The inhibition of potato sprout growth by light.

When potato seed tubers (Solanum tuberosum cv. Pentland Javelin) were stored in darkness or diffuse daylight at 9°C and transferred at intervals to conditions suitable for sprouting, their capacity for sprout growth was unaffected by the presence or absence of light during previous storage. When similar tubers were stored at 10°C, 18°C or 25°C, sprout growth commenced earliest at 25°C, but the date was unaffected by fluorescent light. It was concluded that light did not affect the length of the dormant period, but only the rate of sprout elongation after that period had ceased. When tubers with growing sprouts at 10°C or 18°C were transferred from darkness into fluorescent light, sprout growth virtually ceased. Transfer from light into darkness resulted in immediate sprout growth, at a rate comparable with tubers stored continuously in the dark. Tubers of three Peruvian cultivars, stored in farm-scale diffuse-daylight stores, grew progressively shorter sprouts with increasing daily exposure to light from 30 min to 12 h. Storage of cv. Wilja under 21 Wm^-2 (total) of white fluorescent light for 10 h per day maintained the sprouts at the same length as ten times this light intensity for 1 h per day. In a subsequent experiment with cv. Bintje the 10 h, low-intensity light regime gave slightly shorter sprouts. It appeared that the total light energy falling on the tubers was the dominant factor controlling sprout growth.     

Changes in ribulose-1,5-bisphosphate carboxylase and its translatable small subunit mRNA levels during senescence of mustard (Sinapis alba) cotyledons

During the senescence of detached first leaves of oat ( Avena sativa L. cv. Victory) seedlings (grown in continuous light) the protein is hydrolyzed and the proteases increase, but the expected simple relation between these two factors is not always realized. The present experiments examine the timing, the influence of light and darkness and the action of the protein synthesis inhibitors cycloheximide (CHI) and cordycepin. Transfer from dark to light delays the breakdown of both chlorophyll (Chl) and protein, but some residual proteolysis is ascribed to the enzyme initially present. Transfer to CHI resembles transfer to light, while the action of cordyceptin is similar but much weaker. Repeated determinations of the acid protease, which is the most active one and the first to appear, show that this enzyme is formed in the light about as rapidly as in the dark, though with different kinetics. In spite of this there is little proteolysis in light in the first 5 days. One possible explanation of that could be that protein is rapidly resynthesized in light, but treatment with [¹⁴C]-leucine shows that such resynthesis is no faster in light than in darkness. It is therefore concluded that the protease initially does not have access to its substrates and, as a corollary, that the senescence process must be controlled by the gradual impairment of the vacuolar membrane, allowing protease to enter the cytosol and attack the proteins there and in the organelles. This concept is supported by many observations on the timing and on the known changes in membrane permeability during senescence.     

The Formation and Development of Adventitious Buds on Potato Sprouts

Potato tubers were pre-sprouted in trays at 10 C for 28 d.The scale leaves at the base of the sprouts were marked andtubers then planted in field soil. Twenty-eight days after plantingan average of three adventitious buds were present per sproutcompared with an average of six below-ground axillary buds.Ten per cent of adventitious buds formed stolons compared withalmost 100 per cent of axillary buds. Similar responses wereseen with tubers pre-sprouted either in the light or dark.     

Circadian Timekeeping for the Photoperiodic Control of Budset in Picea abies (Norway Spruce) Seedlings

Romanian populations of Norway spruce are induced to set terminal buds by four inductive cycles of 8 h light/16 h darkness. To distinguish between circadian and hourglass timekeeping for the photoperiodic control of budset, seedlings were raised in continuous light at 300 µmol m^-2s^-1 at 20°C for 10 weeks. They were then exposed to an extended night regime consisting of three cycles of 8 h light/40 h dark with 4-h or 1-h nightbreaks (120 µmol m^-2s^-1) applied to groups of plants at intervals during the extended night. Following a final cycle of 8 h light/16 h dark to maximize budset, the plants were transferred to continuous light. Budset was delayed when the night-break was applied close to the critical nightlength (CNL) of 6-7 h or about 22-23 h later in the extended night, consistent with circadian rather than hourglass timekeeping. Confidence intervals were calculated for the times to maximum effect of the night-breaks.     

