-mas (L.) Schott nach Applikation von Chloramphenicol und Actidion (Cycloheximid)

Summary Morphogenesis and differentiation of the young gametophytes (=sporelings) of Dryopteris filix-mas are controlled by light. Blue light leads to the formation of normal two-dimensional prothallia; under red light, however, the gametophytes grow as cellular filaments. Morphogenesis in blue light is connected with an increase in protein synthesis; in red light the protein content of the sporelings is markedly lower. The size of the chloroplasts is correlated with the protein content of the sporelings.In the present paper the diverse effect of chloramphenicol (CAP) and actidione (cycloheximide, ACT) was studied in connection with the formation of chloroplasts. ACT blocks the growth of the gametophytes, while the chloroplasts remain functional. On the other hand, CAP does not influence morphogenesis of the gametophytes. In particular the activity of the dividing apical cells remains untouched. Even when the light quality is changed during the development the corresponding specific effect of the light quality on morphogenesis is normal. The chloroplasts, however, become smaller, probably by inhibition of synthesis of structural proteins. But their synthetic activity is not completely suppressed. The specific blue or red light dependent morphogenesis is not changed, when protein synthesis in the chloroplasts is inhibited.     

THE RELATIONSHIP BETWEEN THE PROMOTION OF ELONGATION OF FERN PROTONEMATA BY LIGHT AND GROWTH SUBSTANCES

Germinating spores of the fern Onoclea sensibilis L. were grown in darkness, so that they developed as filaments (protonemata). Brief daily exposure of the filaments to red, far-red or blue light increased the rate of filament elongation. Filament elongation was also promoted by indoleacetic acid. When filament elongation was promoted with both indoleacetic acid and exposure to light, the growth promotions caused by red and far-red light were additive to auxin-induced growth. Blue light promoted elongation only at sub-optimal concentrations of auxin. Elongation induced by guanine was additive to red- and far-red-induced elongation. Gibberellic acid had no effect on elongation under any condition. Blue-light-induced elongation resembled auxin-induced elongation in its requirement for exogenous sucrose and sensitivity to inhibition by parachlorophenoxyisobutyric acid. Red and far-red light were active regardless of the presence or absence of sucrose and promoted elongation at a concentration of parachlorophenoxyisobutyric acid which completely inhibited blue-light-induced elongation.     

ACTINOMYCIN D-INDUCED CHANGES IN GROWTH AND RIBONUCLEIC ACID METABOLISM IN THE GAMETOPHYTES OF BRACKEN FERN

Actinomycin D affected the morphological type of growth in the gametophytes of Pteridium aquilinum and the distribution of RNA and protein in their particulate fractions. Increasing concentrations of the drug progressively inhibited two-dimensional growth at the end of a period during which controls had formed typical two-dimensional plants. RNA was lost maximally from the nuclei-rich and ribosome-rich fractions of plants growing in a concentration of actinomycin D which inhibited two-dimensional growth. The magnitude of changes in protein content of the plants was less striking. Presence of actinomycin D in the medium also suppressed incorporation of uridine-H³ into cytoplasmic fractions of gametophytes. The possibility that two-dimensional growth in the gametophytes is under control of a newly synthesized messenger RNA, which is sensitive to actinomycin D, is discussed.     

Inhibition of two-dimensional growth and suppression of ribonucleic acid and protein synthesis in the gametophytes of the fern, Asplenium nidus, by chloramphenicol, puromycin and actinomycin D

Chloramphenicol and puromycin at appropriate concentrations inhibited the induction of two-dimensional growth in the gametophytes of the fern Asplenium nidus without drastically inhibiting germination and continued filamentous growth. Similar responses to actinomycin D were reported earlier. Radioautographic techniques were employed to study the pattern of ribonucleic acid and protein synthesis in gametophytes which were treated with chloramphenicol, puromycin and actinomycin D. Uptake of H³-uridine into ribonucleic acid was strongly inhibited by all three antibiotics. Chloramphenicol and puromycin were not as effective as actinomycin D in inhibiting H³-leucine incorporation. The results are discussed in relation to the quality of light and antibiotics on two-dimensional growth in the gametophytes.     

