Nucleic acid synthesis and effect of glucose on its kinetics in cotyledons ofChenopodium rubrum l. during photoperiodic induction

In cotyledons ofChenopodium rubrum L. polydisperse RNA is synthesized in the region of the low molecular weight RNAs during photoperiodic induction. After short-time labelling the rate of 4s RNA synthesis was always higher in induced plants than in plants having obtained a light-break in the middle of the dark period. When glucose was added to the nutrient medium during the dark period of a single photoperiodic cycle the rate of nucleic acid (NA) synthesis was higher in non-induced plants than in induced ones at the termination of the dark period. In plants induced by two cycles in the absence of glucose the rate of NA synthesis at the termination of the second dark period was higher in induced than in non-induced plants. This difference is due to the differential kinetics of NA synthesis during darkness. In plants induced in the presence of glucose the peak of the rhythm in NA synthesis was advanced by 4 h relative to that found in plants induced in the absence of sugar. Thus, the termination of the dark period coincided with the negative slope of the oscillation in plants induced in the presence of glucose, while in plants having obtained a light-break NA synthesis decreased only slightly after having attained its peak. In plants induced in the absence of glucose the termination of the dark period coincided with the peak in the rhythm in NA synthesis. The rhythm in NA synthesis of the cotyledons during the dark period of an inductive cycle is out of phase with the rhythm in flower initiation.     

Rhythmic changes in ribonuclease activity in relation to nucleic acid synthesis in cotyledons ofChenopodium rubrum L. and to floral induction of the plants

Ribonuclease (RNAse) activity was investigated in cotyledons ofChenopodium rubrum plants subjected to various conditions of illumination (photoperiodic induction, continuous light, induction cancelled by interrupting the dark period by a light-break). At the end of the dark period of the single inductive cycles RNAse activity of induced plants was inferior to that of plants grown in continuous light. At the end of the first two cycles the activity was lowest after the interruption of the dark period by light. The investigation of the enzyme in 6h intervals showed rhythmic changes in activity to occur in induced plants. Enzyme activity followed a pattern opposed to this of nucleic acid (NA) synthesis in the cotyledons. In plants from continuous light the enzyme activity did not show any rhythm and in plants having obtained a light-break during the inductive period the rhythm was less distinct than in the induced ones. The period length of the endogenous rhythm of NA synthesis in the cotyledons is about half as long as this of flowering and the peaks of flowering coincide with the throughs of NA synthesis.     

Changes in nucleic acid synthesis in cotyledons and apical buds of chenopodium rubrum l. associated with photoperiodic induction

The nucleic acid (NA) fractions were analyzed in cotyledons and apical buds ofChenopodium rubrum plants by means of acrylamide electrophoresis at the end of the dark period of a different number of photoperiodic cycles or after transfer of the plants to light for 4 h subsequent to the termination of the dark period. The plants were labelled with³²P three hours prior to sampling. The uptake of³²P into the cotyledons was higher in light than in darkness in all cases, however, it was not in correlation with³²P incorporation into the NA fractions. After one dark period lasting 8 or 16 h NA synthesis in light did not increase in comparison with darkness. After two or more photoperiodic cycles NA synthesis was higher in light than in darkness irrespective of whether the dark period lasted 8 or 16 h. NA synthesis was distinctly highest after two inductive cycles lasting 16 h. In buds NA synthesis was slightly shifted in favour of ribosomal RNA as compared with cotyledons. In the cotyledons the increase in light was mainly duo to a raise of rRNA synthesis whereas in the buds synthesis of sRNA and DNA increased, as well.     

Fluctuations of Uridine Incorporation in the Shoot Apex ofChenopodium rubrum L. during Photoperiodic Induction

Uridine incorporation into the shoot apex of the short-day plantChenopodium rubrum was investigated during a 16 h period of darkness and the following transfer to light. Uridine incorporation during this single inductive cycle was compared to incorporation under non-inductive conditions of continuous light. After transfer of the plants from light to darkness RNA synthesis was reduced to about half after the first two hours. This occurred not only when the plants were precultivated in continuous light but also after an interruption of the dark period by light for 31/2 h. The low level of uridine incorporation was maintained for the whole duration of the dark period. Incorporation regained its initial level after exposure of the plants to light irrespective of the duration of the preceding dark period. After this immediate rise of uridine incorporation in plants transferred from darkness to light a slight temporary decrease was observed in light. In darkness the decrease of incorporation into the nucleoli was still more marked than the reduction of overall incorporation. After the termination of the dark period incorporation into the nucleolus rose slowly and extranucleolar incorporation was relatively enhanced during the first 10 h of light in induced plants. The fluctuations of RNA synthesis observed in the shoot apex during photoperiodic treatment may be regarded as a necessary condition for the transition from the vegetative to the reproductive state.     