Effect of different photoperiods on the ultrastructure of the specific secretory cells and α-subunit mRNA level in the chicken pars tuberalis

The effect of different photoperiods on the specific secretory cells of the pars tuberalis was examined in male chicks. Animals were placed in one of three different photoperiod regimens: (1) normal control (light:dark = 12 h:12 h), (2) continuous light (L:D = 24 h:0), and (3) extended darkness (L:D = 1 h:23 h). The levels of common alpha-subunit mRNA in the pars tuberalis were examined by Northern blot analysis and compared with those in the pars distalis. In chicks exposed to continuous light for 1 week, alpha-subunit mRNA level in the pars tuberalis was decreased, although the level in the pars distalis was increased. Exposure to continuous light for 30 days also induced a decrease in alpha-subunit mRNA level in the pars tuberalis. On the other hand, in chicks exposed to extended darkness for 1 week, the alpha-subunit mRNA level of the pars tuberalis was markedly increased. In situ hybridization with digoxigenin-labeled common alpha-subunit cRNA probe also showed that the hybridization signals for alpha-subunit mRNA in the pars tuberalis cells become weak under continuous light for 30 days but they are very intense under extended darkness. Thus, the synthesis of alpha-subunits in the chick pars tuberalis was inhibited by continuous light but stimulated by extended darkness. These results were confirmed by semiquantitative electron-microscopic analyses. After exposure to continuous light for 30 days, many pars tuberalis (PT)-specific cells were filled with enlarged secretory granules, showing the reduction of secretory activity. On the contrary, extended darkness for 30 days induced hypertrophy of the PT-specific cells; the areas of cytoplasm and nucleus were significantly increased. In addition, secretory granules became small in size and exocytotic features were more frequent. Mitochondria and lysosomes were also increased in number. Thus, the synthetic and secretory activities of the PT-specific cells were increased under extended darkness. The data indicate that the specific cells of the pars tuberalis are responsive to photoperiodic changes in the chick.     

Circadian Timekeeping for the Photoperiodic Control of Budset in Picea abies (Norway Spruce) Seedlings

Romanian populations of Norway spruce are induced to set terminal buds by four inductive cycles of 8 h light/16 h darkness. To distinguish between circadian and hourglass timekeeping for the photoperiodic control of budset, seedlings were raised in continuous light at 300 µmol m^-2s^-1 at 20°C for 10 weeks. They were then exposed to an extended night regime consisting of three cycles of 8 h light/40 h dark with 4-h or 1-h nightbreaks (120 µmol m^-2s^-1) applied to groups of plants at intervals during the extended night. Following a final cycle of 8 h light/16 h dark to maximize budset, the plants were transferred to continuous light. Budset was delayed when the night-break was applied close to the critical nightlength (CNL) of 6–7 h or about 22–23 h later in the extended night, consistent with circadian rather than hourglass timekeeping. Confidence intervals were calculated for the times to maximum effect of the night-breaks.     

Pigment formation in potato tubers (Solanum tuberosum) exposed to light followed by darkness

Potato tubers ( Solanum tubersoum cvs Bintje and King Edward). never exposed to light, lack chlorophyllous pigments. Continuous irradiation results in chlorophyll (Chl) formation and induces the ability for protochlorophyll (Pchl) formation when the tubers are brought back to darkness. Pigment synthesis takes place in both blue and red light, but blue light is more effective than red in starting the greening process. The pigment formation is strongest in the layers just below the periderm with a steep gradient inwards. Small amounts of Chl formed after irradiation. slowly fade away during extended darkness. However, the Chl formed after long time of irradiation is remarkably stable. Irradiated potatoes, placed in darkness, form Pchl with a fluorescence emission peak at 633 nm. A maximal level is reached after ca 7 days. Resolution of the Pchl spectrum suggests the presence of small amounts of a pigment with an emission maximum at around 642 nm. No sign of the Pchl with emission maximum at 657 nm, which dominates in etiolated leaves, is found. A faint Chl fluorescence indicates that some Pchl, probably the 642 nm form, is phototransformed into Chl in weak light. The Chl formation in the potato tuber is discussed in relation to that of roots and leaves.     