ACTION OF PURINE AND PYRIMIDINE ANALOGS ON THE GROWTH AND DIFFERENTIATION OF THE GAMETOPHYTES OF THE FERN ASPLENIUM NIDUS

The purine analogs, 8-azaadenine, 8-azaguanine, 8-azaxanthine and 8-azahypoxanthine, and the pyrimidine analogs, 2-thiocytosine, 5-fluorouracil, 2-thiouracil and 6-azauracil, inhibited the induction of 2-dimensional growth in the gametophytes of the fern Asplenium nidus L. In contrast, thymine analogs such as 5-fluorodeoxyuridine, 2-thiothymine, 6-azathymine and 5-bromouracil caused non-specific growth inhibitions without suppressing 2-dimensional growth. Subinhibitory concentrations of 8-azaxanthine, 8-azahypoxanthine, and 2-thiouracil promoted both 1-dimensional and 2-dimensional phases of growth of the gametophytes. Inhibitory effects of the analogs were observed on treatment of the spores or of gametophytes of different ages. Gametophytes growing in the analogs for different periods of time recovered from inhibition on transfer to the basal medium.     

THE EFFECT OF LIGHT QUALITY ON THE CARBON METABOLISM AND EXTRACELLULAR RELEASE OF CHLAMYDOMONAS REINHARDTII DANGEARD1

The influence of red, blue, green, and white light on growth and photosynthetic rates, carbon metabolism, and rates of release of extracellular compounds in the freshwater alga Chlamydomonas reinhardtii Dangeard was examined. Relative growth constants were 0.28, 0.32, 0.40, and 0.41 in green, white, blue, and red light, respectively. Photosynthetic rates were higher in white, blue, or red than in green light of the same intensity. More than 66% of the ¹⁴CO₂ assimilated by cells grown under blue or green light was incorporated into the ethanol-insoluble fraction, compared with about 50% in cells grown under white or red light. The percentage of sugars in this fraction was significantly higher in cells grown under green or red light than in cells cultured in white or blue light, while the percentage of proteins was highest in blue light. Light quality also influenced the composition of the ethanol-soluble fraction. The percentage of organic acids was highest in cells grown in green and white light, while amino acids were highest in blue and green cultures. The percentage of ethanol-soluble sugars was greatest in cultures grown in blue and red light. The percentage release of dissolved organic carbon into the medium was highest in white light and lowest in blue or red light. The nature of the extracellular products varied according to the quality of light under which the cells were cultured, but had no consistent relation to the nature or concentration or components in the ethanol-soluble fraction.     

PHOTOCONTROL OF THE ORIENTATION OF CELL DIVISION IN ADIANTUM. III. EFFECTS OF METABOLIC INHIBITORS

5-Fluorouracil, 8-azaguanine and ethionine were tested on the orientation of cell division to see whether the two-dimensional development of the fern Adiantum gametophytes was due to newly synthesized protein(s). Using the system in which the orientation of cell division was controlled experimentally by sequential treatment with red light, white light and darkness and by the direction of irradiation, all the inhibitors decreased the rates of cell elongation and cell division of the gametophytes, but did not specifically affect the two-dimensional differentiation at all.     

THE TRANSITION FROM FILAMENTOUS TO TWO-DIMENSIONAL GROWTH IN FERN GAMETOPHYTES. II. KINETIC STUDIES ON PTERIDIUM AQUILINUM

Gametophytes of the fern Pteridium aquilinum were incubated 3 days in red light and then transferred to white light. The sequence of events occurring after the transfer was as follows: a swelling of the apical region within 1 hr; a reduction in cell elongation after 5 hr; a series of one-dimensional cell divisions between 10-25 hr; and the initiation of two-dimensional gametophytes after 25-50 hr. The percentage of two-dimensional gametophytes was proportional to the logarithm of the intensity of white light. The rate of elongation after 5 hr was inversely proportional to the logarithm of the intensity. The rates of cell divisions for one- and two-dimensional gametophytes were proportional to intensity up to 240 and 120 ft-c, respectively.     