Changes in the content of endogenous auxins in apical buds ofchenopodium rubrum L. Induced with respect to the endogenous rhythm in capacity to flower

The content of endogenous auxins was examined in apical buds ofChenopodium rubrum plants induced by a photoperiodic cycle of 16h darkness and 8h light followed by a dark period of various duration so as to correspond with either maximal or minimal flowering response in the endogenous rhythm in capacity to flower initiated by the photoperiodic treatment. Apical buds of potentially generative plants contained less auxins than apical buds of plants which remained in the vegetative state. Apical buds from plants treated with kinetin (1. 10^-3 M) and therefore remaining in the vegetative state showed an auxin level comparable to that of untreated plants exhibiting minimal flowering response irrespective of the duration of the second dark period. Plants cultivated on a sucrose solution (0.6 M) during the second dark period became generative even at the normal minimum of flowering. The auxin content of the apical buds was low, similarly as in untreated plants induced for a period leading to maximal flowering response. On the other hand, apical buds from plants grown on sucrose solution during a dark period leading to the manifestation of maximal flowering response showed a relatively high auxin content comparable to that found in untreated plants which had obtained a more extended induction by three photoperiodic cycles. The results are discussed with respect to the possible role of endogenous auxins in the regulation of the changes in growth correlations occurring in the shoot apex during photoperiodic induction and in the expression of the competence to flower.     

Rhythms as photoperiodic timers in the control of flowring in Chenopodium rubrum L.

Summary In C. rubrum, the amount of flowering that is induced by a single dark period interrupting continuous light depends upon the duration of darkness. A rhythmic oscillation in sensitivity to the time that light terminates darkness regulates the level of flowering. The period length of this oscillation is close to 30 hours, peaks of the rhythm occurring at about 13, 43 and 73 h of darkness.Phasing of the rhythm by 6-, 12- and 18-h photoperiods was studied by exposing plants to a given photoperiod at different phases of the free-running oscillation in darkness. The shift in phase of the rhythm was then determined by varying the length of the dark period following the photoperiod; this dark period was terminated by continuous light.With a 6-h photoperiod the timing of both the light-on and light-off signals is shown to control rhythm phasing. However, when the photoperiod is increased to 12 or 18 h, only the light-off signal determines phasing of the rhythm. In prolonged periods of irradiation-12 to 62 h light—a durational response to light overrides any interaction between the timing of the light period and the position of the oscillation at which light is administered. Such prolonged periods of irradiation apparently suspend or otherwise interact with the rhythm so that, in a following dark period, it is reinitiated at a fixed phase relative to the time of the light-off signal to give a peak of the rhythm 13 h after the dusk signal.In daily photoperiodic cycles rhythm phasing by a 6-h photocycle was also estimated by progressively increasing the number of cycles given prior to a single dark period of varied duration.In confirmation of Bünning's (1936) hypothesis, calculated and observed phasing of the rhythm controlling flowering in c. rubrum accounts for the photoperiodic response of this species. Evidence is also discussed which indicates that the timing of disappearance of phytochrome P_fr may limit flowering over the early hours of darkness.     

Gibberellins in the control of photoperiodic flower transition in Pharbitis nil

The role of gibberellins in the photoperiodic flower induction of short-day plant Pharbitis nil has been investigated. It has been found that the endogenous content of gibberellins in the cotyledons of P. nil is low before and after a 16-h-long inductive dark period. During the inductive night the content of gibberellins is high at the beginning of darkness and about the middle of the dark period. Exogenous GA₃ when applied to the cotyledons of non-induced plants does not replace the effect of the inductive night but it can stimulate the intensity of flowering in plants cultivated on suboptimal photoperiods. GA₃ could also reverse the inhibitory effect of end-of-day far-red light irradiation on P. nil flowering. 2-Chloroethyltri-methylammonium chloride (CCC) applied to the cotyledons during the inductive night also inhibited flowering. GA₃ could reverse the inhibitory effect of CCC. The obtained results strongly suggest that gibberellins are involved in the phytochrome controlled transition of P. nil to flowering. Their effect could be additive to that of photoperiodic induction.     