Content of Adenosine Phosphate Compounds in a Long-Day Duckweed, Lemna gibba G3, under Different Light and Nutritional Conditions

The levels of ATP ADP and AMP as well as the energy charge were examined in the long-day duckweed. Lemna gibba G3, under different light and nutritional conditions. ATP and ADP content, but not AMP, decreased slightly when the plant was cultured with a medium depleted of sucrose or was exposed to continuous darkness. The energy charge was not affected either by changes in the light conditions or by the depletion of sucrose from the medium, although increase of fresh weight changed drastically under these conditions. The levels of ADP and AMP and the value of the energy charge remained nearly constant throughout a 1-day period when the plant was exposed to short-day, continuous light and continuous dark. ATP content decreased gradually during the light period of the short day and thereafter remained constant during the dark period. When the plant was exposed to continuous light. ATP content decreased until the 8th hour after beginning of the continuous light, only to recover rapidly to its original level during the phase coinciding with the preceding dark period of the short day. Under continuous dark conditions the ATP content remained constant throughout the day. It was concluded that the diurnal rhythm of physiological activities previously reported cannot be related to the energy charge.     

CYCLING OF STOMATAL APERTURE IN LEAVES OF PLANTS WITH CRASSULACEAN ACID METABOLISM UNDER CONSTANT CONDITIONS

When leaves of plants with C₃ metabolism are detached and held in darkness, they senesce and the stomata close. Because the relation of senescence and stomatal closure is very close, if not actually causal, the question arose as to whether in the leaves of plants with Crassulacean acid metabolism whose stomata open at night the relationship to senescence would be reversed. Detached leaves of four species of Hoya, floated on water in constant darkness or constant light, were found to show no large differences in stomatal aperture (measured as diffusion resistance) between those in the light or dark, but the aperture changed in a regular circadian rhythm. In some leaves the rhythm was simple, in others the peak showed small secondary peaks, but in all cases the values were nearly the same in the light as in the dark, throughout the cycle. Previous culture of the intact plants under normal day/night conditions gave results similar to those with plants that had had prolonged culture under constant light or darkness. In those cases when the stomata were more open in the dark, the chlorophyll content was greater than when the stomata were more open in the light; but when they were more open in the light, the chlorophyll content showed little difference between light and dark. When the leaves had only their petioles in water they showed greater senescence in the light than in the dark, and the stomata were more tightly closed in the light, especially at the apical ends. All four species of Hoya gave similar results. We deduce that senescence of these leaves is modified by stomatal aperture, and generally in the same direction as in C₃ leaves, but that in continuous light or darkness the primary control over the aperture is the endogenous cycle.     

The effect of light and darkness on hatching in the pomacentrid Abudefduf saxatilis

Synopsis Reports that pomacentrid embryos hatch after dusk are confirmed by photic manipulation of sergeant major eggs. Embryos placed in the dark for 20 minutes or longer prior to their normal hatching after sunset hatched, whereas controls held in light did not hatch. Percent of hatched embryos correlated with increasing exposure to darkness up to one hour after which no further improvement in hatching was observed. Embryos maintained in continuous light during their normal twilight hatching period did not hatch. Also, embryos exposed to 60 minutes of darkness, if interrupted by one minute of light every 10 minutes did not hatch. The percent hatch in dark treatments varied significantly between nests and, in some treatments, correlated negatively with the size of the egg clumps (number of eggs per clump) tested. To initiate hatching in the presence of light required intensities of 0.03 lux or less. These low intensities are not reached until about 20 minutes after sunset on the reef where the embryos occur. We conclude that hatching for some embryos occurs about 30 minutes after sunset but for most is not completed until at least one hour after sunset. Hatching therefore takes place at a time long after potential diurnal fish predators have refuged in the reef structure.     