GROWTH INHIBITION IN GAMETOPHYTES AND OAT COLEOPTILES BY THELYPTERIN A AND B RELEASED FROM ROOTS OF THE FERN THELYPTERIS NORMALIS

Gametophytes, when grown in the immediate vicinity of a Thelypteris normalis sporophyte—in soil or in sterile culture on agar—showed a reduced number of cells and an altered gross morphology. This is attributed to the action of the thelypterins, which are inhibitors released from T. normalis sporophytes. Growth inhibition of the gametophytes was greatest when thelypterins were added during early stages of gametophyte development. Removal of thelypterin A permitted resumption of growth. Thelypterin A noncompetitively inhibited auxin-enhanced elongation of A vena coleoptiles. The growth of T. normalis root segments was not affected by thelypterins. The growth of young sporophytes of T. normalis was inhibited by mature sporophytes.     

HYBRIDIZATION,POLYPLOIDY AND ADVENTITIOUS GROWTH IN THE GENUS ASPLENIUM

Gametophytes of Asplenium platyneuron, A. rhizophyllum and A. monlanum were grown from spores under controlled conditions. Factors inhibiting and promoting germination and growth were determined. Leaves cut from sexually produced sporophytes of A. platyneuron, A. rhizophyllum and A. montanum were cultured under controlled conditions on agar slants. The leaves produced neoplastic growths of two kinds: (1) two-dimensional growths, (2) three-dimensional growths. The former developed into normal-appearing, rhizoid- and gametangia-producing, heart-shaped gametophytes having a diploid chromosome complement. The formation of two-dimensional growths was favored by conditions drastically reducing the energy supply of the initiating sporophytic cells and by destroying the correlation of the leaf. Three-dimensional growths most often developed into sporophytes unless conditions were limiting; in the latter case they transformed into two-dimensional growths. Both two- and three-dimensional growths developed from fragmented leaves excised from buds growing near the tips of acuminate fronds of A. ebenoides. The high frequency of two-dimensional adventitious growth leads to the conclusion that apospory may provide a mechanism for the production of fertile allotetraploids, by the fusion of diploid gametes of adventitiously produced diploid gametophytes, in the genus Asplenium and perhaps other fern genera, in contrast to that which has been previously suggested in the literature.     

QUANTIFICATION OF THE EFFECTS OF SPORELING COALESCENCE ON THE EARLY DEVELOPMENT OF GRACILARIA CHILENSIS (RHODOPHYTA)1

Sporeling coalescence in species of Gracilariales and Gigartinales is predicted to result in larger basal areas of growing disks as well as earlier initiation, increased abundance, and faster growth rates of erect shoots as compared to noncoalescent sporelings. These responses have been interpreted as providing mutual benefits for organisms living in aggregation, counterbalancing disadvantages associated with crowding. Quantitative evaluations of sporelings of Gracilaria chilensis failed to support several of these predictions. Sporelings were grown in the laboratory from a range of single sporelings to coalescent masses of 20 sporelings. Coalescent sporeling masses of G. chilensis exhibited larger basal areas than noncoalescent ones, but because the specific growth rates were inversely related to the original number of carpospores, no significant differences in actual area increments, during most of the experiment, were found among sporelings derived from one, two, or three to five coalescing sporelings. Initiation of erect shoots occurred at a similar time, regardless of their origin, i.e. coalescent or noncoalescent. Abundance of erect shoots was only loosely related to the number of coalescing sporelings. Even though by the end of the experiment (week 18), the total length of the longer erect shoots arising from coalescent sporeling masses was significantly greater than that of shoots arising from noncoalescent sporelings, total length was independent of the original number of coalescing sporelings. Furthermore, specific elongation rates between week 12 and week 18 were significantly greater for noncoalescent sporelings than for coalescent sporeling masses. Quantitative screening of other species seems necessary before generalizations on the ecological advantages of sporeling coalescence in seaweeds can be made.     