Fluctuation of free IAA under inductive and non-inductive photoperiods in Chenopodium rubrum

Fluctuation in levels of endogenous free IAA has been followed in the SD plant Chenopodium rubrum under photoperiodic conditions inductive or not inductive of flowering. Endogenous IAA was measured fluorimetrically as -pyrone. The level of IAA shows little fluctuation under continuous illumination. An endogenous rhythm of IAA fluctuation was found in plants transferred from light to continuous darkness, with a natural period of 30 hrs. The troughs of minimum IAA level within the endogenous rhythm coincided with the peaks in the endogenous rhythm of flowering response, which possessed the same period length. The concentration of IAA in the shoot always decreased at the end of cycles of dark period that induce flowering. The results are discussed in relation to the role of IAA in flowering of SD plants.     

Photoperiodic time measurement in the male lizardAnolis carolinensis

Summary Photoperiod plays an important role in controlling the annual reproductive cycle of the male lizardAnolis carolinensis. Groups of anoles were exposed to various experimental lighting regimens to determine how the lizards were measuring the length of the day. The experimental regimens were designed to discriminate between the following two general classes of hypotheses: (1) Photoperiodic time measurement is based on an hourglass or interval timer which measures the length of the light or dark or (2) Photoperiodic time measurement is based on an endogenous circadian rhythm of photoperiodic photosensitivity. The experiments demonstrated thatAnolis uses an hourglass mechanism which measures the absolute length of the light period. This is in contrast to the higher vertebrates (birds and mammals) which measure photoperiodic time by means of a circadian oscillation of responsiveness to light.     

Changes of the diurnal and circadian (endogenous) mRNA oscillations of the chlorophyll a/b binding protein in tomato leaves during altered day/night (light/dark) regimes

Characteristic steady-state mRNA level oscillations were monitored for the chlorophyll a/b-binding (cab) protein in tomato plants grown under the natural day/night (light/dark) regime as well as under constant environmental conditions. This typical expression pattern was altered when plants were transferred to different light/dark regimes. For example, by shifting the light phase by six hours, a change of the time points of maximum and minimum of expression level was monitored, while the principal oscillation pattern remained the same. It appeared that the transition from dark to light is involved in determining the time points of minima and maxima of mRNA accumulation.After exposing tomato plants to an abnormal light/dark periodicity (e.g. six hours of alternating light/dark) an altered oscillation pattern was determined: within 24 hours two maxima of cab mRNA levels were detected. However, this entrained abnormal rhythm was not manifested at the molecular level and the circadian pattern reappeared under constant environmental conditions (e.g. darkness). This result favours the hypothesis that the oscillation pattern of the cab mRNA in tomato plants is not only endogenous but also hereditary.     

Light Rquirements for Photoperiodic Sensitivity in Cotyledons of Dark-grown Pharbitis nil

The light requirements for induction of flowering by a long dark period were investigated in dark-grown seedlings of Pharbitis nil Chois, cv. Violet. The cotyledons bcame photoperiodically sensitive to a 24 h dark period by two 1 min red irradiations (6.3 μmol m⁻² S⁻¹) separated by a 24 h dark period. The reversibility of the effect of brief red irradiations, and the effectiveness of low energies of red irradiation suggest the involvement of phytochrome in the induction of photoperiodic sensitivity. Partial de-etiolation occurred after these brief periods of red irradiation but the seedlings were not capable of net CO₂ uptakeeven 7 h after the start of the main light period that followed the critical dark period. A changing response to the duration of the priod of darkness given between the two short red irradiations showed the the correct phasing of an endogenous photoperiodic rhythm is needed for the attainment of photoperiodic snsitivity.     