Circadian and senescence-enhanced expression of a tobacco cysteine protease gene

A cDNA clone encoding a cysteine protease was isolated from a tobacco cDNA library, utilizing as a probe a PCR fragment obtained from degenerated primers based on the conserved sequences of plant cysteine protease genes. A putative protein encoded by the clone NTCP-23 had an amino acid sequence with significant similarities to those of plant senescence-associated cysteine proteases and mammalian cathepsin H. Northern blot analysis showed that NTCP-23 mRNA is expressed in all organs and the mRNA and protein expression is enhanced during natural senescence. We propose that NTCP-23 is responsible for amino acid remobilization especially in senescencing leaves. Furthermore, it was found that the mRNA expression follows a circadian rhythm and is reduced by continuous darkness, wounding and hypersensitive reaction (HR). NTCP-23 is the first cysteine protease whose mRNA expression has been shown to be temporarily reduced by wounding.     

Effects of light and spermine on senescence of hydrilla and spinach leaves

Light treatment markedly accelerated chlorophyll loss in Hydrilla (Hydrilla verticillata [L.f.] Royle) over dark treatment whereas such acceleration could not be observed in spinach (Spinacia oleracea L.) leaf segments. Spermine, a polyamine, retarded the loss of chlorophyll in the dark but markedly accelerated this loss in the light during senescence of Hydrilla leaves. However, such effect of spermine in the dark was not so pronounced in spinach. The loss of protein was slower in the light than in the dark in both the species. Spermine arrested the loss of protein (as in spinach) or even raised the protein level over initial (as in Hydrilla). Loss of both soluble and insoluble protein was slower in light than in darkness. Spermine treatment, either in light or darkness, markedly accelerated the loss of soluble protein but raised the level of insoluble protein over initial in both the species. The pattern of change in α-amino nitrogen in either species could be correlated well with that of protein level. In Hydrilla, light increased the soluble protein fraction over initial and this rise was prevented by cycloheximide and not by chloramphenicol. Also, spermine augmented the protease activity (both acid and neutral) while light retarded the rise in protease activity during senescence of either species. Although spermine treatment reduced the leaching of α-amino nitrogen and electrolytes in Hydrilla, it augmented the same in spinach.     

Vacuolar cysteine proteases of wheat (Triticum aestivum L.) are common to leaf senescence induced by different factors

Cellular proteins are extensively degraded during leaf senescence, and this correlates with an up-regulation of protease gene expression, particularly cysteine proteases. The objectives of this work were (i) to detect cysteine proteases associated with senescence of wheat leaves under different conditions and (ii) to find out their subcellular location. Activity labelling of cysteine proteases with the biotinylated inhibitor DCG-04 detected five bands at 27, 36, 39, 42, and 46 kDa in leaves of wheat senescing under continuous darkness. In-gel activity assays showed that these proteases are only active in an acid milieu (pH 4), and their activity increased several-fold in senescing leaves. Fractionation experiments showed that the senescence-associated cysteine proteases of 36, 39, 42, and 46 kDa localize to a vacuolar-enriched fraction. The vacuolar cysteine proteases of 36, 39, and 42 kDa increased in activity in attached flag leaves senescing naturally during post-anthesis, and in attached leaves of plants subjected to a period of water deficit. Thus, the activity of these vacuolar cysteine proteases is associated with developmental (post-anthesis) senescence and with senescence induced by stress factors (i.e. protracted darkness or drought). This suggests that vacuoles are involved in senescence-associated cellular degradation, and that different senescence-inducing factors may converge on a single degradation pathway.     

Characterization of senescence-associated proteases in postharvest broccoli florets.