THE INFLUENCE OF CULTURAL CONDITIONS ON THE INDUCTION OF APOGAMY IN PTERIDIUM GAMETOPHYTES

Apogamous sporophytes formed on Pteridium gametophytes in response to concentrations of certain sugars which supported gametophytic growth. High osmotic concentration of the medium inhibited apogamy, while variations in the basic medium were not stimulatory. Agar, autoclaving, the ammonium ion, and dry media were not required for apogamy. Renewing the medium during an experiment enhanced the apogamous response. Changing the medium at set intervals facilitated the separation of apogamous plant development into gametophytic, initiative, and developmental phases, thus enabling testing of various factors at each of these stages. Apogamy was light-initiated, while the actual development of apogamous sporophytes was caused by light, succinic acid or sugar.     

Eine Analyse der durch Blaulicht bewirkten Steigerung der Proteinsynthese bei Farnvorkeimen auf der Ebene der Aminosäuren

Zusammenfassung Bei der quantitativen Analyse der Protein-Aminosäuren von Farnvorkeimen (Dryopteris filix-mas) ergab sich, daß die Steigerung des Proteingehalts durch Blaulicht mit einer Zunahme aller untersuchten Aminosäuren einhergeht. Dabei bleibt der prozentuale Anteil der einzelnen Aminosäuren am Gesamtprotein immer annähernd gleich, abgesehen vom Prolin. — Offenbar kommt es unter dem Einfluß von Blaulicht hauptsächlich zu einer quantitativen Vermehrung und nur im geringen Maße zu einer qualitativen Veränderung der Proteinfraktion. — Da die pools der erfaßbaren freien Aminosäuren im Hellrot stets größer sind als im Blaulicht, ist der Schluß berechtigt, daß das Blaulicht seine Wirkung im Zusammenhang mit der Polypeptidsynthese und nicht über eine Steigerung der Aminosäuresynthese ausübt.

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An analysis of the blue light mediated increase of protein synthesis in fern gametophytes on the level of amino acids

Summary Morphogenesis and metabolism of the sporelings (= young gametophytes) of the common male fern Dryopteris filix-max are controlled by visible radiation. Short wavelengths visible radiation (= blue light) leads to an increase in protein synthesis and makes possible the formation of normal two-dimensional prothallia. Under long wavelengths visible radiation (= red light) the sporelings grow as cellular filaments, the protein contents of which are markedly lower than under blue light even under conditions of equal rate of dry matter accumulation in red and blue light (Fig. 2). — Quantitative amino acid analysis of the total protein of blue and red grown sporelings did not reveal any striking difference. The contents of all those amino acids which can be measured quantitatively after protein hydrolysis show about the same increase under the influence of blue light (Figs. 3, 4). Only in the case of proline are the differences between red grown and blue grown sporelings indicative of a qualitative change in the nature of the protein fraction (Fig. 3/h). —The pools of the free amino acids are always smaller in blue grown sporelings than in red grown ones (Figs. 6, 7). — The facts reported in this paper indicate that blue light leads mainly to a quantitative increase in the rate of protein synthesis. The influence of blue light on the qualitative nature of the protein fraction seems to be slight. On the other hand, the data on the pool sizes of the free amino acids support the conclusion that blue light controls the rate of protein synthesis at the stage of polypeptide synthesis and not through amino acid synthesis.