Capacity of cytochrome and alternative path in coupled and uncoupled root respiration of Pisum and Plantago

A critical duration of darkness must be exceeded for the photoperiodic induction of flowering in short-day plants. This requires detection of the light/dark transition at dusk and the coupling of this information to a time-measuring system.
Lowering the P_fr/P_tot, ratio photochemically at the end of the day did not accelerate the onset of dark timing in Pharbitis nil Choisy cv. Violet. Time-measurement was initiated when, with no change in spectral quality, the irradiance fell below a threshold value. Thus, if the light/dark transition at dusk is sensed by a reduction in P_fr, this reduction can be achieved as rapidly through thermal reactions as through photochemical ones. When given at hourly intervals during a 6-h extension of a 24-h main light period in white light, pulses of red light were as effective as continuous red light in delaying the onset of timing; pulses every 2 or 3 h were less effective. The effectiveness of intermittent red light indicates that phytochrome is the photoreceptor and the requirement for frequent exposures suggests that P_fr is lost rapidly in the dark. However, the red light pulses could not be reversed by far-red light, which argues against this hypothesis. An alternative explanation is that the perception of light as being continuous occurs only when "new" P_fr is regenerated sufficiently frequently.
The nature of the coupling of the dusk signal to the time-measuring system is discussed and it is suggested that the effect of each red light pulse is to delay the phase of the photoperiodic rhythm by 1–3 h.     

Jasmonates Inhibit Flowering in Short-Day Plant Pharbitis nil

The role of jasmonates in the photoperiodic flower induction of short-day plant Pharbitis nil was investigated. The plants were grown in a special cycle: 72 h of darkness, 24 h of white light with lowered intensity, 24-h long inductive night, 14 days of continuous light. At 4 h of inductive night the cotyledons of non-induced plants contained about two times the amount of endogenous jasmonates (JA/JA-Me) compared to those induced. A 15-min long pulse of far red light (FR) applied at the end of a 24-h long white light phase inhibited flowering of P. nil. The concentration of jasmonates at 2 and 4 h of inductive night in the cotyledons of the plants treated with FR was similar. Red light (R) could reverse the effect of FR. R light applied after FR light decreased the content of jasmonates by about 50%. Methyl jasmonate (JA-Me) applied to cotyledons, shoot apices and cotyledon petioles of P. nil inhibited the formation of flower buds during the first half of a 24-h long inductive or 14-h long subinductive night. Application of JA-Me to the cotyledons was the most effective. None of the plants treated with JA-Me on the cotyledons in the middle of the inductive night formed terminal flower buds. The aspirin, ibuprofen and phenidone, jasmonates biosynthesis inhibitors partially reversed the effect of FR, stimulating the formation of axillary and terminal flower buds. Thus, the results obtained suggests that phytochrome system control both the photoperiodic flower induction and jasmonates metabolism. Jasmonates inhibit flowering in P. nil.     

Photoperiodic control of tetrasporangium formation in the red alga Rhodochorton purpureum

Six geographical isolates of Rhodochorton purpureum (Lightfoot) Rosenvinge (Rhodophyta, Nemalionales) formed tetrasporangia only in short days at 10°C. For most isolates, the critical day-length increased with latitude of origin from 9.5 h for an isolate from California to 14.5 h for one from Antarctica. Tetrasporangium production could be induced by 9–15 short-day cycles followed by a further 22–28 cycles in long days. A night-break consisting of 1 h of white light in the middle of a 16-h dark period inhibited the short-day response of isolates from low latitudes, but not those from higher latitudes. When a similar night-break was given in the middle of a 14-h dark period, however, the response of all isolates was at least partially inhibited. Night-breaks given at any time in the central 7 h of a 14-h dark period were equally inhibitory. Broad-band red light (0.3–0.4 mmol m^-2), given as a night-break, caused 50% inhibition of the short-day response. At a slightly higher photon exposure (0.6 mmol m^-2, given as 1 μmol m^-2 s^-1 for 10 min), narrow-band red (662 nm) and blue (448 nm) light caused similar inhibition, but green (547 nm) and far-red (731 nm) were ineffective as night-breaks. The inhibitory effect of a 10-min night-break with red light could not be reversed by subsequent exposure to an equal photon exposure of far-red light. These results add to the existing evidence that the pigments mediating photoperiodic responses among algae are more varied than those among flowering plants.     