We characterized the senescence-associated proteases of postharvest broccoli (Brassica oleracea L. var Green King) florets, using class-specific protease inhibitors and gelatin-polyacrylamide gel electrophoresis. Different classes of senescence-associated proteases in broccoli florets were partially characterized for the first time. Protease activity of broccoli florets was depressed by all the inhibitors and showed different inhibition curves during postharvest. The hydrolytic activity of metalloprotease (EC 3.4.24. - ) and serine protease (EC 3.4.21. - ) reached a maximum, 1 day after harvest (DAH), then decreased, while the hydrolytic activity of cysteine protease (EC 3.4.22. - ) and aspartic protease (EC 3.4.23. - ) increased throughout the postharvest senescence based on the calculated inhibition percentage of protease activity. The senescence-associated proteases were separated into seven endoprotease (EP) groups by gelatin-polyacryamide gel electrophoresis and classified into EP1 (metalloprotease), EP2 (metalloprotease and cysteine protease), EP3 (serine protease and aspartic protease), EP4, EP5, EP7 (cysteine protease), and EP6 (serine protease) based on the sensitivity of class-specific protease inhibitors. The proteases EP2, EP3, and EP4 were present throughout the postharvest stages. EP3 was the major EP at all times during senescence; EP4 intensity of activity increased after 2 DAH; EP6 and EP7 clearly increased after 4 DAH. Our results suggest that serine protease activity contributes to early stage (0-1 DAH) and late stage (4-5 DAH) of senescence; metalloprotease activity was involved in the early and intermediate stages (0-3 DAH) of senescence; and cysteine protease and aspartic protease activities participated in the whole process of broccoli senescence.     

Induction of a Carbon-Starvation-Related Proteolysis in Whole Maize Plants Submitted to Light/Dark Cycles and to Extended Darkness

Three-week-old maize (Zea mays L.) plants were submitted to light/dark cycles and to prolonged darkness to investigate the occurrence of sugar-limitation effects in different parts of the whole plant. Soluble sugars fluctuated with light/dark cycles and dropped sharply during extended darkness. Significant decreases in protein level were observed after prolonged darkness in mature roots, root tips, and young leaves. Glutamine and asparagine (Asn) changed in opposite ways, with Asn increasing in the dark. After prolonged darkness the increase in Asn accounted for most of the nitrogen released by protein breakdown. Using polyclonal antibodies against a vacuolar root protease previously described (F. James, R. Brouquisse, C. Suire, A. Pradet, P. Raymond [1996] Biochem J 320: 283–292) or the 20S proteasome, we showed that the increase in proteolytic activities was related to an enrichment of roots in the vacuolar protease, with no change in the amount of 20S proteasome in either roots or leaves. Our results show that no significant net proteolysis is induced in any part of the plant during normal light/dark cycles, although changes in metabolism and growth appear soon after the beginning of the dark period, and starvation-related proteolysis probably appears in prolonged darkness earlier in sink than in mature tissues.     

Relation between Respiration and Senescence in Oat Leaves

The respiration of excised oat (Avena sativa cv Victory) leaves and their sensitivity to inhibitors was followed during senescence under varied conditions. The respiration rate, which in controls reaches its peak on the third day in darkness, is lowered at the time of fastest loss of chlorophyll (as reported earlier) by seven unrelated reagents that all delay dark senescence. When senescence is delayed by white light or by cytokinins, the respiratory rise is correspondingly delayed. Kinetin and l-serine, which act as antagonists on senescence, also act as antagonists on the respiratory rate. However, an exception to this close correspondence between senescence and the respiratory rise is offered by the lower aliphatic alcohols, which delay dark senescence and yet accelerate the onset of the respiratory rise.     