     

SPORELING COALESCENCE IN CHONDRUS CRISPUS (RHODOPHYCEAE)1

Coalescence of developing sporelings of Chondrus crispus Stackhouse was observed. Juvenile tetra-sporophytes showed a higher proportion of coalescence than developing gametophytes. Stages of complete coalescence between different sporelings are illustrated. Coalesced sporelings exhibit vertical and horizontal alignment of cells, as well as “cuticular” continuity and secondary pit connections between adjacent, coalesced sporelings. Ultimately the cells in the center of the coalesced sporelings produce upright, multiaxial fronds that grow more rapidly than fronds of non-coalesced sporelings. Other red algae, such as Gracilaria verrucosa (Hudson) Papenfuss and Gigartina stellata (Stackhouse) Batters also show a similar sequence of sporeling coalescence and enhanced growth. The ecological significance of sporeling coalescence is discussed.     

RESPONSE OF PROROCENTRUM MARIAE-LEBOURIAE (DINOPHYCEAE) TO LIGHT OF DIFFERENT SPECTRAL QUALITIES AND IRRADIANCES: GROWTH AND PIGMENTATION1

Growth and pigment concentrations of the, estuarine dinoflagellate, Prorocentrum mariae-lebouriae (Parke and Ballantine) comb. nov., were measured in cultures grown in white, blue, green and red radiation at three different irradiances. White irradiances (400–800 nm) were 13.4, 4.0 and 1.8 W · m^?2 with photon flux densities of 58.7 ± 3.5, 17.4 ± 0.6 and 7.8 ± 0.3 μM quanta · m^?2· s^?1, respectively. All other spectral qualities had the same photon flux densities. Concentrations of chlorophyll a and chlorophyll c were inversely related to irradiance. A decrease of 7- to 8-fold in photon flux density resulted in a 2-fold increase in chlorophyll a and c and a 1.6- to 2.4-fold increase in both peridinin and total carotenoid concentrations. Cells grown in green light contained 22 to 32% more peridinin per cell and exhibited 10 to 16% higher peridinin to chlorophyll a ratios than cells grown in white light. Growth decreased as a function of irradiance in white, green and red light grown cells but was the same at all blue light irradiances. Maximum growth rates occurred at 8 μM quanta · m^?2· s^?1 in blue light, while in red and white light maximum growth rates occurred at considerably higher photon flux densities (24 to 32 μM quanta · m^?2· s^?1). The fastest growth rates occurred in blue and red radiation. White radiation producing maximum growth was only as effective as red and blue light when the photon flux density in either the red or blue portion of the white light spectrum was equivalent to that of a red or of blue light treatment which produced maximum growth rates. These differences in growth and pigmentation indicate that P. mariae-lebouriae responds to the spectral quality under which it is grown.     

Photocontrol of the orientation of cell division inAdiantum

The rate of transition from one- to two-dimensional growth of fernAdiantum gametophytes under white light depends on the age of gametophyte cultured under red light. When gametophytes were cultured for longer period under red light, the rate of transition decreased and the number of abnormal gametophytes increased. Although the first step of the transition was the first longitudinal cell division following the two transverse ones (Wada and Furuya, 1970), the time-lapse-video study revealed that the apical cell of protonemata became flattened in the plane perpendicular to the incident ray of white light before the first longitudinal cell division. Analytical study of growing part of the apical cell with grains of activated charcoal as markers revealed that the apical cell flattening occurred evenly throughout the equatorial circumference of the cell even in the shaded side of the protonemata as well as in the side irradiated with white light.     

Growth Promotion of Vegetative Gametophytes of <Emphasis Type="Italic">Undaria pinnatifida</Emphasis> by Blue Light

Through an acclimation period of 10 days, compared to white light, the maximal net photosynthetic rates were significantly higher for gametophytes of Undaria pinnatifida cultivated under blue light (400–500 nm), and were lower under red light (600–700 nm). Chlorophyll c and the carotenoid content of gametophytes were similar under blue light and red light but were much lower under white light. The growth rate of female gametophytes under blue light was higher than that under other lights, and the growth rate of male gametophytes showed little variation with respect to blue and white light. Male and female gametophytes were mixed together to form sporophytes under white, blue and red light. After approximately 5 days, 50% gametophytes became fertile under blue and white light, but remained vegetative under red light after 10 days.