The photoperiodic clock in the flesh-fly, Sarcophaga argyrostoma

Larval cultures of the flesh-fly, Sarcophaga argyrostoma, were raised in experimental light cycles with periods (T) of 21 to 72 hr, each cycle containing a photoperiod of 4 to 20 hr of white light. This ‘resonance’ technique revealed periodic maxima (～24 hr apart) of pupal diapause, thereby demonstrating an endogenous circadian component in the photoperiodic clock. The positions of these maxima of pupal diapause suggested that the oscillation, like that controlling the pupal eclosion rhythm in Drosophila pseudoobscura, is ‘damped out’ by photoperiods longer than about 11 to 12 hr, but restarts at dusk whereupon it runs with circadian periodicity in a protracted dark period. With photoperiods shorter than 12 hr, however, the two diapause maxima were less than 24 hr apart, suggesting that an additional component, possibly a ‘dawn hour-glass’, was modifying the position of the first peak.Both photoperiod and the period of the driving light cycle (T) were shown to affect the length of larval development (the sensitive period) and the number of calendar days needed to raise the incidence of pupal diapause to 50 per cent (the required day number, RDN). Peaks of diapause induction were shown to be the result of an interaction between a long sensitive period (slow development) and a low RDN, whereas troughs in diapause induction were the result of an interaction between a short sensitive period (fast development) and a higher RDN.Larvae of S. argyrostoma are unable to distinguish (in a photoperiodic sense) between 12 and 18 hr of red light (600 nm).     

Changes in cotyledon mRNA during floral induction of Pharbitis nil CV. Violet

Floral induction in seedlings of Pharbitis nil Choisy cv. Violet, with one cotyledon removed, was manipulated by applying various photoperiodic treatments to the remaining cotyledon. Populations of polyadenylated RNA from treated cotyledons were examined to identify messages specifically involved in floral induction. The RNA was translated in vitro using a wheat-germ system, and the resulting translation products were analysed by two-dimensional polyacrylamide gel electrophoresis. Substantial qualitative and quantitative differences were found between mRNA from cotyledons of seedlings kept in continuous light (non-induced) and of seedlings given a 16-h dark period (induced). In contrast, inhibition of flowering with a night-break resulted only in one detectable, quantitative difference in mRNA.Abbreviations CL continuous light - kDa kilodalton - NB 16 h darkness+10 min red-light break, 8 h into the dark period - poly(A)⁺ RNA polyadenylated RNA (isolated by binding to a cellulose oligodeoxythymidine affinity column) - SD short day (16 h dark) - SDP short-day plant - SDS sodium dodecyl sulfate     

Growth correlations and rna synthesis in different parts of the shoot apical meristem ofChenopodium rubrunt L. induced to flowering

Uridine-³H incorporation and RNA concentration were investigated in different parts of the shoot apical meristem ofChenopodium rubrum using autoradiography and cytophotometry. A single inductive cycle was sufficient to bring about postinductive first events in the shoot apex but not for complete flower differentiation. The initial activation of RNA synthesis manifested itself in all zones of the apex. The first increase was more conspicuous in the peripheral than in the central zone. The indications of the first events in the apices after a single inductive cycle disappear prior to morphological reversal to the vegetative state. Induction by three short days led to rapid flower differentiation. The increase in RNA synthesis and concentration was most conspicuous in the central zone in this case. The ratio of RNA synthesis and content between bud and leaf primordia (B/L) also change in relation to photoperiodic induction. In vegetative plants the B/L ratio was low while after induction it increased. The shifts in activity of RNA synthesis observed in the shoot apical meristem are related to the changes in growth activity of the different parts of the apex. The growth ratios in the apices bear the character of growth correlations. The change in the growth correlations following photoperiodic induction together with the total activation of RNA synthesis are considered to represent one of the first events of the transition to the reproductive state.     

Photoperiodic control of reproduction in the male lizardAnolis carolinensis

Summary In contrast to the higher vertebrates the photoperiodic time measuring system in the male lizardAnolis carolinensis seems to rely on an hourglass timer which lacks endogenous rhythmicity. This timer appears to measure the absolute length of the light portion of light-dark (LD) cycles. The present study further characterized the nature of theAnolis photoperiodic timer and demonstrated: (1) The gonadal response is quite sensitive to photostimulation. Exposure to as few as three 16 h photoperiods (over a 3 week period) can maintain testicular function in summer anoles whereas exposure to as few as six 16 h photoperiods (over a 3 week period) can elicit maximal testicular development in the fall. (2) The photoperiodic timer does not have to be reset daily by a dark interruption. (3) The dark portion of LD cycles may be involved in a complex fashion in reversing a light-initiated reaction and (4) Comparisons of entrained circadian activity rhythms with testicular responses to various light cycles argue against the participation of a circadian clock in photoperiodic time measurement.Abbreviation CRPP circadian rhythm of photoperiodic photo-sensitivity