Metabolism of Oat Leaves during Senescence : VII. The Interaction of Carbon Dioxide and Other Atmospheric Gases with Light in Controlling Chlorophyll Loss and Senescence

In air largely freed from CO₂, senescence of isolated oat (Avena sativa cv Victory) seedling leaves is no longer prevented by white light; instead, the leaves lose both chlorophyll and protein as rapidly as in the dark. Senescence in light is also accelerated in pure O₂, but it is greatly delayed in N₂; 100% N₂ preserves both protein and chlorophyll in light and in darkness. In light in air, most of the compounds tested that had previously been found to delay or inhibit senescence in darkness actually promote the loss of chlorophyll, but they do not promote proteolysis. Under these conditions, proteolysis can therefore be separated from chlorophyll loss. But in light minus CO₂, where chlorophyll loss is rapid in controls, two of these same reagents prevent the chlorophyll loss. Unlike the many reagents whose action in light is thus the opposite of that in darkness, abscisic acid, which promotes chlorophyll loss in the dark, also promotes it in light with or without CO₂. Kinetin, which prevents chlorophyll loss in the dark, also prevents it in light minus CO₂. In general, therefore, the responses to light minus CO₂ are similar to the responses to darkness, and (with the exception of abscisic acid and kinetin) opposite to the response to light in air.     

Analysis of secreted protease inhibitors after water stress in potato tubers

Potato tubers (Solanum tuberosum) secrete two kinds of proteinase inhibitors after a water stress. The polypeptides have differing inhibitory activities but are Kunitz-type inhibitors based on amino-terminal sequences homologies. A proteolysis maturation type of a cell protease inhibitor was observed. They can constitute high MW complex, sometimes with another type of protein. The function of these protease inhibitors is discussed in relation to plant defence.     

Possible mechanisms of light-induced chlorophyll degradation in senescing leaves of Hydrilla verticillata

Light treatment markedly accelerated the chlorophyll loss in senescing leaves of Hydrilla verticillata [(L.f.) Royle] as compared to dark treatment, whereas such acceleration could not be observed in senescing spinach (Spinacia oleracea L.) leaves. The light-induced cholorophyll loss in Hydrilla was retarded slightly by chloramphenicol and markedly by cycloheximide. Catalase (EC 1.11.1.6) activity did not change appreciably in Hydrilla leaves either in light or in darkness, while in spinach it declined markedly in the dark, and light retarded such decline. Peroxidase activity in Hydrilla showed faster increase in light than in darkness, while in spinach it increased only in light during senescence. The activity of phenol(pyrogallol)-specific peroxidase increased markedly in light, and that of ascorbate-specific peroxidase decreased slightly both in light and darkness during senescence of Hydrilla leaves. This rise in phenolspecific peroxidase activity was prevented by cycloheximide treatment. Pretreatment of Hydrilla leaves with monophenol (2,4-dichlorophenol) and o-diphenol (hydroquinone) accelerated and retarded, respectively, the light-induced cholorophyll loss. Pretreatment of Hydrilla leaves with H₂O₂ augmented the chlorophyll loss more markedly in light than in darkness. The endogenous level of H₂O₂ increased more in light than in dark during senescence of Hydrilla leaves. Treatment of Hydrilla leaves with 3-(3.4-dichlorophenyl)-l,l-dimethylurea. a photosystem II inhibitor, prevented both light-induced rise in H₂O: level and chlorophyll loss, but it was without effect in the dark. Retardation of light-induced chlorophyll loss occurred during senescence of Hydrilla leaves when light was given in different photoperiods in a 24-h daily cycle for 6 days instead of as continuous irradiance. There was a negative correlation between the length of the photoperiod and the extent of cholorophyll loss.     

Relationship between light conditions and behavior of the freshwater snail Biomphalaria glabrata (Say)

The influence of light upon behavior of Biomphalaria glabrata was investigated in snails submitted for 48 h to one of the following regimes: normal light cycle, continuous darkness, continuous light. Time-lapse cinematography was used to provide data about snail locomotor activity in response to (a) continuous light or darkness; (b) light or dark phases; (c) light transitions. Snails were significantly less active under continuous light than under continuous or intermittent darkness. Under the normal light cycle, the activity rate was significantly higher in the dark than in the light. Changes from light to dark corresponded to increases in the activity rate which persisted long afterwards. No significant variation in activity occurred upon changes from dark to light.